S. cerevisiae

Mutat Res, 1982 Dec, 105(6), 403 - 7
Testing of Endosulfan and Fenitrothion for genotoxicity in Saccharomyces cerevisiae; Yadav AS et al.; Two insecticides, Endosulfan and Fenitrothion, were tested for their ability to induce mitotic crossing-over, mitotic gene conversion and reverse mutation in Saccharomyces cerevisiae . Treatment of cells with Endosulfan increased the frequencies of gene convertants and revertants . However, Fenitrothion treatment did not induce any of these genetic events.

Fed Proc, 1982 Dec, 41(14), 3084 - 8
The biochemistry, genetics, and regulation of polyamine biosynthesis in Saccharomyces cerevisiae; Tabor CW et al.; We have studied the enzymes and genes involved in the biosynthesis of putrescine, spermidine, and spermine in Saccharomyces cerevisiae . Mutants have been isolated with defects in the biosynthetic pathway as follows: spe10 mutants, deficient in ornithine decarboxylase, cannot make putrescine, spermidine, or spermine; spe2 mutants, lacking S-adenosylmethionine decarboxylase, cannot make spermidine or spermine; spe3 mutants, lacking putrescine aminopropyltransferase, cannot make spermidine or spermine; and spe4 and spe40 mutants, lacking spermidine aminopropyltransferase, contain no spermine and permit growth of spe10 mutants . Studies with these mutants have shown that in yeast: 1) polyamines are absolutely required for growth; 2) putrescine is formed only by decarboxylation or ornithine; 3) two separate aminopropyltransferases are required for spermidine and spermine synthesis; 4) spermine and spermidine are important in the regulation of ornithine decarboxylase and the amines exert this control by a posttranslational modification of the enzyme; and 5) spermidine or spermine is essential for sporulation of yeast and for the maintenance of the double-stranded RNA killer plasmid . Recent studies in amine-deficient mutants of Escherichia coli have shown an important role of the polyamines in protein synthesis in vivo.

J Bacteriol, 1982 Dec, 152(3), 1255 - 64
Activation of chitin synthetase in permeabilized cells of a Saccharomyces cerevisiae mutant lacking proteinase B; Fernandez MP et al.; Digitonin treatment at 30 degrees C of a Saccharomyces cerevisiae mutant lacking proteinase B permeabilized the cells and caused rapid and extensive activation of chitin synthetase in situ . The same result was obtained with a mutant generally defective in vacuolar proteases . By lowering the temperature and using different permeabilization procedures, we showed that increases in permeability and activation are distinct processes . Activation was inhibited by the protease inhibitors antipain and leupeptin, but by pepstatin or chymostatin . Metal chelators were also inhibitory, and their effect was reversed by the addition of Ca2+ but not by Mg2+ . Antipain added together with Ca2+ after incubation of the cells in the presence of a chelating agent prevented reversal of inhibition, a result that was interpreted as indicating that antipain acts either on the same step affected by Ca2+ or on a subsequent step . Efforts to obtain activation in cell-free extracts were unsuccessful, but it was possible to extract the synthetase, once activated, by breaking permeabilized cells with glass beads . Treatment of the cell-free extracts with trypsin led not only to increased activity of chitin synthetase, but also to a change in the pH-activity curve and a diminished requirement by the enzyme for free N-acetylglucosamine . These observations suggest that the modification undergone by the synthetase during endogenous activation is different from that brought about by trypsin treatment.

Nucleic Acids Res, 1982 Nov 25, 10(22), 7283 - 93
Transcripts of mitochondrial tRNA genes in Saccharomyces cerevisiae; Frontali L et al.; The transcription of a group of tRNA genes from the large tRNA gene cluster of mitochondrial DNA from Saccharomyces cerevisiae has been investigated by hybridization with DNA probes carrying tRNA coding sequences and small portions of the A + T rich intergenic regions . Results have shown that in some rho- mutants (DS502, F11) mature tRNA was absent, but a few transcripts could be detected . Some high molecular weight species actually hybridized with DNA probes carrying different tRNA coding sequences . Low molecular weight transcripts (100-150 nucleotides, carrying one tRNA sequence) were also present in these mutants . A high molecular weight transcript was also observed in the wild type, though in much more limited amount . The low molecular weight transcripts were analysed by the S1 mapping technique and found to include both a tRNA sequence and the upstream 5' flanking region extending as far as the 3' end of the preceding tRNA gene . The results suggest the existence of a common transcript bearing several tRNA sequences and indicate a possible mechanism of processing, which might be defective in mutants.

Biochim Biophys Acta, 1982 Nov 24, 719(2), 356 - 62
Transport and metabolic effects of alpha-aminoisobutyric acid in Saccharomyces cerevisiae; Kim KW et al.; alpha-Aminoisobutyric acid is actively transported into yeast cells by the general amino acid transport system . The system exhibits a Km for alpha-aminoisobutyric acid of 270 microM, a Vmax of 24 nmol/min per mg cells (dry weight), and a pH optimum of 4.1-4.3 . alpha-Aminoisobutyric acid is also transported by a minor system(s) with a Vmax of 1.7 nmol/min per mg cells . Transport occurs against a concentration gradient with the concentration ratio reaching over 1000:1 (in/out) . The alpha-aminoisobutyric acid is not significantly metabolized or incorporated into protein after an 18 h incubation . alpha-Aminoisobutyric acid inhibits cell growth when a poor nitrogen source such as proline is provided but not with good nitrogen sources such as NH+4 . During nitrogen starvation alpha-aminoisobutyric acid strongly inhibits the synthesis of the nitrogen catabolite repression sensitive enzyme, asparaginase II . Studies with a mutant yeast strain (GDH-CR) suggest that alpha-aminoisobutyric acid inhibition of asparaginase II synthesis occurs because alpha-aminoisobutyric acid is an effective inhibitor of protein synthesis in nitrogen starved cells.

Eur J Biochem, 1982 Nov 15, 128(2-3), 589 - 95
Regulation of the phosphatidylethanolamine methylation pathway in Saccharomyces cerevisiae; Yamashita S et al.; 1 . Phosphatidyl-N-methylethanolamine methyltransferase mutants of Saccharomyces cerevisiae were isolated . Genetic analysis showed that phosphatidylethanolamine methyltransferase and phosphatidyl-N-methylethanolamine methyltransferase are coded for by separate genes . Phosphatidyl-N-methylethanolamine methyltransferase activity and phosphatidyl-N,N-dimethylethanolamine methyltransferase activity appeared to be catalyzed by the same enzyme . 2 . Phosphatidyl-N-methylethanolamine methyltransferase was found to be repressed by myo-inositol and choline . Both myo-inositol and choline at concentrations of 10 micrograms/ml were required for repression . The decreased enzyme level was restored by the removal of myo-inositol or choline or both . 3 . Both myo-inositol and choline were required for the maximum repression of phosphatidylethanolamine methyltransferase in wild-type cells . In contrast, choline was not required for the repression of the enzyme in mutant strain 172 . This was due to a single nuclear gene mutation in the genome of strain 172 . 4 . The activity of the phosphatidylethanolamine methylation pathway in cells decreased with time on incubation of cells with myo-inositol and choline, myo-Inositol could not be replaced by other structurally related compounds, such as scyllo-inositol or mannitol . 5 . The physiological significance of the repression of the phosphatidylethanolamine methylation pathway is discussed with respect to the mechanism for maintaining the contents of phosphatidylethanolamine and phosphatidylcholine at normal levels.

J Biol Chem, 1982 Nov 10, 257(21), 13081 - 7
The amino acid sequence of cytochrome c oxidase subunit VI from Saccharomyces cerevisiae; Gregor I et al.; The complete amino acid sequence of the nuclearly coded cytochrome c oxidase subunit VI was determined for a genetically defined haploid strain of Saccharomyces cerevisiae . The subunit contains 108 amino acids, has Mr = 12,627, is acidic (net charge of -9.7 at pH 7) and is quite polar (polarity index, 50.9%) . Distribution of charges within the polypeptide chain is highly non-random . The NH2- and COOH-terminal regions are predominantly acidic whereas an apolar and a basic region are found in the interior, Subunit VI shows between 28 and 40% sequence homology (depending on the method of alignment) with subunit V of bovine cytochrome c oxidase; since the yeast subunit VI lacks methionine and contains only a single histidine residue very close to the NH2 terminus, it is unlikely that either of the two subunits carries heme alpha in the native enzyme.

Mol Cell Biol, 1982 Nov, 2(11), 1399 - 409
Chromosomes XIV and XVII of Saccharomyces cerevisiae constitute a single linkage group; Klapholz S et al.; We present several lines of evidence that chromosomes XIV and XVII of Saccharomyces cerevisiae are not independent chromosomes, but rather constitute a single linkage group . Studies which made use of a new mapping method based on the haploidization-without-recombination meiotic phenotype of the spoll mutant initially indicated that markers on chromosomes XIV and XVII were linked . Tetrad analysis was used to establish gene-gene distances, and a new chromosome XIV map incorporating markers originally assigned to chromosome XVII was derived . During the course of trisomic segregation studies, we discovered that a 2n + 2 homothallic diploid, originally believed to be tetrasomic for chromosome XVII (now XIV), carries two normal chromosome XIV homologs and two aberrant homologs which appear to be deficient for a large portion of the right arm of XIV . The previous evidence that established chromosome XVII as an independent linkage group is discussed in the light of these findings.

Mol Cell Biol, 1982 Nov, 2(11), 1299 - 303
Propranolol, atenolol, and trifluoperazine reduce the spontaneous occurrence of meiotic diploid products in Saccharomyces cerevisiae; Sora S et al.; The effect of atenolol, propranolol, trifluoperazine, and caffeine on the occurrence of meiotic diploid and disomic products in Saccharomyces cerevisiae was investigated . We demonstrated that atenolol, propranolol, and trifluoperazine reduce the occurrence of meiotic diploid products and that propranolol also slightly decreases the spontaneous frequency of disomics . On the other hand, caffeine appears to be a powerful inducer of diploid meiotic products, but also shows a lesser effect on disomic induction . Since spontaneous or caffeine-induced diploids arise from a failure of the second meiotic division, it appears that the target of these drugs is at the beginning of the second meiotic division . The only common effect of trifluoperazine and propranolol, mainly investigated in mammals, was an inhibition of calmodulin activity via direct interaction . We tend, therefore, to believe that calmodulin activity must be a crucial point for the second meiotic division to begin . The increased induction of diploids, due to caffeine, may be interpreted as a consequence of an increased cyclic AMP level.

Mikrobiologiia, 1982 Nov-Dec, 51(6), 901 - 4
{Increased permeability of the intracellular membranes in the dehydration and rehydration of Saccharomyces cerevisiae yeasts}; Rapoport AI et al.; A considerable quantity of potassium and magnesium ions was found to be released from Saccharomyces cerevisiae cells being in the state of anabiosis upon their rehydration . The nearly maximal (for each of the experiments) quantity of ions was released as early as when cells dehydrated to a residual humidity of ca . 20% were rehydrated . A further decrease of the residual humidity down to 8-10% did not increase the leakage of the ions when the cells were rehydrated . It was concluded that the permeability of the cytoplasmic and vacuolar membranes for the ions increased when the cells were dehydrated and that this phenomenon should be attributed to the removal of free water from the cells.

J Gen Microbiol, 1982 Nov, 128 (Pt 11), 2591 - 600
Accumulation of 2'-O-methylguanosine deficient tRNATrp in tryptophan limited Saccharomyces cerevisiae; Staheli P et al.; Saccharomyces cerevisiae synthesizes one major tryptophan transfer ribonucleic acid (tRNATrp) species (isoacceptor A) carrying characteristic base modifications . Under tryptophan limited growth conditions wild-type strain X2180-1A exhibited a second important tRNATrp species (isoacceptor B) . The total amount of tRNATrp, approximately 2.5 pmol (mg dry wt)-1, stayed essentially constant during amino acid shift-down experiments . The amount of isoacceptor B relative to total tRNATrp was 10 to 15% during amino acid sufficient exponential growth conditions, but increased during tryptophan limitation three- to four-fold . Analysis of the base compositions showed that isoacceptor B differed from isoacceptor A in one respect only: 2'-O-methylguanosine, a modified guanosine base occurring at position 17 of the major isoacceptor A tRNATrp, was not detectable in hydrolysates of purified isoacceptor B . The biological significance of isoacceptor B is discussed.

Genetics, 1982 Nov, 102(3), 361 - 78
Frameshift suppression in Saccharomyces cerevisiae . V . Isolation and genetic properties of nongroup-specific suppressors; Culbertson MR et al.; Two classes of frameshift suppressors distributed at 22 different loci were identified in previous studies in the yeast Saccharomyces cerevisiae . These suppressors exhibited allele-specific suppression of +1 G:C insertion mutations in either glycine or proline codons, designated as group II and group III frameshift mutations, respectively . Genes corresponding to representative suppressors of each group have been shown to encode altered glycine or proline tRNAs containing four base anticodons.--This communication reports the existence of a third class of frameshift suppressor that exhibits a wider range in specificity of suppression . The suppressors map at three loci, suf12, suf13, and suf14, which are located on chromosomes IV, XV, and XIV, respectively . The phenotypes of these suppressors suggest that suppression may be mediated by genes other than those encoding the primary structure of glycine or proline tRNAs.

J Bacteriol, 1982 Nov, 152(2), 874 - 9
Growth inhibition by alpha-aminoadipate and reversal of the effect by specific amino acid supplements in Saccharomyces cerevisiae; Winston MK et al.; The growth of Saccharomyces cerevisiae wild-type strain X2180 in minimal medium was inhibited by the addition of higher-than-supplementary levels of alpha-aminoadipate . This inhibitory effect was reversed by the addition of arginine, asparagine, aspartate, glutamine, homoserine, methionine, or serine as single amino acid supplements . Mutants belonging to the lys2 and lys14 loci were able to grow in lysine-supplemented alpha-aminoadipate medium, although not as well as when selected amino acids were added . Growth in alpha-aminoadipate medium by all strains was accompanied by an accumulation of alpha-ketoadipate . Glutamate:keto-adipate transaminase levels were derepressed two- to fivefold in lys2 mutants using alpha-aminoadipate as a nitrogen source . Wild-type strain X2180 growing in amino acid-supplemented AA medium exhibited higher levels of alpha-aminoadipate reductase . Mutants unable to use alpha-aminoadipate without amino acid supplementation were obtained by treatment of lys2 strain MW5-64 and were shown to have glutamate: ketoadipate transaminase activity and to lack alpha-aminoadipate reductase activity . Altered cell morphologies, including increased size, multiple buds, pseudohyphae, and germ tubes, evidenced by cells grown in alpha-aminoadipate medium suggest that higher-than-supplementary levels of alpha-aminoadipate result in an impairment of cell division.

J Bacteriol, 1982 Nov, 152(2), 747 - 56
Effects of unsaturated fatty acid deprivation on neutral lipid synthesis in Saccharomyces cerevisiae; Buttke TM et al.; The effects of unsaturated fatty acid deprivation on lipid synthesis in Saccharomyces cerevisiae strain GL7 were determined by following the incorporation of {14C}acetate . Compared to yeast cells grown with oleic acid, unsaturated fatty acid-deprived cells contained 200 times as much 14C label in squalene, with correspondingly less label in 2,3-oxidosqualene and 2,3;22,23-dioxidosqualene . Cells deprived of either methionine or cholesterol did not accumulate squalene, demonstrating that the effect of unsaturated fatty acid starvation on squalene oxidation was not due to an inhibition of cell growth . Cells deprived of olefinic supplements displayed additional changes in lipid metabolism: (i) an increase in 14C-labeled diacylglycerides, (ii) a decrease in 14C-labeled triacylglycerides, and (iii) increased levels of 14C-labeled decanoic and dodecanoic fatty acids . The changes in squalene oxidation and acylglyceride metabolism in unsaturated fatty acid-deprived cells were readily reversed by adding oleic acid . Pulse-chase studies demonstrated that the {14C}squalene and 14C-labeled diacylglycerides which accumulated during starvation were further metabolized when cells were resupplemented with oleic acid . These results demonstrate that unsaturated fatty acids are essential for normal lipid metabolism in yeasts.

Genetics, 1982 Nov, 102(3), 341 - 59
Mutations in the pho80 gene confer permeability to 5'-mononucleotides in Saccharomyces cerevisiae; Bisson LF et al.; Yeast mutants permeable to dTMP (tup) were selected and two new complementation groups (tup5 and tup7) were identified . Assay of the levels of both acid and alkaline phosphatase in cells grown under either repressing (5 mM PO4(-3) or derepressing (0.03 mM PO4(-3) conditions indicated that, in general, tup mutations cause cells to be defective in their regulation of phosphatase synthesis . In addition, three of the tup mutations (tup1, tup4 and tup7) displayed markedly elevated rates of inorganic phosphate transport . The tup7 locus was found to be tightly centromere-linked on the right arm of chromosome XV, and was shown to be allelic with the pho80 regulatory locus on the basis of both genetic and biochemical criteria . Analysis of other mutations known to affect phosphatase levels (pho) indicated that some also conferred permeability to dTMP . Possible allelic relationships between tup genes and certain of these pho mutations are discussed . Regardless of the culture conditions, wild-type strains were not permeable to dTMP; in contrast, it was found in the course of this work that normal yeast cells were permeable to dUMP and that dUMP permeability was regulated by the concentration of inorganic phosphate present in the medium used to grow the cells . Thus, permeability to 5'-mononucleotides appears to be under coordinate control with phosphatase synthesis.

Eur J Biochem, 1982 Nov, 128(1), 179 - 84
Regulation of synthesis of catalases and iso-1-cytochrome c in Saccharomyces cerevisiae by glucose, oxygen and heme; Hortner H et al.; The regulation of the hemoproteins catalase T, catalase A and iso-1-cytochrome c was studied in the yeast Saccharomyces cerevisiae . Levels of catalase T and catalase A mRNAs are low or undetectable in anaerobic and heme-deficient cells, and in wild type strains grown on high glucose concentrations . Regulatory mutants (cgr4 and cas1), which have previously been shown to have high catalase T activity when grown in the absence of oxygen or on high glucose concentrations, have high levels of catalase T mRNA when grown under glucose repression conditions . Whereas no catalase T mRNA could be detected in a heme-deficient (ole3) single mutant, double mutants (ole3 cgr4) and (ole3 cas1) contain mature catalase T mRNA . Catalase T and A mRNAs are accumulated rapidly during adaptation of anaerobic cells to oxygen . Anaerobic and heme-deficient cells lack or have extremely low levels of iso-1-cytochrome c mRNA, which, like catalase mRNAs, is accumulated rapidly during oxygen adaptation . The results obtained demonstrate that glucose, oxygen and heme regulate the synthesis of the hemoproteins studied by controlling mRNA levels . In addition, posttranscriptional, probably translational control has to be postulated at least in the case of catalases, to explain the results obtained.

Biochim Biophys Acta, 1982 Oct 14, 713(1), 86 - 93
Lipid synthesis in inositol-starved Saccharomyces cerevisiae; McCammon MT et al.; Lipid synthesis was analyzed in an inositol-requiring mutant of Saccharomyces cerevisiae (MC13) . Both rates and cellular amounts of {U-14C}acetate incorporation into phospholipids, triacylglycerols, free sterols and steryl esters were elevated in an inositol-starved culture compared to the supplemented control at a time when the deprived culture was losing viability (inositol-less death) . The rates at a later time were greatly reduced . During the period when de novo lipid synthesis was high in the starved culture, phospholipid turnover and presumed conversion to triacylglycerols was also accelerated; no differences were apparent in the turnover of the sterol fractions between the two cultures . No change in the fractional percent of ergosterol or of the sterol precursors could be attributed to inositol starvation . The synthesis and maintenance of membrane lipids (phospholipids and free sterols) and their coupling in cellular metabolism are discussed in light of these results.

J Biol Chem, 1982 Oct 10, 257(19), 11203 - 6
In vivo biosynthesis of the vacuolar proteinases A and B in the yeast Saccharomyces cerevisiae; Mechler B et al.; Proteinase A and proteinase B, two vacuolar enzymes in Saccharomyces cerevisiae, are synthesized as larger precursors with apparent molecular weights of approximately 52,000 and 42,000, respectively . These precursor molecules are processed to their mature forms of 42,000 molecular weight for proteinase A and 33,000 molecular weight for proteinase B . In the presence of tunicamycin, an inhibitor of the synthesis of protein-asparagine linked carbohydrate moieties, two smaller molecular forms each of precursor and mature proteinase A were synthesized, indicating that proteinase A contains N-linked carbohydrate which is apparently not required for processing . Tunicamycin interferes also with the glycosylation of the proteinase B precursor, whereas no unglycosylated mature proteinase B could be detected.

Biochimie, 1982 Oct, 64(10), 859 - 65
{Biosynthesis and metabolism of phosphonolipids and phospholipids in rat hepatocytes and Saccharomyces cerevisiae}; Baraud J et al.; In rat hepatocytes, ciliatine (2 aminoethylphosphonic acid) is incorporated into phosphonolipid (PnE) by the same pathway leading from phosphorylethanolamine to phospholipid (PE) . The two resulting lipids are isolated from mitochondria and microsomes . The rates of biosynthesis are quite comparable; the processes of trimethylation and of in vitro transfer in the presence of a specific exchange protein are very similar . In yeast, on the other hand, the uptake of the two precursors is very slight, suggesting that the direct cytidylic pathway of phospholipid biosynthesis is strongly repressed . Despite small amounts of PE and PnE produced, methylation occurs with a good yield . The good incorporation of ethanolamine may be understood by a base-exchange mechanism . The natural PE biosynthesis is achieved through the decarboxylation of phosphatidylserine, and followed by methylation leading to phosphatidylcholine . The use of very small amounts of precursors does not modify the natural course of phospholipid biosynthesis.

Z Naturforsch {C}, 1982 Oct, 37(10), 916 - 20
The Triton X-100 and high salt resistant residue of Saccharomyces cerevisiae nuclear membranes; Mann K et al.; Saccharomyces cerevisiae nuclear membranes were prepared from isolated nuclei by digesting chromatin with deoxyribonuclease after an initial treatment of nuclei with very diluted buffers . When the nuclear membranes were treated with 5% Triton X-100 and 1M NaCl an insoluble fibrous net was obtained which consisted mainly of protein with Mr values of 85 000, 48 000, 45 000, 39 000 and 31 000 . Lamins, a set of proteins with Mr = 65 000--75 000, which were shown to be the major proteins of the insoluble nuclear membrane residue of higher eukaryotes, were not found.

Mol Cell Biol, 1982 Oct, 2(10), 1205 - 11
Further evidence that the rna2 mutation of Saccharomyces cerevisiae affects mRNA processing; Bromley S et al.; The relative rate at which ribosomal protein 51 (rp51) mRNA is synthesized was measured by pulse-labeling cells in vivo with {3H}adenine . Two strains of Saccharomyces cerevisiae were compared: A364A (wild type) and ts368 (rna2), a temperature-sensitive strain in which the level of rp51 mRNA decreases and an intron-containing rp51 precursor RNA increases . When cells were shifted up to the nonpermissive temperature (36 degrees C), the rate of rp51 RNA synthesis was only marginally affected (75% of wild type) by the presence of the rna2 mutation . The precursor RNA was the predominant transcription product at 36 degrees C . This precursor could be converted into RNA equal in size to mature mRNA by further incubation at either 36 or 23 degrees C in the presence of unlabeled adenine . The relative half-life of the rp51 transcripts at 36 degrees C also decreased approximately twofold in ts368 as compared with A364A . All of these data imply that the precursor (intron-containing) RNA is processed inefficiently to mature mRNA and that the rp51 precursor RNA is continuously synthesized and degraded in the mutant strain at 36 degrees C.

Mol Cell Biol, 1982 Oct, 2(10), 1199 - 204
Sporulation and rna2 lower ribosomal protein mRNA levels by different mechanisms in Saccharomyces cerevisiae; Kraig E et al.; In Saccharomyces cerevisiae, the levels of ribosomal protein mRNAs are regulated coordinately . Vegetative strains carrying the temperature-sensitive rna2 mutation exhibit a dramatic decrease in the levels of most ribosomal protein mRNAs at the restrictive temperature . Similarly, in wild-type cells induced to sporulate by nitrogen starvation, there is a fivefold reduction in the relative synthesis rate of ribosomal proteins . Using Northern gel analysis and cloned ribosomal protein genes, we compared the way in which ribosomal protein mRNA is affected under these two conditions . In vegetative rna2 cells, incubation at 34 degrees C led to the disappearance of ribosomal protein mRNAs and the accumulation of higher-molecular-weight precursor RNAs . A different phenotype was observed during sporulation . Although sporulating conditions led to a significant reduction in the relative abundance of ribosomal protein mRNA, there was no detectable accumulation of precursor RNAs even in rna2/rna2 diploids at 34 degrees C . A suppressor of rna2 and of other rna mutations, SRN1, at least partially relieved the block in the splicing of the ribosomal protein 51 intron in vegetative rna2 cells but did not detectably affect the level of ribosomal protein mRNA in sporulating cells . We concluded that the rna2 mutation and sporulation conditions affected ribosomal protein mRNA metabolism in two quite different ways . In vegetative cells the mutant rna2 effected a block which occurred primarily in post-transcriptional processing, whereas in sporulating cells the ribosomal protein mRNA levels were decreased by some other mechanism, presumably a change in the relative rate of transcription or mRNA turnover . Furthermore, the data suggest that the mutation rna2 has no additional effect on ribosomal protein mRNA metabolism in sporulating cells.

Proc Natl Acad Sci U S A, 1982 Oct, 79(20), 6191 - 5
Specific interactions of Saccharomyces cerevisiae proteins with a promoter region of eukaryotic tRNA genes; Klemenz R et al.; The specific binding of one or several Saccharomyces cerevisiae proteins to a segment of genes that code for different yeast tRNAs has been demonstrated with the use of the DNase I-protection "footprint" assay of Galas and Schmitz . The analyzed binding occurs near the 3' ends of the genes and is centered on an 11-base-pair DNA sequence that has been well conserved among eukaryotic tRNA genes . Others have shown the involvement of this sequence in initiating the transcription of tRNA genes by RNA polymerase III . The adenovirus gene that codes for VAI RNA also contains this conserved sequence element, and we detect binding of yeast protein(s) to this gene . Competition experiments show that a common set of proteins binds to different tRNA genes . The DNA-protein complex is quite stable at 20 degrees C and low ionic strength.

Mutat Res, 1982 Oct-Nov, 102(3), 249 - 59
Genetic effects of ozone: induction of point mutation and genetic recombination in Saccharomyces cerevisiae; Dubeau H et al.; The mutagenicity and recombinogenicity of the atmospheric pollutant, ozone, were investigated in several strains of Saccharomyces cerevisiae . It was observed that ozone induced a variety of genetic events, such as forward and reverse mutations as well as gene conversion and mitotic crossing-over . However, when compared to known mutagens like ultraviolet light, X-rays and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), ozone appeared to be a weak mutagen.

Eur J Biochem, 1982 Oct, 127(2), 411 - 6
Mitochondrial transcripts in glucose-repressed cells of Saccharomyces cerevisiae; Baldacci G et al.; We have compared mitochondrial transcripts from yeast Saccharomyces cerevisiae strain D273-10B grown in the presence of 2% galactose (non-repressed cells) or 15% glucose (glucose-repressed cells) . The ethidium-bromide-stained electrophoretic pattern of mitochondrial RNAs from glucose-repressed cells shows a clear decrease of tRNAs . In addition, some RNA bands appear to be specific for a single growth condition . To identify these RNA species we have performed hybridization experiments with 32P-labelled mitochondrial DNA from petite mutant cells . The mitochondrial repeat units of the mutants retained only one of the following genes: oxi1, oxi2, oxi3, oli2, cob and oli1 . Unchanged amounts of oxi2 and oli2 transcripts and reduced concentrations of oli1 and oxi1 putative mRNAs are present in glucose-repressed cells . In the same growth condition we observe a decreased processing of a precursor RNA species from the split cob gene and reduced amounts of transcripts corresponding to the first, second and fifth intron of the split oxi3 gene . The oxi3 first and second introns, whose transcripts are the most variable, include long open reading frames in their nucleotide sequence, but at present it is not known whether the corresponding RNA species have a functional role . Our results show that their concentrations are related to the growth condition.

Eur J Biochem, 1982 Oct, 127(2), 339 - 42
Reduction of respiratory-chain cytochrome b by lactate in Saccharomyces cerevisiae; Briquet M et al.; Cytochrome b of yeast mitochondria can be reduced by a part of the electrons resulting from the oxidation of lactate enantiomers . 1 . The respiration of D-lactate and L-lactate is 30-40% inhibited by antimycin A . 2 . Reduction of cytochrome b is observed in submitochondrial particles in the presence of low concentration of D-lactate and L-lactate (half-optimal concentration of 4.7 mM and 2.4 mM respectively) in the presence of different bc1 inhibitors . 3 . Reduction of cytochrome b and c1 occurs in purified complex III of yeast in the presence of L-lactate and added L-lactate: NAD+ oxidoreductase . 4 . In the particles obtained from yeast grown in lactate the oxidation of L-lactate involves the reduction of a pigment absorbing at 558 nm.

J Bacteriol, 1982 Oct, 152(1), 517 - 20
DNA replication in a diploid strain of Saccharomyces cerevisiae homozygous for the rad6-1 mutation; Haladus E et al.; The generation time of a diploid strain homozygous for the rad6-1 mutation was 160 min, and the duration of the S phase was 80 min; in the parental heterozygote, these values were 90 and 40 min, respectively . Analysis of DNA sedimentation in an alkaline sucrose gradient revealed that heterozygote high-molecular-weight DNA appeared after 60 min, and homozygote high-molecular weight DNA only after a 100-min pulse.

J Bacteriol, 1982 Oct, 152(1), 111 - 9
Exogenous dTMP utilization by a novel tup mutant of Saccharomyces cerevisiae; Bisson LF et al.; The rate and extent of entry of dTMP were measured in strains of Saccharomyces cerevisiae carrying two new tup mutations (tup5 and tup7) and most of the other tup mutations which have been reported previously by others . The tup7 mutation allowed dramatically greater accumulation of dTMP than any of the other mutations tested . Specific labeling of DNA by {CH3-3H}dTMP, fate of the dTMP pool inside of the cells, and degradation of the dTMP in the culture medium were investigated in strains carrying the tup7 mutation . The extracellular dTMP was not appreciably degraded, and that accumulated intracellularly was readily phosphorylated to dTDP and dTTP . Under optimum labeling conditions, 60 to 80% of the total thymidylate residues in newly synthesized DNA were derived from the exogenously provided dTMP, even in the absence of a block in de novo dTMP biosynthesis . An apparent Km for entry of 2 mM dTMP was found . The tup7 mutation increased permeability to dTMP (and some other 5'-mononucleotides), but did not affect uptake of nucleosides and purine and pyrimidine bases . Uptake of dTMP could be almost completely inhibited by moderate concentrations of Pi . These findings and other observations suggest that entry of dTMP in strains carrying the tup7 mutation is mediated by a permease whose function in normal cells is the transport of Pi.

J Biol Chem, 1982 Sep 25, 257(18), 10846 - 51
Analysis of temperature-sensitive mutant ts 187 of Saccharomyces cerevisiae altered in a component required for the initiation of protein synthesis; Feinberg B et al.; Postpolysomal extracts have been prepared from wild type haploid Saccharomyces cerevisiae cells (wt A364A) and from a temperature-sensitive mutant strain (ts 187, gene prt 1) . The extracts, prepared via spheroplasts and depleted of endogenous mRNA with nuclease, translate exogenous natural mRNA and polyuridylic acid . The activity of wt A364A with respect to translation of yeast mRNA, poly(U)-dependent synthesis of polyphenylalanine which measures elongation components, reactions involved in the initiation of protein synthesis, and termination and release of polypeptides, is not significantly affected when spheroplasts are incubated at 39 degrees C for relatively short periods of time, prior to the preparation of the cell-free system . With extracts obtained from ts 187 cells, preincubation of spheroplasts at 39 degrees C prior to the preparations of the cell-free system markedly decreases the ability to translate natural mRNA but not poly(U) . Compared to extracts from unheated spheroplasts, the following activities in ts 187 extracts from spheroplasts preincubated at 39 degrees C are not significantly affected: activation of methionine and methionylation of tRNAMet; formation of (eukaryotic initiation factor 2.Met-tRNAf.GTP} ternary complex; binding of mRNA to 40S preinitiation intermediate containing Met-tRNAf, and joining of 60S subunits to form the 80S initiation complex; elongation factor 1- and elongation factor 2-dependent elongations reactions; and termination and release of completed polypeptide chains . However, the interaction between the {eukaryotic initiation factor 2.Met-tRNAf.GTP} ternary complex and 40 S subunits, to form the 40 S preinitiation complex, is drastically inhibited by treatment of the spheroplasts at 39 degrees C.

Biochim Biophys Acta, 1982 Sep 24, 691(1), 144 - 50
L-Proline transport in Saccharomyces cerevisiae; Horak J et al.; Transport of L-proline into Saccharomyces cerevisiae K is mediated by two systems, one with a KT of 31 microM and Jmax of 40 nmol . s-1 . (g dry wt.)-1, the other with KT greater than 2.5 mM and Jmax of 150-165 nmol . s-1 . (g dry wt.)-1 . The kinetic properties of the high-affinity system were studied in detail . It proved to be highly specific, the only potent competitive inhibitors being (i) L-proline and its analogs L-azetidine-2-carboxylic acid, sarcosine, D-proline and 3,4-dehydro-DL-proline, and (ii) L-alanine . The other amino acids tested behaved as noncompetitive inhibitors . The high-affinity system is active, has a sharp pH optimum at 5.8-5.9 and, in an Arrhenius plot, exhibits two inflection points at 15 degrees C and 20-21 degrees C . It is trans-inhibited by most amino acids (but probably only the natural substrates act in a trans-noncompetitive manner) and its activity depends to a considerable extent on growth conditions . In cells grown in a rich medium with yeast extract maximum activity is attained during the stationary phase, on a poor medium it is maximal during the early exponential phase . Some 50-60% of accumulated L-proline can leave cells in 90 min (and more if washing is done repeatedly), the efflux being insensitive to 0.5 mM 2,4-dinitrophenol and uranyl ions, the pH between 3 and 7.3, as well as to the presence of 10-100 mM unlabeled L-proline in the outside medium . Its rate and extent are increased by 1% D-glucose and by 10 micrograms nystatin per ml.

J Bacteriol, 1982 Sep, 151(3), 1123 - 8
New genes involved in carbon catabolite repression and derepression in the yeast Saccharomyces cerevisiae; Entian KD et al.; A mutation causing resistance to carbon catabolite repression in gene HEX2, mutant allele hex2-3, causes an extreme sensitivity to maltose when in combination with the genes necessary for maltose metabolism . This provided a convenient system for the selective isolation of mutations in genes specifically required for maltose metabolism and other genes involved in general carbon catabolite repression . In addition to reversion of the hex2-3 allele, mutations in three other genes were detected . These genes were called CAT1, CAT3, and MUR1 and in a mutated form abolished maltose inhibition caused by mutant allele hex2-3 . Mutant alleles cat1 and cat3 also restored normal repression in the presence of the hex2-3 allele . Segregants having only mutant alleles cat1 or cat3 were obtained by tetrad analysis . These segregants could not grow on nonfermentable carbon sources . Mutant alleles of gene CAT1 were allelic to a mutant allele cat1-1 previously isolated (Zimmermann et al., Mol . Gen . Genet . 151:95-103) . Such mutants prevented derepression not only of the maltose catabolizing system, the selected property, but also of glyoxylate shunt and gluconeogenic enzymes . However, respiratory activities and invertase formation were not affected under derepressing conditions . cat3 mutants had the same phenotypic properties as cat1 mutants . This showed that carbon metabolism in yeast cells is under a very complex and ramified control of repressing and derepressing genes, which are interdependent.

Mol Cell Biol, 1982 Sep, 2(9), 1088 - 95
Isolation and characterization of mutants that produce the allantoin-degrading enzymes constitutively in Saccharomyces cerevisiae; Chisholm G et al.; Degradation of allantoin, allantoate, or urea by Saccharomyces cerevisiae requires the participation of four enzymes and four transport systems . Production of the four enzymes and one of the active transport systems is inducible; allophanate, the last intermediate of the pathway, functions as the inducer . The involvement of allophanate in the expression of five distinct genes suggested that they might be regulated by a common element . This suggestion is now supported by the isolation of a new class of mutants (dal80) . Strains possessing lesions in the DAL80 locus produce the five inducible activities at high, constitutive levels . Comparable constitutive levels of activity were also observed in doubly mutant strains (durl dal80) which are unable to synthesize allophanate . This, with the observation that arginase activity remained at its uninduced, basal level in strains mutated at the DAL80 locus, eliminates internal induction as the basis for constitutive enzyme synthesis . Mutations in dal80 are recessive to wild-type alleles . The DAL80 locus has been located and is not linked to any of the structural genes of the allantoin pathway . Synthesis of the five enzymes produced constitutively in dal80-1-containing mutants remains normally sensitive to nitrogen repression even though the dal80-1 mutation is present . From these observations we conclude that production of the allantoin-degrading enzymes is regulated by the DAL80 gene product and that induction and repression of enzyme synthesis can be cleanly separated mutationally.

Mol Cell Biol, 1982 Sep, 2(9), 1080 - 7
Mitotic chromosome loss induced by methyl benzimidazole-2-yl-carbamate as a rapid mapping method in Saccharomyces cerevisiae; Wood JS; Mitotic chromosome loss induced by methyl benzimidazole-2-yl-carbamate has been utilized as a rapid and simple method for assigning genes to individual chromosomes in Saccharomyces cerevisiae . This technique relied on the segregation of heterozygous markers in a diploid strain after methyl benzimidazole-2-yl-carbamate treatment due to loss of whole chromosomes . Correlations between the expression of an unmapped gene and that of a previously mapped recessive marker indicated chromosomal linkage . Depending on whether the unmapped gene and the marker were located in coupling or in repulsion, either positive or negative correlations were seen . The chromosomal location of several previously mapped genes were confirmed as a test of the method, and one previously unmapped gene, nib1, was mapped.

Mol Cell Biol, 1982 Sep, 2(9), 1064 - 79
Genetic effects of methyl benzimidazole-2-yl-carbamate on Saccharomyces cerevisiae; Wood JS; The genetic effects of the mitotic inhibitor methyl benzimidazole-2-yl-carbamate (MBC) have been studied in Saccharomyces cerevisiae . MBC had little or no effect on the frequency of mutation . In some experiments MBC caused an increase in the frequency of mitotic recombination; however, this effect was small and not reproducible . The primary genetic effect of MBC was to induce mitotic chromosome loss at a high frequency . Chromosome loss occurred at equal frequencies for all chromosomes tested (13 of 16) . Cells which had lost multiple chromosomes were found more frequently than predicted if individual chromosome loss events were independent . The probability of loss for a particular chromosome increased with length of time cells were incubated with MBC . MBC treatment also increased the frequency at which polyploid cells were found . These results suggested that MBC acted to disrupt the structure or function of the mitotic spindle and cause chromosome nondisjunction.

Mol Cell Biol, 1982 Sep, 2(9), 1052 - 63
Mating-defective ste mutations are suppressed by cell division cycle start mutations in Saccharomyces cerevisiae; Shuster JR; Temperature-sensitive mutants which arrest in the G1 phase of the cell cycle have been described for the yeast Saccharomyces cerevisiae . One class of these mutants (carrying cdc28, cdc36, cdc37, or cdc39) forms a shmoo morphology at restrictive temperature, characteristic of mating pheromone-arrested wild-type cells . Therefore, one hypothesis to explain the control of cell division by mating factors states that mating pheromones arrest wild-type cells by inactivating one or more of these CDC gene products . A class of mutants (carrying ste4, ste5, ste7, ste11, or ste12) which is insensitive to mating pheromone and sterile has also been described . One possible function of the STE gene products is the inactivation of the CDC gene products in the presence of a mating pheromone . A model incorporating these two hypotheses predicts that such STE gene products will not be required for mating in strains carrying an appropriate cdc lesion . This prediction was tested by assaying the mating abilities of double mutants for all of the pairwise combinations of cdc and ste mutations . Lesions in either cdc36 or cdc39 suppressed the mating defect due to ste4 and ste5 . Allele specificity was observed in the suppression of both ste4 and ste5 . The results indicate that the CDC36, CDC39, STE4, and STE5 gene products interact functionally or physically or both in the regulation of cell division mediated by the presence or absence of mating pheromones . The cdc36 and cdc39 mutations did not suppress ste7, ste11, or ste12 . Lesions in cdc28 or cdc37 did not suppress any of the ste mutations . Other models of CDC and STE gene action which predicted that some of the cdc and ste mutations would be alleles of the same locus were tested . None of the cdc mutations was allelic to the ste mutations and, therefore, these models were eliminated.

J Gen Microbiol, 1982 Sep, 128 (Pt 9), 2133 - 40
Abnormalities in cell division induced by diepoxybutane in rad1-1 and rad3 mutants of Saccharomyces cerevisiae; Zaborowska D et al.; Saccharomyces cerevisiae mutants rad1-1 and rad3 differ from the wild-type and from other UV-sensitive rad mutants in their behaviour after transfer from medium containing 1,2:3,4-diepoxybutane (DEB) to DEB-free medium . In both mutants several post-treatment cell cycles proceed in the absence of cell wall separation, resulting in the formation of multicellular chains or aggregates . In this study, electron and light microscopy revealed that at least one post-treatment budding cycle is accompanied by nuclear division while subsequent cell cycles can proceed in the absence of regular nuclear cycles . At low percentage survival levels, the first post-treatment budding cycle was not delayed and was accompanied by significant incorporation of radioactivity into DNA and protein . In contrast, subsequent cell cycles were found to be accompanied by only protein synthesis and not DNA synthesis . The wild-type strain, unlike the mutants, responded to DEB treatment by a dose-dependent lag in the onset of macromolecular synthesis and cell proliferation, and after prolonged incubation in mutagen-free medium the culture consisted of single budded and unbudded cells.

Mutat Res, 1982 Sep, 102(2), 123 - 36
Genetic effects of fresh cigarette smoke in Saccharomyces cerevisiae; Gairola C; Ability of fresh cigarette smoke from University of Kentucky reference cigarette 2R1 to induce gene conversion, reverse mutation and mitotic crossing-over in strain D7 of Saccharomyces cerevisiae was examined . A closed cell suspension-recycle system using 2 peristaltic pumps interconnected to a single-port reverse-phase smoking machine was developed to provide complete exposure of cells to smoke within 0.2--10 sec of its generation . The exposed cells showed a dose-dependent increase in the frequency of all the 3 genetic endpoints examined . Cell age was an important factor with younger cells being more sensitive than older . Filtration studies showed that the gas phase possessed as much as 25% of the total whole-smoke activity . Activated charcoal reduced the activity of smoke in direct proportion to its amount in the filter . Acetate filter did not appreciably alter the activity . A comparison of whole smoke from various cigarettes showed that: (1) the nicotine content of a cigarette does not affect the genetic activity of smoke; (2) burley and flue-cured tobaccos have differential activity in gene conversion and reverse mutation systems; and (3) the genetic effects of whole smoke are not peculiar to tobacco pyrolysis because similar effects are produced by smokes from lettuce and other non-tobacco cigarettes . It is concluded that the yeast D7 system can be used effectively for the quantitative evaluation of genetic effects of smoke from different cigarettes, and both whole cigarette smoke and its gas phase possess mutagenic as well as recombinogenic activity that can be modified by the use of filters.

Lipids, 1982 Sep, 17(9), 662 - 5
Effect of altered sterol composition on the osmotic behavior of sphaeroplasts and mitochondria of Saccharomyces cerevisiae; McLean-Bowen CA et al.; The effect of sterols on the osmotic stability of mitochondrial and plasma membranes of yeast wild-types and mutants that are defective in ergosterol biosynthesis has been studied . Incorporation of the nonfungal sterol, cholesterol, into yeast membranes reduces membrane elasticity which is observed as an increased susceptibility to osmotic lysis . However, the wild-type and nystatin-resistant strains which were examined indicate that qualitative alterations in endogenously generated sterols do not affect resistance to swelling . Although these strains exhibit differences in membrane fluidity, which is influenced by the sterol accumulated by the organisms, the membrane stretching capacity shows no distinct dependence on sterol structure or bilayer fluidity.

Mol Cell Biol, 1982 Sep, 2(9), 1025 - 32
Genetic selection for reciprocal translocation at chosen chromosomal sites in Saccharomyces cerevisiae; Potier S et al.; We have constructed viable Saccharomyces cerevisiae strains containing a reciprocal translocation between the URA2 site of chromosome X and the HIS3 site of chromosome XV . Our methodology is an extension of the method originally developed to introduce an altered cloned sequence at the chromosomal location from which the parent sequence was derived (S . Scherer and R.W . Davis, Proc . Natl . Acad . Sci . U.S.A . 76:4951-4955, 1979) . It comprises three essential steps . First, a nonreverting ura2- strain was constructed by deleting a 3.7-kilobase fragment from the coding sequence of the wild-type URA2 gene . Second, part of the coding sequence of the wild-type URA2 gene (without promotor) was inserted at the HIS3 locus of the ura2- strain . Third, after several generations of growth on uracil-supplemented medium, ura2+ colonies were selected which resulted from mitotic recombination between the nonoverlapping deletions of URA2 located on chromosomes X and XV.

Eur J Biochem, 1982 Sep 1, 126(3), 617 - 22
Effect of caffeine on glucose-induced inactivation of gluconeogenetic enzymes in Saccharomyces cerevisiae . A possible role of cyclic AMP; Tortora P et al.; The mechanism of catabolite inactivation of three gluconeogenetic enzymes, fructose-1,6-bisphosphatase, cytoplasmic malate dehydrogenase and phosphoenolpyruvate carboxykinase, has been studied in the yeast Saccharomyces cerevisiae . The glucose-induced inactivation of the three enzymes is remarkably retarded by preincubation of the cells with different caffeine concentrations; however, a full conservation of activity has never been obtained, even at the highest drug concentration . Caffeine modifies the metabolic effects produced in the yeast cell by exposure to glucose . It reduces the consumption rate of glucose; changes the glycolytic intermediate pattern, giving rise to a crossover point at the level of the phosphofructokinase/fructose-bisphosphatase cycle; and increases the ATP level and the energy charge . Moreover, it substantially reduces the peak of intracellular cAMP content that immediately follows glucose entry; the magnitude of this effect is dependent on the drug concentration . The effect on the change of intracellular cAMP level appears, among all metabolic effects determined by caffeine, the only plausible one to explain the interference with catabolite inactivation of enzymes . Actually a strong negative correlation between residual activity of each of the three investigated enzymes and intracellular cAMP level has been demonstrated . The existence of a common mechanism of action of cAMP, as the mediating factor for catabolite inactivation of all three enzymes, is proposed.

Biochim Biophys Acta, 1982 Aug 12, 689(3), 429 - 36
The influence of uncouplers on facilitated diffusion of sorbose in Saccharomyces cerevisiae; Van den Broek PJ et al.; Sorbose uptake in Saccharomyces cerevisiae, strain Delft 1, proceeds via mediated passive transport . In the cell sorbose is distributed in at least two compartments . Efflux studies showed that sorbose uptake in one of these compartments is not readily reversible . Uncouplers of oxidative phosphorylation inhibit both transport velocity and steady-state uptake level . It could be shown that these two effects are caused by different modes of action of the uncouplers . None of these two effects could be ascribed to changes of the electrochemical H+ gradient or of the intracellular pH . It is suggested that the inhibition of uptake velocity is caused by binding of the uncoupler to the sorbose translocator, thus lowering the transport activity . The uncoupler binding site is probably located at the intracellular fragment of the carrier . The second effect, reduction of the steady-state uptake level, is probably due to blocking of sorbose influx into the compartment that exhibits poor reversibility.

Biochim Biophys Acta, 1982 Aug 6, 717(2), 355 - 68
Biochemical characterization of the OXI mutants of the yeast Saccharomyces cerevisiae; Keyhani E et al.; OXI mutants in Saccharomyces cerevisiae lack a functional cytochrome c oxidase . Wild type and OXI mutants were grown in the presence of radioactive delta-amino{14C}levulinic acid, a precursor of porphyrin and heme, and {3H}mevalonic acid, a precursor of the alkyl side-chain of heme a . SDS polyacrylamide gel electrophoresis of the delipidated mitochondria showed that delta-amino{14C}levulinic acid was distributed into three bands migrating in the regions of Mr 28 000, 13 500, and 10 000, while {3H}mevalonic acid was found in a single band with apparent Mr of 10 000 . The immunoprecipitates obtained by incubating the solubilized mitochondria of any OXI mutant with antibodies against cytochrome c oxidase, showed, after delipidation, a high specific radioactivity due to delta-amino{14C}levulinic acid and {3H}mevalonic acid . This suggested that a prophyrin a was present in all these OXI mutants . HCl fractionation confirmed the presence of porphyrin a in the apooxidase of these mutants . Atomic absorption spectra of the immunoprecipitate of cytochrome c oxidase showed that copper was not detectable in the mutant OXI IIIa which lacked subunit 1, but was present in the mutant OXI IIIb, which exhibited a minor alteration in the electrophoretic mobility of subunit 1 . In OXI I and II mutants there was a 50% reduction in the amount of copper in the immunoprecipitated cytochrome c oxidase . These observations may be interpretable as follows: (1) alterations in polypeptide biosynthesis due to the OXI mutations lead to an improper configuration of cytochrome c oxidase, so that ferrochelatase cannot transfer iron into porphyrin a; (2) subunit I is the binding site for copper, but the mutations in subunits II and III alter the binding site of one of the two copper atoms in subunit I.

Mol Cell Biol, 1982 Aug, 2(8), 939 - 48
Genetic control of excision of Saccharomyces cerevisiae interstrand DNA cross-links induced by psoralen plus near-UV light; Miller RD et al.; Excision of interstrand DNA cross-links induced by 4,5',8-trimethyl psoralen plus 360-nm light was examined in wild type (RAD+) and various radiation-sensitive (rad) mutants of Saccharomyces cerevisiae known to be defective in the excision of UV light-induced pyrimidine dimers . Alkaline sucrose sedimentation of DNA after incubation of psoralen-plus-light-treated cells indicated little or no nicking of cross-linked DNA in rad1-2, rad2-5, rad3-2, rad4-4, rad10-2, and mms19-1 mutants . In the rad14-2 mutant, substantial nicking was observed but to a much lesser extent than in the RAD+ strains, whereas the rad16-1 mutant was as proficient in nicking as the RAD+ strain . Removal of cross-links was also examined in RAD+, rad3-2, and rad14-2 strains by determining the sensitivity of alkali-denatured and -neutralized DNA to hydrolysis by S1 nuclease . No cross-link removal was observed in the rad3-2 mutants, and the rad14-2 mutant was much less efficient than the RAD+ strain in removing cross-links.

Mol Cell Biol, 1982 Aug, 2(8), 897 - 903
A and alpha supernatant pretreatment of Saccharomyces cerevisiae cells affects both the kinetics and efficiency of mating; Sena EP; The effects of culture supernatant treatment on subsequent matings between pretreated a and alpha Saccharomyces cerevisiae cells were studied . For each experiment, pairs of a and alpha {rho+} or {rho- rho0} cells in the logarithmic growth phase in defined minimal medium were pretreated for a total of 15 min (by exchanging their cell-free supernatants or by mixing samples of a and alpha cell cultures) and then mated in defined minimal (YNB) or enriched (YEP) liquid medium . All pretreated cells, regardless of treatment procedure, initiated cell fusion 15 to 35 min faster than did their nontreated counterparts . In all cases, pretreated cells mated 8 to 20% more efficiently than did nonpretreated ones . Regardless of the strains, the hierarchy of mating efficiency was always treated YEP greater than untreated YEP greater than treated YNB greater than untreated YNB . The cell fusion kinetics in alpha {rho+} X a {rho-} crosses were most affected by pretreatment (delta 30 to 35 min), whereas {rho+} X {rho+} crosses were least affected (delta 15 min) . These results are discussed in relation to the functions known for a and alpha pheromones . The successful pretreatment regimes were used to design new rapid and efficient techniques for mating YNB-grown log-phase cells in either YNB or YEP liquid media . These techniques can be used for small- or large-scale mating, and because of their inherent media flexibility, they have many potential applications to future studies on mating-specific or intrazygotic phenomena.

Eur J Cell Biol, 1982 Aug, 28(1), 98 - 102
Ultrastructure of mitotic spindles isolated from a cell division cycle mutant of the yeast, Saccharomyces cerevisiae; King SM et al.; Mitotic spindles were isolated from a temperature-sensitive cell division cycle mutant of Saccharomyces cerevisiae arrested in medical nuclear division by incubation at 36.5 degrees C for 5 h . Cell walls were removed and the resulting sphaeroplasts lyzed on an air water interface . Spindles were collected on electron microscope grids for examination following positive or negative staining . The poles of the spindle consisted of a pair of quadrilaminar spindle pole bodies (SPBs) which measured 160 nm in diameter . The outer, cytoplasmic layer of the SPB was resolved into distinct subunits from which 1 to 3 cytoplasmic microtubules were occasionally seen to emerge . The inner, nuclear, face of the SPB was associated with two families of spindle microtubules; a bundle of 5 to 10 continuous microtubules 1.5 to 2.5 microns long with both ends associated with an SPB, and up to 17 shorter discontinuous microtubules radiating from each SPB and ending at no obvious structure.

Biokhimiia, 1982 Aug, 47(8), 1401 - 8
{Mitochondrial proteinases of the yeast Saccharomyces cerevisiae: electrophoretic detection, substrate specificity and sensitivity to inhibitors}; Novikova LA et al.; The proteins of submitochondrial particles solubilized with 0.1% Triton X-100 were separated by polyacrylamide gel electrophoresis . Hydrolysis of several proteinase substrates was registered directly in the gel after completion of electrophoresis . According to the data obtained the inner mitochondrial membrane contains one or two enzymes which catalyze hydrolysis of cytochrome c as well as one or two enzymes splitting synthetic substrate of trypsin-like proteinases, e . g . N-alpha-benzoyl-L-arginine-p-nitroanilide (BAPA) and N-alpha-benzoyl-L-arginine-beta-naphthylamide (BANA) . Submitochondrial particles were shown to catalyze hydrolysis of 3H-labelled cytochrome c . This activity is suppressed by the same inhibitors as the hydrolysis of mitochondrial translation products, i . e . phenyl-methylsulfonylfluoride, p-chloromercuribenzosulfonate, leupeptin and antipain . Presumably these two processes are catalyzed by the same enzyme localized in the inner mitochondrial membrane . Physiological functions of BAPA- and BANA-hydrolyzing enzyme(s) are still unclear.

Arch Microbiol, 1982 Aug, 132(2), 144 - 8
Metabolism of Saccharomyces cerevisiae envelope mannoproteins; Pastor FI et al.; By pulse and chase labeling experiments, two independent mannoprotein pools have been found associated with the Saccharomyces cerevisiae envelope . One of them probably corresponds to mannoproteins localized in the periplasmic space . These molecules showed a high turnover rate at 28 degrees C . The second pool is formed by intrinsic wall mannoproteins which are apparently stable for long periods of time, after a small initial turnover . These results suggest that at least part of the mannoproteins initially found in the periplasmic space may move into the wall . The time lag between the addition of the radioactive precursors and their incorporation in the cell envelope (20-30 min for amino acids and about 10 min for carbohydrate) indicates that protein formation and carbohydrate incorporation take place in succession . Moreover, bulk glycosylation of mannoproteins seems to occur close in time to the moment of secretion into the periplasmic space.

Arch Microbiol, 1982 Aug, 132(2), 141 - 3
Isolation and characterization of a mutant of Saccharomyces cerevisiae defective in phosphoenolpyruvate carboxykinase; Perea J et al.; A mutant of Saccharomyces cerevisiae lacking phosphoenolpyruvate carboxykinase (E.C . 4.1.1.32) was isolated . The mutant did not grow on gluconeogenic sources except glycerol . The mutation was recessive and apparently affected the structural gene of the enzyme . Intracellular levels of metabolites related to the metabolic situation of the enzyme were not significantly affected after transfer of the mutant from a medium with glycerol to a medium with ethanol as carbon source . In these conditions only AMP decreased 3 to 5 times . A search for mutants affected in the other gluconeogenic enzyme, fructose 1,6 bisphosphatase, remained unsuccessful.

Proc Natl Acad Sci U S A, 1982 Aug, 79(15), 4706 - 8
Ribosomal protein L3 is involved in replication or maintenance of the killer double-stranded RNA genome of Saccharomyces cerevisiae; Wickner RB et al.; Ability to secrete the K1 (or K2) toxin protein and immunity to that toxin {the K1 (or K2) killer trait} are determined by a double-stranded (ds) RNA, called M1 (or M2), whose replication and maintenance depend on at least one of the larger (L) ds RNAs and 29 chromosomal genes, called MAK genes (maintenance of killer) . The location of the MAK8 gene near TCM1 (trichodermin resistance) on the yeast map suggested the possible identity of these two genes . Of six independently isolated tcm1 mutants, five were clearly mak-, and the sixth was weakly mak- . In each case, the mak- phenotype and the trichodermin-resistant phenotypes cosegregated in meiosis and showed the expected tight linkage to pet17 . The mak- mutations in the trichodermin-resistant strains did not complement mak8-1, indicating that MAK8 and TCM1 are the same gene . The mak8-1 mutation does not make strains resistant to trichodermin, and one tcm1 mutation is only slightly mak- . Whereas tcm1 mutants lose M1 or M2 ds RNA, they do not lose L ds RNA . Because TCM1 codes for ribosomal protein L3 {Fried, H . M . & Warner, J . R . (1981) Proc . Natl . Acad . Sci, USA 78, 238--242}, we conclude that ribosomal protein L3 is involved in the replication and maintenance of M ds RNA . Mutations in cyh2 or cry1, producing resistance to cycloheximide and crytopleurine due to mutant ribosomal proteins, do not produce a mak- phenotype . In analogy with bacterial ribosome assembly mutants, yeast low-temperature-sensitive (lts) mutants may have defective ribosomes . We thus examined mutants for an effect on the killer system . An lts5 mutant, unable to grow at 5 degrees C, also has a mak- phenotype (at 30 degrees C) that cosegregates in meiosis with the lts- phenotype . Mutations in seven other lts genes do not result in the mak- phenotype.

Mutat Res, 1982 Aug, 95(2-3), 213 - 24
Genetic control of budding-cell resistance in the diploid yeast Saccharomyces cerevisiae exposed to gamma-radiation; Rao BS et al.; The gamma-radiation response of stationary and budding cells of wild-type diploid strains (RAD) and radiation-sensitive strains rad2, 6, 9, 18, 50-55, 57 and rec4 was studied . As compared with the wild-type strains, mutants generally showed enhanced sensitivity in both stages of the cell cycle . Budding-cell resistance was totally absent from rad50-55 strains . Mutants rad6, 9 and 18 showed some degree of budding-cell resistance . The response of rad2 and rec4 strains was identical with that of the corresponding wild-type strains . These results suggest that the pathway dependent upon the expression of RAD50-55 loci functions more efficiently in budding cells compared with the pathway dependent on RAD2 and RAD6, 9 and 18 loci . Recombination between sister chromatids appears to play an important role in budding-cell resistance, and this process is under the control of the RAD52 repair pathway . The relationship between the repair pathways associated with budding-cell resistance and post-irradiation cellular recovery (LHR) is discussed.

J Biol Chem, 1982 Jul 25, 257(14), 8177 - 82
Exonuclease II from Saccharomyces cerevisiae . An enzyme which liberates 5'-deoxyribomononucleotides from single-stranded DNA by a 5' goes to 3' mode of hydrolysis; Villadsen IS et al.; A DNase, designated Exonuclease II, has been purified 2,000-fold from whole cell extracts of bakers' yeast . It degrades single-stranded but not double-stranded DNA . The enzyme has a pH optimum around 8 and requires at least 2 mM Mg2+ for activity . It is slightly stimulated by dithiothreitol and inhibited by N-ethylmaleimide, suggesting that --SH groups are essential for activity; heparin also inhibits the activity . With 0.1 enzyme unit the Km has been determined to 4 nM of DNA ends and Vmax to 0.52 pmol of liberated nucleotides per min . The Mr is around 120,000 . The enzyme does not degrade circular single-stranded M13 DNA, whereas linearized M13 DNA is degraded . The products are 5'-deoxyribomononucleotides exclusively . Using 5'-end labeled and 3'-end labeled DNA fragments as substrates, partially degraded DNA is only detectable in the latter case, showing that the exonuclease solely attacks DNA from the 5'-end . This distinguishes Exonuclease II from other exonucleases degrading DNA from the 5'-end, since they all either produce a mixture of 5'-mono- and oligonucleotides or 3'-mononucleotides . Analysis of 3'-end labeled fragments after incubation shows that the rate of exonucleolytic cleavage was dependent on the DNA sequence at the 5'-end.

J Biol Chem, 1982 Jul 25, 257(14), 8405 - 11
Alterations of transcription during heat shock of Saccharomyces cerevisiae; Finkelstein DB et al.; The pattern of polyadenylated RNA of the yeast Saccharomyces cerevisiae changes dramatically upon heat shock . By in vitro translation, we have demonstrated that a 2.9-kilobase heat shock RNA encodes the Mr = 90,000 yeast heat shock protein . Heat shock-responsive genes have been isolated by differential plaque filter hybridization of a recombinant library of yeast DNA inserted in the vector lambda Charon 4 . The putative yeast gene products of a number of these recombinants molecules has been determined by in vitro translation of hybrid-selected RNA . We have used one of these hybrid phages (lambda Yhsil) to demonstrate that the heat shock-induced alteration in the level of the 2.9-kilobase polyadenylated RNA which encodes the Mr = 90,000 yeast heat shock protein is regulated at the level of transcription.

Biochim Biophys Acta, 1982 Jul 14, 689(1), 38 - 44
Turnover of protein components of the plasma membrane of Saccharomyces cerevisiae; Herrero E et al.; The peptide composition of plasma membrane in Saccharomyces cerevisiae cells growing at different temperatures between 18 and 38 degrees C was studied using SDS-polyacrylamide gel electrophoresis . Stability of the proteins, both qualitative and quantitative, was observed at the tested temperatures . Treatment for 2 h with cycloheximide decreased by about 50% the amount of a 80 kDa membrane peptide at 18, 23, 28 and 33 degrees C, with no other apparent effects . At 38 degrees C the 80 kDa peptide was not affected by the presence of the drug . Addition of tunicamycin to cultures at concentrations partially inhibitory to growth caused a large accumulation of the 80 kDa peptide in the plasma membrane, which cycloheximide did not modify . Pulse-chase experiments indicated a low rate of turnover of total plasma membranes in cells growing at 18 and 28 degrees C . In contrast, at 38 degrees C about 50% of the radioactivity in plasma membranes disappeared after a 2 h chase . The 80 kDa protein band was the only one with significant differential decay.

J Bacteriol, 1982 Jul, 151(1), 311 - 27
Quantitative analysis of the heat shock response of Saccharomyces cerevisiae; Miller MJ et al.; Transient protein synthesis in Saccharomyces cerevisiae, after shift from 21-23 degrees C to 37 degrees C, was quantitatively analyzed . Pulse-labeled proteins were separated by two-dimensional gel electrophoresis, and autoradiograms of the gels were analyzed by a recently described method involving a computer-coupled film scanning system . In this way, the rate of incorporation of L-{35S}methionine into approximately 500 proteins was followed . The synthesis of more than 80 of these proteins was transiently induced at 37 degrees C, with about 20 being classified as major heat shock proteins (defined as those whose rate of labeling was increased at least eightfold at some time during the response) . The synthesis of more than 300 of the proteins was transiently repressed at 37 degrees C, and several general temporal patterns of repression could be distinguished . The influence of temperature-sensitive mutations affecting RNA synthesis and transport on the heat shock response was also examined . A protein whose induction in response to heat shock has a post-transcriptional component could be identified . As previously pointed out, the heat shock repression of certain proteins is so rapid that it also must involve post-transcriptional effects.

Mol Cell Biol, 1982 Jul, 2(7), 731 - 6
Genetic and biochemical study of threonine-overproducing mutants of Saccharomyces cerevisiae; Delgado MA et al.; Three threonine-overproducing mutants were obtained as prototrophic revertants of a hom3 mutant strain of Saccharomyces cerevisiae . The gene HOM3 codes for aspartokinase (aspartate kinase; EC 2.7.2.4), the first enzyme of the threonine-methionine biosynthetic route, which is subjected to feedback inhibition by threonine . Enzymatic studies indicated that aspartokinase from the revertants has lost the feedback inhibition, resulting in overproduction of threonine . These revertants also bore one or two additional mutations, named tex1-1 and tex2-1, which alone or jointly made possible the excretion of the threonine accumulated . The effect of these two genes on excretion is potentiated by excess inositol in the medium.

Proc Natl Acad Sci U S A, 1982 Jul, 79(14), 4243 - 7
Purification of the cdc8 protein of Saccharomyces cerevisiae by complementation in an aphidicolin-sensitive in vitro DNA replication system; Kuo CL et al.; DNA synthesis in vitro in Brij-treated Saccharomyces cerevisiae requires the product of the CDC8 gene (Hereford, L . M . & Hartwell, L . H . (1971) Nature (London) New Biol . 234, 171-172) . Extracts of wild-type A364a yeast restore DNA synthesis in Brij-treated cdc8, a mutant containing a thermolabile cdc8 gene product . This constitutes a complementation assay by which the cdc8 gene product can be monitored during purification . A heat-stable protein responsible for this complementation has been partially purified from both wild-type A364a cells and from a cdc8 temperature-sensitive mutant . The complementation activity from the mutant is thermolabile when compared to the wild-type activity, indicating that CDC8 is the structural gene for the protein.

J Antibiot (Tokyo), 1982 Jul, 35(7), 875 - 81
Inhibition of protein synthesis in Saccharomyces cerevisiae by the 12,13-epoxytrichothecenes trichodermol, diacetoxyscirpenol and verrucarin A . Reversibility of the effects; Hernandez F et al.; Inhibition of protein synthesis by trichodermol, diacetoxyscirpenol and verrucarin A in cells and spheroplasts of Saccharomyces cerevisiae was investigated . Inhibition was reversible for trichodermol and diacetoxyscirpenol, both drugs being removed from their target site(s) by washing, but was reversible for verrucarin A . These results are interpreted in relation to variations in chemical structure between these trichothecenes.

Genetics, 1982 Jul-Aug, 101(3-4), 369 - 404
Recombination between genes located on nonhomologous chromosomes in Saccharomyces cerevisiae; Mikus MD et al.; We constructed strains of Saccharomyces cerevisiae that contained two different mutant alleles of either the leu2 gene or the ura3 gene . These repeated genes were located on chromosomes V and XII and the two leu2- alleles were located on chromosomes III and XII . Genetic interactions between the two mutant copies of a gene were detected by the generation of either Leu+ or Ura+ revertants . Both spontaneous and ultraviolet irradiation-induced revertants were examined . By genetic and physical analysis, we have shown that Leu+ or Ura+ revertants can arise by a variety of different genetic interactions . The most common type of genetic interaction is the nonreciprocal transfer of information from one repeat to the other . We also detected reciprocal recombination between repeated genes, resulting in reciprocally translocated chromosomes.

Genetics, 1982 Jul-Aug, 101(3-4), 345 - 67
Frameshift suppression in Saccharomyces cerevisiae . IV . New suppressors among spontaneous co-revertants of the Group II his4-206 and leu 2-3 frameshift mutations; Gaber RF et al.; ICR-induced frameshift mutations at the his4 locus in Saccharomyces cerevisiae have been classified into several groups on the basis of their reversion and suppression properties . One group of externally suppressible his4 mutations, designated Group II, have been shown to contain +1 G:C insertions in glycine codons and are suppressed by any one of five suppressor mutations described previously (SUF1, SUF3, SUF4, SUF5, and SUF6) . The suppressor genes are believed to encode glycine tRNAs containing four base anticodons.--An analysis of spontaneous co-revertants of the Group II frameshift mutation his4-206 and leu2-3 has revealed the existence of eleven new Group II-specific suppressor genes (SUF15 through SUF25) . The locations of the new suppressor loci on the yeast genetic map have been determined.--By comparing the ability or inability of Group II-specific suppressors mapping at 16 different Group II his4 mutations, two subclasses of suppressors have been defined . One subclass suppresses his4-38 and his4-519, which contain the altered four base mRNA codons 5'-GGGU-3' and 5'-GGGG-3', respectively . The other subclass suppresses his4-38, but fails to suppress his4-519 . The mechanism of tRNA-mediated frameshift suppression and the molecular basis for this division of the suppressors into two subclasses is discussed.

Mutat Res, 1982 Jul-Aug, 102(1), 59 - 69
Genetic activity in Saccharomyces cerevisiae and thin-layer chromatographic comparisons of medical grades of pyrvinium pamoate and monopyrvinium salts; Mehta RD et al.; The pamoate, chloride, and iodide salts of pyrvinium, a cyanine dye with anthelmintic properties, were studied in a diploid mitotic recombination and gene conversion assay system (strain D5 of Saccharomyces cerevisiae) and a haploid yeast reversion assay (strain XV185-14C) . With the use of a thin-layer chromatographic (TLC) detection technique, samples of pyrvinium pamoate from several sources were found to contain different numbers and quantities of impurities . All samples of pyrvinium pamoate and the monopyrvinium salts were recombinogenic in strain D5 and mutagenic in strain XV185-14C; the degree of genetic activity varied among the tested medical grades of pyrvinium pamoate . Monopotassium pamoate was found to be genetically inactive in both strains . Light-catalyzed degradation did not enhance the genetic activity of pyrvinium in either of the yeast strains; the degraded samples were not mutagenic.

J Bacteriol, 1982 Jul, 151(1), 334 - 42
Effect of inositol starvation on the in vitro syntheses of mannan and N-acetylglucosaminylpyrophosphoryldolichol in Saccharomyces cerevisiae; Hanson BA et al.; An early consequence of starvation for inositol in yeast is inhibition of synthesis of the major cell wall components mannan and glucan . In looking for the mechanism of this inhibition, we found that the activity of the enzyme catalyzing the synthesis of N-acetylglucosaminylpyrophosphoryldolichol was diminished in particular membrane preparations from cells starved for inositol . This loss of reactivity was observed under a variety of in vitro assay conditions and could be restored by the addition of phosphatidylinositol but not by other phosphoinositol-containing sphingolipids known to occur in yeast . When assayed in the presence of high concentrations of Triton X-100, enzyme preparations from both control and inositol-starved cells required phosphatidylinositol for maximal activity . Since this enzyme catalyzed an early step in the synthesis of mannan that is N-linked to protein, a reasonable hypothesis is that inhibition of mannan synthesis in inositol-starved cells results from the depletion of the necessary cofactor phosphatidylinositol.

Mol Cell Biol, 1982 Jul, 2(7), 800 - 4
Post-translational processing of urea amidolyase in Saccharomyces cerevisiae; Sumrada RA et al.; Urea amidolyase catalyzes the two reactions (urea carboxylase and a allophanate hydrolase) associated with urea degradation in Saccharomyces cerevisiae . Past work has shown that both reactions are catalyzed by a 204-kilodalton, multifunctional protein . In view of these observations, it was surprising to find that on induction at 22 degrees C, approximately 2 to 6 min elapsed between the appearance of allophanate hydrolase and urea carboxylase activities . In search of an explanation for this apparent paradox, we determined whether or not a detectable period of time elapsed between the appearance of allophanate hydrolase activity and activation of the urea carboxylase domain by the addition of biotin . We found that a significant portion of the protein produced immediately after the onset of induction lacked the prosthetic group . A steady-state level of biotin-free enzyme was reached 16 min after induction and persisted indefinitely thereafter . These data are consistent with the suggestion that sequential induction of allophanate hydrolase and urea carboxylase activities results from the time required to covalently bind biotin to the latter domain of the protein.

J Bacteriol, 1982 Jul, 151(1), 29 - 35
Nitrogen catabolite repression in a glutamate auxotroph of Saccharomyces cerevisiae; Kang L et al.; The biosynthesis of asparaginase II in Saccharomyces cerevisiae is subject to nitrogen catabolite repression . In the present study we examined the physiological effects of glutamate auxotrophy on cellular metabolism and on the nitrogen catabolite repression of asparaginase II . Glutamate auxotrophic cells, incubated without a glutamate supplement, had a diminished internal pool of alpha-ketoglutarate and a concomitant inability to equilibrate ammonium ion with alpha-amino nitrogen . In the glutamate auxotroph, asparaginase II biosynthesis exhibited a decreased sensitivity to nitrogen catabolite repression by ammonium ion but normal sensitivity to nitrogen catabolite repression by all amino acids tested.

Biochim Biophys Acta, 1982 Jun 11, 711(3), 403 - 10
Purification and properties of a phospholipid acyl hydrolase from plasma membranes of Saccharomyces cerevisiae; Witt W et al.; The properties of a phospholipid acyl hydrolase bound to yeast plasma membranes are described in detail . The enzyme is capable of splitting all phospholipids which can be extracted from yeast cells . The specific activity with lysophosphatidylcholine as substrate was much higher than with phosphatidylcholine . With dipalmitoyl phosphatidylcholine as substrate a broad pH optimum was measured between pH 3.0 and 4.5 . The membrane-bound enzyme was activated strongly by the anionic detergents SDS, deoxycholate and, to a lesser extent, by cholate . The uncharged detergent Triton X-100 and the zwitterionic detergent SB12 exerted an only slightly activating effect . KCl, NaCl, MgCl2, and CaCl2 were inhibitory in the presence of glycine/acetic acid buffer at pH 4.0 . THe enzyme was solubilized by cholate or by SB12 in an active form from the plasma membrane and purified by acetone and (NH4)2SO4 precipitation or gel filtration with Sephadex G-200 . THe phospholipid acyl hydrolase was identified as a glycoprotein with an apparent molecular weight of 145,000 by SDS slab gel electrophoresis.

J Biol Chem, 1982 Jun 10, 257(11), 6268 - 74
Assembly of the mitochondrial membrane system . Processing of the apocytochrome b precursor RNAs in Saccharomyces cerevisiae D273-10B; Bonitz SG et al.; The DNA sequence of the apocytochrome b gene in Saccharomyces cerevisiae D273-10B contains two intervening sequences (Nobrega, F . G., and Tzagoloff, A . (1980) J . Biol . Chem . 255, 9828-9837) . The exon-intron boundaries of the gene have been determined in this study from the sequence of the DNA which was copied from the mRNA . A protein of 385 amino acid residues is predicted from the 1155-nucleotide long coding regions . Northern blot analysis of total mitochondrial RNA, probed with restriction fragments from both exon and intron regions of the gene, reveals a 4.3-kilobase (kb) transcript containing both introns and two partially spliced intermediates, one (2.9 kb) lacking the first intron and the other (3.6 kb) lacking the second intron . The most abundant transcript (2.1 kb) hybridizes only to exon probes and is presumed to the fully spliced mRNA . S1 nuclease mapping of the purified mRNA indicates existence of two separate RNAs with identical 3' termini but differing by approximately 217 nucleotides at their 5' ends . The larger transcript has a 950-nucleotide nontranslated leader . Analyses of the RNA species present in various rho- and mit- mutants indicate that: 1) exon mutants process both introns, albeit not as efficiently as wild type, 2) intron mutants blocked in the excision of the first or second intron are capable of processing the alternate intron, suggesting a non-obligatory order of excision of the two intervening sequences, and 3) excision of the second intron occurs in rho- mutants and therefore does not require a mitochondrial translation product.

J Biol Chem, 1982 Jun 10, 257(11), 6581 - 7
Characterization of transcripts from the Var1 region on mitochondrial DNA of Saccharomyces cerevisiae; Farrelly F et al.; We have identified transcripts with sequence homology to the var1 region on yeast mtDNA . In wild type, and in cytoplasmic petite strains retaining the var1 region, we detect four RNA species, 19 S, 16 S, 14 S, and 13 S, which hybridize to var1 DNA probes . The 16 S RNA is by far the most abundant of these RNAs in wild type cells . We also observe hybridization of the 15 S rRNA and a 10 S RNA species to var1 DNA probes . This hybridization is most likely due to the presence in these RNAs of a GC-rich cluster which has near perfect sequence homology to a GC-rich cluster in the var1 region . We find that the 16 S RNA, estimated to be 2000 to 2100 nucleotides long, varies in size in direct correspondence with the size of var1 polypeptide; it is about 100 nucleotides longer in strains with the var1 (44,000) allele than in strains with var1 (40,000) . The amount of the 16 S RNA also varies in correspondence with the amount of var1 polypeptide made; it is increased in an oxi-3 mit- strain (CAD245) which makes about 10 times more var1 protein than wild type, and is barely detected in PZ200, a strain with very low levels of mitochondrial protein synthesis harboring two mutations within var1 . We have purified the 16 S RNA following chromatography over oligo(dT) cellulose columns . When the RNA is end-labeled and hybridized to a HincII digest of wild type mtDNA, we observe hybridization only to the fragment containing the entire var1 region, HincII fragment 10.

Biochemistry, 1982 Jun 8, 21(12), 2957 - 63
In vivo phosphorus-31 nuclear magnetic resonance saturation transfer studies of adenosinetriphosphatase kinetics in Saccharomyces cerevisiae; Alger JR et al.; Phosphorus-31 saturation transfer NMR techniques have been employed to measure the unidirectional Pi consumption rate by respiration competent suspensions of the yeast Saccharomyces cerevisiae while the levels of ATP, ADP, and Pi are constant . These experiments are performed by saturating the ATP gamma phosphate resonance and observing the changes in the Pi resonance intensity while the yeast are respiring on endogenous substrates . The unidirectional Pi consumption rate is 3.5 +/- mumol s-1 (g of wet cells)-1 . The rate is reduced 10-fold upon addition of oligomycin (80 micrograms/ML), suggesting that at least 90% of the Pi consumption activity is due to the mitochondrial F1-F0 ATPase . We have not been able to conclusively assign the remaining 10% . When the yeast are glycolyzing anaerobically, the unidirectional Pi consumption rate was 1.0 +/- 0.2 mumol s-1 (g of wet cells)-1 . At most, 80% of this is due to Pi consumption by the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase leaving a residual activity of at least 0.2 mumol s-1 (g of wet cells)-1 . Thus the activity in the oligomycin-inhibited cells under respiratory conditions and the nonglycolytic activity in anaerobic cells are equal to within the experimental errors . Furthermore the unidirectional rate of Pi consumption during anaerobic glycolysis is insensitive to oligomycin . These data suggest that the mitochondrial adenosinetriphosphatase is not turning over during anaerobic glycolysis . Possible explanations for this inhibition are discussed.

Biokhimiia, 1982 Jun, 47(6), 962 - 70
{Inhibiting effect of concanavalin A on certain biosynthetic processes in spheroplasts of the yeast Saccharomyces cerevisiae}; Riazanova LP et al.; The effect of concanavalin A on biosynthesis of nucleic acids, proteins, structural polysaccharides and glycoproteins of the yeast cell membrane and of enzymes having different localization in the cell as well as on other processes occurring in spheroplasts of the yeasts Saccharomyces cerevisiae IBPhM-350 and CCY 21-4-13 were studied . In both yeast strains lectin strongly inhibited total protein synthesis and produced a weaker inhibiting effect on DNA and RNA synthesis . This was accompanied by a decrease of the activity of the majority of already known enzymes (acid phosphatase, invertase, alpha-glucosidase, polyphosphatase, pyrophosphatase, ATPase) and glucose consumption . In addition concanavalin A inhibited the synthesis of structural components of the yeast cell membrane, i.e . mannane and glucane . The data obtained suggest that lectin (50 microgram/ml or higher) has a toxic effect on yeast spheroplasts (or protoplasts).

Arch Microbiol, 1982 Jun, 131(4), 298 - 301
Localization of trehalase in vacuoles and of trehalose in the cytosol of yeast (Saccharomyces cerevisiae); Keller F et al.; Protoplasts of Saccharomyces cerevisiae synthesized and degraded trehalose when they were incubated in a medium containing traces of glucose and acetate . Such protoplasts were gently lyzed by the polybase method and a particulate and soluble fraction was prepared . Trehalose was found in the soluble fraction and the trehalase activity mostly in the particulate fraction which also contained the vacuoles besides other cell organelles . Upon purification of the vacuoles, by density gradient centrifugation, the specific activity of trehalase increased parallel to the specific content of vacuolar markers . This indicates that trehalose is located in the cytosol and trehalase in the vacuole . It is suggested that trehalose, in addition to its role as a reserve may also function as a protective agent to maintain the cytosolic structure under conditions of stress.

Proc Natl Acad Sci U S A, 1982 Jun, 79(11), 3565 - 9
Nucleotide sequence of the SUF2 frameshift suppressor gene of Saccharomyces cerevisiae; Cummins CM et al.; To elucidate the molecular mechanism of frameshift suppression by the SUF2 gene of yeast, the sequences of DNA fragments carrying the SUF2-1 and suf2+ alleles of the gene and surrounding regions have been determined . Comparison of the suppressor and wild-type sequences indicates that the SUF2 gene product is a proline tRNA . Disregarding possible base modifications, we find that the wild-type suf2+ anticodon of the tRNA inferred from the DNA sequence is 3'-GGA-5' . The SUF2-1 mutation represents the insertion of a G-C base pair at a position in the gene that corresponds to the anticodon loop of the tRNA . Replacement of the wild-type suf2+ anticodon by a 3'-GGGA-5' fourbase anticodon enables the SUF2-1 tRNA to suppress the 5'-CCCU-3' four-base codons generated as the result of the his4-712 and his4-713 frameshift mutations . This nontriplet codon-anticodon interaction restores the correct reading frame and allows synthesis of a functional his4 protein.

J Bioenerg Biomembr, 1982 Jun, 14(3), 191 - 205
Transmembrane ferricyanide reduction by cells of the yeast Saccharomyces cerevisiae; Crane FL et al.; Both respiratory-competent and respiratory-deficient yeast cells reduce external ferricyanide . The reduction is stimulated by ethanol and inhibited by the alcohol dehydrogenase inhibitor, pyrazole . The reduction of ferricyanide is not inhibited by inhibitors of mitochondrial or microsomal ferricyanide reduction . Cells in exponential-phase growth show a much higher rate of ferricyanide reduction . The reduction of ferricyanide is accompanied by increased release of protons by the yeast cells . We propose that the ferricyanide reduction is carried out by a transmembrane NADH dehydrogenase.

J Bioenerg Biomembr, 1982 Jun, 14(3), 159 - 69
The reconstitution of oxidative phosphorylation in mitochondria isolated from a ubiquinone-deficient mutant of Saccharomyces cerevisiae; De Santis A et al.; Mitochondria, isolated from the ubiquinone-deficient nuclear mutant of Saccharomyces cerevisiae E3-24, are practically unable to oxidize exogenous substrates . Respiratory activity, coupled to ATP synthesis, can, however, be reconstituted by the simple addition of ethanolic solutions of ubiquinones . A minimal length of the isoprenoid side chain (greater than or equal to 3) was required for the restoration . Saturation of the reconstitution required a large amount of exogeneous ubiquinone, in excess over the normal content present in the mitochondria of the wild type strain . A similar pattern of reconstituted activities could be also obtained using sonicated inverted particles . Mitochondria and sonicated particles are also able to carry out a dye-mediated electron flow coupled to ATP synthesis in the absence of added ubiquinone, using ascorbate or succinate as electron donor . This demonstrates that the energy conserving mechanism at the third coupling site of the respiratory chain is fully independent of the presence of the large mobile pool of ubiquinone in the membrane.

J Gen Microbiol, 1982 Jun, 128(Pt 6), 1309 - 17
Genetic block of outer plaque morphogenesis at the second meiotic division in an hfd1-1 mutant of Saccharomyces cerevisiae; Okamoto S et al.; An hfd1-1 mutant of Saccharomyces cerevisiae, SOS4, characterized by predominant production of two-spored asci at 29 degrees C, undergoes normal meiotic nuclear divisions and produces four haploid nuclei, but only two non-sister nuclei among them are incorporated into mature ascospores . Spindle pole bodies and prospore wall membranes at the second meiotic division at 29 degrees C were observed in this mutant by electron microscopy . The spindle pole body at one pole of each spindle had a normal outer plaque which was larger than the inner plaque . At the other pole, the outer plaque was entirely absent and a normal prospore wall membrane was not detected . It was concluded that at 29 degrees C the hfd1-1 mutation blocks the morphogenesis of outer plaques and prospore wall membranes at the two non-sister poles in the second meiotic division, and consequently only non-sister nuclei in the resulting meiotic cell are incorporated into ascospores.

Can J Biochem, 1982 Jun, 60(6), 659 - 67
Electroimmunochemical analysis of plasma membrane vesicles from Saccharomyces cerevisiae; Gerlach JH et al.; Plasma membrane vesicles of Saccharomyces cerevisiae were extracted with 1% (w/v) Triton X-100 and the solubilized proteins examined by crossed immunoelectrophoresis using rabbit antibodies against the vesicles . Solubilization was shown to be nonselective and 23 immunoprecipitates were observed reproducibly . Four glycoproteins were identified by interaction with concanavalin A and lentil lectin, either immobilized on agarose beads in an intermediate gel or incorporated in the free form in the first dimension gel . One glycoprotein was stainable by the periodic acid--Schiff procedure . None of the glycoproteins had their origin in the cell wall . Five amphiphilic proteins were identified on the basis of charge-shift and hydrophobic interaction crossed immunoelectrophoresis as well as {14C}Triton X-100 and Sudan black B binding . Three of the amphiphilic proteins were also glycoproteins . Based on the carbohydrate content and amphiphilic properties of the proteins, purification schemes using concanavalin A-Sepharose and phenyl-Sepharose were proposed . Trial separations using 1-mL columns were monitored by fused rocket and crossed immunoelectrophoresis.

J Biol Chem, 1982 May 25, 257(10), 5568 - 75
Biosynthetic control of the natural abundance of carbon 13 at specific positions within fatty acids in Saccharomyces cerevisiae . Isotopic fractionation in lipid synthesis as evidence for peroxisomal regulation; Monson KD et al.; Measurements of the natural abundance of 13C at C-1, C-9, and C-10 in fatty acids synthesized by Saccharomyces cerevisiae grown aerobically at 30 degrees C show that alkyl chain positions derived from the carboxyl group of the acetate precursor must be enriched in 13C by 2.5 +/- 0.6 parts per thousand while those derived from the methyl group in acetate must be depleted in 13C by an equal amount . Selective depletions of 13C observed at the C-9 and C-10 positions of palmitoleate and oleate require that (i) the carbon kinetic isotope effect associated with the action of desaturase at C-9 must be between 1.2 and 1.6% in vivo, (ii) at C-10 the effect must be between 0.9 and 1.3%, and (iii) less than 20% of the C18 carbon skeletons synthesized are preserved within the cell, the remainder apparently being degraded . It is shown that the novo synthesis (i.e . by fatty acid synthetase) is responsible for the production of more than 95% of the supply of 18-carbon acyl groups, the remainder being provided by all other elongation pathways . In an ancillary study designed to test the accuracy and generality of these results, it was observed that still larger specific depletions occurred at olefinic carbon position in fatty acids from soybeans, thus suggesting that the degradation of substantial quantities of C18 carbon skeletons may be a widespread feature of fatty acid metabolism in eukaryotes . It is suggested that the required degradation is associated with the action of peroxisomes.

J Biol Chem, 1982 May 25, 257(10), 5730 - 7
Effects of mannoprotein mutations on Saccharomyces cerevisiae core oligosaccharide structure; Cohen RE et al.; By the combined actions of an endo-alpha-1 leads to 6-mannanase and an endo-beta-N-acetylglucosaminidase, the core oligosaccharides can be released from Saccharomyces cerevisiae X2180 mnn2 mannoproteins . The effects of various mannoprotein mutations were evaluated by structural comparison of these core oligosaccharides with those prepared from double mutant strains with the genotypes mnn1 mnn2, mnn2 mnn3, mnn2 mnn4, and mnn2 mnn5 . The results indicate that only the mnn1 lesion has a major effect on the mannoprotein core structure . Whereas the mnn2 mannoprotein yields a core composed of 6 fragments that differ in size from each other by single mannose units, only the two smallest species predominate in the mnn1 mnn2 preparation . This change is correlated with a loss of terminal alpha 1 leads to 3-mannosyl residues, an effect on the mnn1 lesion that is found also in the polysaccharide outer chain and hydroxyamino acid-linked mannooligosaccharides . The mnn3 and mnn5 mutations also had slight effects on the core size, but clear differences in linkage composition were not apparent . The results suggest that core oligosaccharides have an average composition of Man11GlcNAc, whereas Man9GlcNAc is the major oligosaccharide in strains containing the mnn1 defect . These values are 2 to 3 sugars less than those estimated previously (Nakajima, T., and Ballou, C . E . (1975) Biochem . Biophys . Res . Commun . 66, 870-879) . Detailed analysis of the major core oligosaccharide from the mnn1 mnn2 mutant revealed that the two mannoses in alpha 1 leads to 3 linkage to the backbone were adjacent to each other and that the oligosacccharide is nearly identical with one isolated from chinese hamster ovary cell membranes (Li, E., and Kornfeld, S . (1979) J . Biol . Chem . 254, 1600-1605) . This finding provides strong evidence for the evolutionary conservation of this structural feature of the high mannose core oligosaccharides.

Mol Cell Biol, 1982 May, 2(5), 571 - 77
A suppressor of temperature-sensitive rna mutations that affect mRNA metabolism in Saccharomyces cerevisiae; Pearson NJ et al.; We have isolated a dominant suppressor of rna mutation (SRN1) that relieves the temperature-sensitive inhibition of mRNA synthesis of ribosomal protein genes in the yeast Saccharomyces cerevisiae . The suppressor was selected for its ability to alleviate simultaneously the temperature-sensitive growth phenotypes of rna2 and rna6 . Several independently isolated suppressors appeared to be recessive lethal mutations . One suppressor, SRN1, was recovered as viable in haploid strains . SRN1 can suppress rna2, rna3, rna4, rna5, rna6, and rna8 singly or in pairs, although some combinations of rna mutations are less well suppressed than others . The suppressor allows strains with rna mutations to grow at 34 degrees C but is unable to suppress at 37 degrees C; however, SRN1 does not, by itself, prevent growth at 37 degrees C . In addition, SRN1 suppresses the rna1 mutation which affects general mRNA levels and also leads to the accumulation of precursor tRNA for those tRNAs that have intervening sequences . SRN1 can suppress the rna1 mutation as well as the rna1 rna2 double mutation at 34 degrees C . The suppressor does not affect the temperature-sensitive growth of two unrelated temperature-sensitive mutations, cdc4 and cdc7.

Mol Cell Biol, 1982 May, 2(5), 490 - 7
Endogenous read-through of a UGA termination codon in a Saccharomyces cerevisiae cell-free system: evidence for involvement of both a mitochondrial and a nuclear tRNA; Tuite MF et al.; Globin mRNA, translated in a Saccharomyces cerevisiae cell-free protein synthesizing system prepared from a {psi+ rho+} strain, primarily directed the synthesis of alpha- and beta-globin . A third globin mRNA-specific polypeptide was also synthesized, representing approximately 10% of the total translation products . This polypeptide (beta') was synthesized by translational read-through of the beta- globin mRNA UGA terminator and was mediated primarily by an endogenous tRNA coded for by the mitochondria . This mitochondrial tRNA, when charged, could be preferentially bound, in high salt, to benzoylated DEAE-cellulose, a characteristic of a tRNATrp . The synthesis of beta- mediated by this mitochondrial tRNATrp was significantly reduced when the translation system was prepared from an isogenic {psi-} strain . Evidence for a nuclear-coded tRNA, also able to suppress the beta-globin mRNA UGA terminator in {psi+} but not {psi-} lysates, was also obtained . The presence of these endogenous UGA suppressor activities in the yeast cell-free system should allow successful in vitro translation of mitochondrial mRNAs.

Antonie Van Leeuwenhoek, 1982 May, 48(2), 145 - 57
Mutants of Saccharomyces cerevisiae cell division cycle defective in cytokinesis . Biosynthesis of the cell wall and morphology; Dominguez A et al.; The four temperature-sensitive mutants of Saccharomyces cerevisiae in the cell division cycle defective in cytokinesis (cdc, 3, 10, 11 and 12), have been analyzed with respect to the biosynthesis of the cell wall polymers . After 3 hours of incubation at the non-permissive temperature (37 degrees C) these strains stop growing . The synthesis of glucan, mannan and chitin (wall polymers) level off in a similar time, but glucan, mannan and chitin synthases remained active for at least 4 hours . If the mutants are analyzed by transmission and scanning electron microscopy different pictures emerge . Two of the mutants cdc 10 and cdc 12, after 3 hours of incubation at 37 degrees C present apparently normal cytoplasms and cell wall surfaces with multiple elongated buds . The other two mutants, cdc 3 and cdc 11, present a completely disarranged cytoplasmic content and damage at the level of the plasma membrane is evident . These and other observations, suggest that between the execution points of cdc 3 (0.27) and cdc 10 (0.58), essential processes in the assembly of cell membrane occur.

J Bacteriol, 1982 May, 150(2), 969 - 72
Evidence that alpha-isopropylmalate synthase of Saccharomyces cerevisiae is under the "general" control of amino acid biosynthesis; Hsu YP et al.; The specific activity and the immunoreactive amount of alpha-isopropylmalate synthase were more than three times above wild-type values in a Saccharomyces cerevisiae mutant (cdr1) with constitutively derepressed levels of enzymes known to be under the "general" control of amino acid biosynthesis . The specific activity was also higher in lysine- and arginine-leaky strains when these were grown under limiting conditions, and in wild-type cells grown in the presence of 5-methyltryptophan . A low specific activity was found in a mutant (ndr1) unable to derepress enzymes of the general control system . Neither isopropylmalate isomerase nor beta-isopropylmalate dehydrogenase responded to general control signals.

J Bacteriol, 1982 May, 150(2), 890 - 9
Repression and induction of flocculation interactions in Saccharomyces cerevisiae; Miki BL et al.; The biological control of flocculation interactions by factors related to growth under different conditions of aeration was documented with a new assay for flocculence . The degree of flocculence expressed in a genetically defined Saccharomyces cerevisiae strain (FLO1/FLO1 ade1/ade1) remained constant during aerobic growth but varied with aeration . Flocculence was repressed in anaerobically growing cells but was induced in stationary cells or cells returned to aerobic growth . Repression was correlated with the selective inactivation of cell surface lectin-like components . The changes in flocculence were accompanied by changes in 16 extractable proteins separated by electrophoresis; however, a clear correlation between specific protein bands and flocculence could not be established . The study clearly demonstrated that the phenotypic expression of FLO1 could be reproducibly manipulated for experimental purposes by aeration alone.

J Bacteriol, 1982 May, 150(2), 878 - 89
Possible mechanism for flocculation interactions governed by gene FLO1 in Saccharomyces cerevisiae; Miki BL et al.; A model is proposed for the mechanism of flocculation interactions in yeasts in which flocculent cells have a recognition factor which attaches to alpha-mannan sites on other cells . This factor may be governed by the expression of the single, dominant gene FLO1 . Isogenic strains of Saccharomyces cerevisiae, differing only at FLO1 and the marker genes ade1 and trp1, were developed to examine the components involved in flocculene . Electron microscopy and concanavalin Aferritin labeling of aggregated cells showed that extensive and intense interactions between cell wall mannan layers mediated cell aggregation . The components of the mannan layer essential for flocculence were Ca2+ ions, alpha-mannan carbohydrates, and proteins . By studying the divalent cation dependence at various pH values and in the presence of competing monovalent cations, flocculation was found to be Ca2+ dependent; however, Mg2+ and Mn2+ ions substituted for Ca2+ under certain conditions . Reversible inhibition of flocculation by concanavalin A and succinylated concanavalin A implicated alpha-branched mannan carbohydrates as one essential component which alone did not determine the strain specificity of flocculence, since nonflocculent strains interacted with and competed for binding sites on flocculent cells . FLO1 may govern the expression of a proteinaceous, lectin-like activity, firmly associated with the cell walls of flocculent cells, which bind to the alpha-mannan carbohydrates of adjoining cells . It was selectively and irreversibly inhibited by proteolysis and reduction of disulfide bonds . The potential of this system as a model for the genetic and biochemical control of cell-cell interactions is discussed.

J Bacteriol, 1982 May, 150(2), 545 - 51
Co-curing of plasmids affecting killer double-stranded RNAs of Saccharomyces cerevisiae: {HOK}, {NEX}, and the abundance of L are related and further evidence that M1 requires L; Sommer SS et al.; We describe two sets of plasmid-plasmid interactions in the yeast Saccharomyces cerevisiae . {HOK}, {EXL}, {NEX}, and {KIL-k1} are genetically defined plasmids, and M1 and L are biochemically defined double-stranded RNA plasmids . We show that (i) {HOK}, {NEX}, and the abundance of L are related, and (ii) under submaximal curing conditions, all colonies retaining M1 also retain L . There are three pieces of evidence that either {NEX} required {HOK} for replication or {NEX} and {HOK} are on the same plasmid . The evidence is as follows . (i) The great majority of strains containing {HOK} also contain {NEX} . However, two {HOK} {NEX-o} strains do exist . (ii) Growth at 39 degrees C or growth at 34 degrees C with 3% ethanol or 2-propanol cures {HOK} and {NEX} . In a {HOK} {NEX} strain, the two plasmids are always co-cured . (iii) {HOK} and {NEX} are both maintained in mak4, mak6, and mak27 strains (mak = maintenance of {KIL-k1}), but not in mak3, mak10, and pet18 strains . Strains containing {HOK} and {NEX} have about fourfold more L double-stranded RNA than their isochromosomal, cured derivatives . In addition, a cytoductant which has acquired {HOK} and {NEX} has fourfold more L than its parent . These results are consistent with either {HOK} being a form of L or {HOK} increasing the copy number of L . Using a K1 killer strain in which L, as well as M1, could be cured by growth at 38 degrees C, we examined the distribution of loss of M1 and L under conditions giving 98% M-o colonies and at least 50% L-o colonies . No M1L-o colonies were observed, supporting the previous suggestion by others that M1 requires L.

Nucleic Acids Res, 1982 Apr 24, 10(8), 2625 - 37
Conservation of high efficiency promoter sequences in Saccharomyces cerevisiae; Dobson MJ et al.; The position of the yeast phosphoglycerate kinase (PGK) gene has been mapped on a 2.95kb Hind III fragment . We have determined the nucleotide sequence of the 5' flanking region and compared this sequence with those from 16 other yeast genes . PGK, like all other yeast genes has an adenine residue at position -3 . It has two possible TATA boxes at positions -114 and -152 and a CAAT box at -129 . In addition we have defined a structure at position -63 to -39 that is common to all yeast genes that encode an abundant RNA . This structure is a CT-rich block followed, about 10 nucleotides later, by the sequence CAAG.

Biochim Biophys Acta, 1982 Apr 23, 687(1), 57 - 62
The isolation of Saccharomyces cerevisiae nuclear membranes with nuclease and high-salt treatment; Mann K et al.; Saccharomyces cerevisiae nuclear membranes were prepared from isolated nuclei by digesting chromatin with deoxyribonuclease and ribonuclease, washing of residual nuclei with 0.5 M MgCl2, and discontinuous gradient centrifugation in buffered Ficoll solutions . Electron microscopic examination of the preparations showed single membrane and double membrane vesicles and membrane sheets . Pores or residual pores were often visible . In double membrane profiles the two unit membranes were often separated by the remains of the perinuclear cistern . The nuclear membrane fragments contained 58% protein, 23.8% phospholipid, 6% sterols, 7.1% neutral acylglycerols, 4.8% RNA, and 0.3% DNA . The phospholipid content of the membrane preparations was influenced by a phospholipase activity with acidic pH optimum.

Mol Cell Biol, 1982 Apr, 2(4), 457 - 66
Influence of the nuclear gene tmp3 on the loss of mitochondrial genes in Saccharomyces cerevisiae; Zelikson R et al.; The Saccharomyces cerevisiae tmp3 mutant is deficient in the mitochondrial enzyme complex that participates in the formation of one-carbon-group-tetrahydrofolate coenzymes, serine transhydroxymethylase, dihydrofolate reductase, and thymidylate synthetase, thus leading to multiple nutritional requirements of dTMP, adenine, histidine, and methionine . The tmp3 mutant quickly loses its mitochondrial genome even when grown on fully supplemented medium or on a high concentration of 5-formyl tetrahydrofolate, which replaces all the four requirements . A study of the loss of the mitochondrial genome by following several mitochondrial genetic markers showed that there was a preferential specific loss of a large region of the mitochondrial genome, covering mit ts983, Er, Cr, and mit ts982 up to OrI, and retention of the region of Pr and mit tscs1297 . A kinetic study showed that there was a preferentially rapid loss of the region covering the mit+ alleles ts983 to tscs902 at the rate of 10% per generation.

Mol Cell Biol, 1982 Apr, 2(4), 450 - 6
Oligoadenylate is present in the mitochondrial RNA of Saccharomyces cerevisiae; Yuckenberg PD et al.; We examined Saccharomyces cerevisiae mitochondrial RNA for polyadenylate . Using hybridization to {3H}polyuridylate as the assay for adenylate sequences, we found adenylate-rich oligonucleotides approximately 8 residues long . Longer polyadenylate was not detected . Most of the adenylate-rich sequence is associated with the large mitochondrial rRNA . The remainder is associated with the 10-12S group of transcripts.

Mol Cell Biol, 1982 Apr, 2(4), 361 - 8
Size control models of Saccharomyces cerevisiae cell proliferation; Wheals AE; By using time-lapse photomicroscopy, the individual cycle times and sizes at bud emergence were measured for a population of saccharomyces cerevisiae cells growing exponentially under balanced growth conditions in a specially constructed filming slide . There was extensive variability in both parameters for daughter and parent cells . The data on 162 pairs of siblings were analyzed for agreement with the predictions of the transition probability hypothesis and the critical-size hypothesis of yeast cell proliferation and also with a model incorporating both of these hypotheses in tandem . None of the models accounted for all of the experimental data, but two models did give good agreement to all of the data . The wobbly tandem model proposes that cells need to attain a critical size, which is very variable, enabling them to enter a start state from which they exit with first order kinetics . The sloppy size control model suggests that cells have an increasing probability per unit time of traversing start as they increase in size, reaching a high plateau value which is less than one . Both models predict that the kinetics of entry into the cell division sequence will strongly depend on variability in birth size and thus will be quite different for daughters and parents of the asymmetrically dividing yeast cells . Mechanisms underlying these models are discussed.

Eur J Biochem, 1982 Apr, 123(3), 571 - 6
Occurrence of a catabolic L-serine (L-threonine) deaminase in Saccharomyces cerevisiae; Ramos F et al.; Saccharomyces cerevisiae mutants lacking the anabolic L-threonine deaminase, the ilv1- mutants, have been found to exhibit a normal ability to grow, without auxotrophy towards isoleucine, on L-threonine of L-serine as only nitrogen nutrient . Starting from a strain carrying a ilv1- mutation, a new mutation affecting the ability to utilize L-threonine as nitrogen source was selected . This mutation, which also impairs the ability to utilize L-serine, has been denominated cha-, for 'catabolism of hydroxyamino acids' and was found to result in the lack of a catabolic L-serine (L-threonine) deaminase . This enzyme which, unlike the anabolic threonine deaminase, is more active towards serine than towards threonine, differs from the latter enzyme by a number of biochemical and regulatory properties . Whereas the anabolic enzyme is an allosteric enzyme sensitive to feedback inhibition by isoleucine, the catabolic enzyme exhibits Michaelian kinetics: no control of its activity has been detected . Its synthesis is induced by L-serine and L-threonine . These two enzymes, which thus can be easily differentiated by means of their regulations, display a limited ability to compensate for one another's absence and appear to play clearly distinct roles under normal physiological conditions.

Mol Cell Biol, 1982 Apr, 2(4), 437 - 42
Isolation of the thymidylate synthetase gene (TMP1) by complementation in Saccharomyces cerevisiae; Taylor GR et al.; The structural gene (TMP1) for yeast thymidylate synthetase (thymidylate synthase; EC 2.1.1.45) was isolated from a chimeric plasmid bank by genetic complementation in Saccharomyces cerevisiae . Retransformation of the dTMP auxotroph GY712 and a temperature-sensitive mutant (cdc21) with purified plasmid (pTL1) yielded Tmp+ transformants at high frequency . In addition, the plasmid was tested for the ability to complement a bacterial thyA mutant that lacks functional thymidylate synthetase . Although it was not possible to select Thy+ transformants directly, it was found that all pTL1 transformants were phenotypically Thy+ after several generations of growth in nonselective conditions . Thus, yeast thymidylate synthetase is biologically active in Escherichia coli . Thymidylate synthetase was assayed in yeast cell lysates by high-pressure liquid chromatography to monitor the conversion of {6-3H}dUMP to {6-3H}dTMP . In protein extracts from the thymidylate auxotroph (tmp1-6) enzymatic conversion of dUMP to dTMP was barely detectable . Lysates of pTL1 transformants of this strain, however, had thymidylate synthetase activity that was comparable to that of the wild-type strain.

Mol Cell Biol, 1982 Apr, 2(4), 412 - 25
Preliminary characterization of the transcriptional and translational products of the Saccharomyces cerevisiae cell division cycle gene CDC28; Reed SI et al.; The CDC28 gene was subcloned from a plasmid containing a 6.5-kilobase-pair segment of Saccharomyces cerevisiae DNA YRp7(CDC28-3) by partial digestion with Sau3A and insertion of the resulting fragments into the BamHI sites of YRp7 and pRC1 . Recombinant plasmids were obtained containing inserts of 4.4 and 3.1 kilobase pairs which were capable of complementing a cdc28(ts) mutation . R-loop analysis indicated that each yeast insert contained two RNA coding regions of about 0.8 and 1.0 kilobase pairs, respectively . In vitro mutagenesis experiments suggested that the smaller coding region corresponded to the CDC28 gene . When cellular polyadenylic acid-containing RNA, separated by agarose gel electrophoresis after denaturation with glyoxal and transferred to nitrocellulose membrane, was reacted with labeled DNA from the smaller coding region, and RNA species of about 1 kilobase in length was detected . Presumably, the discrepancy in size between the R-loop and electrophoretic determinations is due to a segment of polyadenylic acid which is excluded from the R-loops . By using hybridization of the histone H2B mRNAs to an appropriate probe as a previously determined standards, it was possible to estimate the number of CDC28 mRNA copies per haploid cell as between 6 and 12 molecules . Hybrid release translation performed on the CDC29 mRNA directed the synthesis of a polypeptide of 27,000 daltons, as determined by polyacrylamide gel electrophoresis in sodium dodecyl sulfate . This polypeptide was not synthesized when mRNA prepared from a cdc28 nonsense mutant was translated in a parallel fashion . However, if the RNA from a cell containing the CDC28 gene on a plasmid maintained at a high copy number was translated, the amount of in vitro product was amplified fivefold.

Proc Natl Acad Sci U S A, 1982 Apr, 79(7), 2157 - 61
Acid phosphatase polypeptides in Saccharomyces cerevisiae are encoded by a differentially regulated multigene family; Rogers DT et al.; Two clones from a lambda phage collection containing yeast genes regulated by inorganic phosphate were shown by low-stringency hybridization to select three mRNAs that direct the in vitro synthesis of repressible acid phosphatase (EC 3.1.3.2) polypeptides p60, p58, and p56 . By higher stringency hybridization one yeast fragment {8 kilobases (kb)} selects p60 mRNA and the other (5 kb) selects p56 mRNA . These EcoRI digestion fragments were subcloned in yeast transformation vectors and hybridization selection assignments were confirmed by measuring enzyme and mRNA levels in transformants . Enzyme and mRNA levels in (8-kb) high copy number transformants grown in high inorganic phosphate medium revealed a hitherto undetected acid phosphatase protein, P57, which is believed to correspond to the constitutive enzyme encoded by PHO3 . The identify of the 8-kb fragment purported to contain the PHO5/PHO3 genes was confirmed by genetic mapping of an integrated copy of this fragment . The site of integration of the 5-kb fragment was demonstrated to be unlinked to the PHO5/PHO3 genes.

J Biol Chem, 1982 Mar 25, 257(6), 3218 - 24
Evidence for translated intervening sequences in the mitochondrial genome of Saccharomyces cerevisiae; Hanson DK et al.; In yeast, the mitochondrial genes for subunit I of cytochrome oxidase (oxi3) and for apocytochrome b (cob) are known to be split . In some strains, the latter contains five intervening sequences, three of which coincide with clusters of mutational sites referred to in their order of transcription as the loci box3, 10, and 7, respectively . Mutations at the first of these result in the accumulation of novel, large polypeptides (apparent Mr congruent to 43,000) believed to originate from a fusion of sequences found in the NH2-terminal segment of apocytochrome b to others encoded in the intervening sequence itself . We now provide evidence for close similarities of at least a part of translated intron sequences between (a) mutants in box7 in "long" form and "short" form strains (which lack the first three introns including the one for the box3 locus); (b) mutants in a subset of box7 mutants and those in box3, and (c) between intron sequences in box7 and a sequence presumably encoded in oxi3 . These structural homologies have been analyzed and shown to be referable to sequence homologies in two proteins, one derived from the second intron (box3) in cob and the other from oxi3 . The accumulation in certain cob mutants of proteins and of a transcript containing a sequence specified by oxi3 provides additional strong evidence for the previously suggested regulation of oxi3 by the penultimate, box7-containing intron of cob.

Biochim Biophys Acta, 1982 Mar 8, 685(3), 329 - 39
The plasma membrane of Saccharomyces cerevisiae . Molecular structure and asymmetry; Santos E et al.; The molecular structure of the plasma membrane of the haploid strain Saccharomyces cerevisiae X-2180 1 A has been studied by means of sodium dodecyl sulfate polyacrylamide gel electrophoresis . Protein and glycoprotein components have been identified and their apparent Mr determined . A glycoprotein showing an apparent Mr of 27500 has been shown to be the main structural component . Treatment of the cells with cycloheximide prior to plasma membrane isolation resulted in a redistribution of the relative amounts of each protein band and a drastic reduction in the number of Schiff positive bands . It is postulated that treatment with this drug rids the plasma membrane of glycoprotein secretory components which are in the process of being secreted to the periplasmic space, thus allowing the study of the basic structural components of the organelle . The electrophoretic pattern of the internal membranes revealed close similarities with that of the plasma membrane and though two-dimensional electrophoresis might disclose greater differences, these similarities suggest a common origin for most of the components of both membranous systems . Finally, radioiodination techniques, have been used in studying the asymmetric disposition of some of the components of the plasma membrane . At least five polypeptides were identified as located to the outer layer of the plasma membrane and two more glycopeptides were shown to span across the bilayer.

Biochim Biophys Acta, 1982 Mar 4, 701(3), 334 - 8
Partial purification and comparison of some properties of L-serine sulfhydro-lyase of Saccharomyces cerevisiae; Yamagata S et al.; In order to ascertain the role of L-serine sulfhydro-lyase (L-serine hydro-lyase (adding homocysteine) EC 4.2.1.22) which also catalyzes sulfhydrylation of O-acetyl-L-serine (Yamagata, S . (1981) J . Bacteriol . 147, 688-690), the enzyme was partially purified from a wild-type strain and three cysteine auxotrophs of Saccharomyces cerevisiae, and the molecular and enzymatic properties of these preparations were compared . The results showed no significant difference in properties investigated, indicating that cysteine synthesis is exclusively performed in this organism through sulfhydrylation of O-acetyl-L-serine, catalyzed not by serine sulfhydro-lyase but by O-acetylserine . O-acetylhomoserine sulfhydro-lyase (Yamagata, S., Takeshima, K . and Naiki, N . (1974) J . Biochem . 75, 1221-1229) . Insensitivity of the former enzyme to L-methionine also supported this conclusion.

J Cell Biol, 1982 Mar, 92(3), 790 - 4
Nuclear localization of aspartate transcabamoylase in Saccharomyces cerevisiae; Nagy M et al.; The cytochemical technique using the in situ precipitation of orthophosphate ions liberated specifically by the aspartate carbamoyltransferase (ATCase) (EC 2.1.3.2) reaction indicated that in Saccharomyces cerevisiae this enzyme is confined to the nucleus . This observation is in accordance with the result reported by Bernhardt and Davis (1972), Proc . Natl . Acad . Sci . U . S . A . 69:1868-1872) on Neurospora crassa . The nuclear compartmentation was also observed in a mutant strain lacking proteinase B activity . This finding indicates that this proteinase is not involved in the nuclear accumulation of ATCase, and that the activity observed in the nucleus corresponds to the multifunctional form associated with the uracil path-specific carbamoylphosphate synthetase and sensitive to feedback inhibition by UTP . In a ura2 strain transformed by nonintegrated pFL1 plasmids bearing the URA2-ATCase activity encoding gene, the lead phosphate precipitate was observed predominantly in the cytoplasm . This finding enhances the reliability of the technique used by eliminating the possibility of an artifactual displacement of an originally cytoplasmic reaction product during the preparation of the material for electron microscopy . On the other hand, nuclei isolated under hypoosmotic conditions do not exhibit the ATCase activity that is recovered in the cytosolic fractions after differential centrifugation of the lysate in Percoll gradient . A release of the protein from the nuclei during the lysis step, consistent with its nucleoplasmic localization, is postulated.

J Biochem (Tokyo), 1982 Mar, 91(3), 911 - 21
Formation of dehydrosqualene catalyzed by squalene synthetase in Saccharomyces cerevisiae; Takatsuji H et al.; When microsomal fraction of Saccharomyces cerevisiae was incubated with farnesyl pyrophosphate or presqualene pyrophosphate in the presence of Mn2+, 12,13-cis-dehydrosqualene (DeH2Sq) and some related compounds were found to be formed . Incubation in the presence of NADPH gave rise to only squalene . By heat treatment of the microsomal fraction, the DeH2Sq- and squalene-forming activities were inactivated at approximately the same rate . The elution patterns of both activities upon Sephacryl S-200 chromatography of the enzyme solubilized from the microsomal fraction with taurodeoxycholate coincided completely . These results indicate that DeH2Sq formation in yeast is catalyzed by squalene synthetase . Divalent cation was essential for this reaction and Mn2+ was six times more effective than Mg2+ . DeH2Sq formation was also observed when microsomes of pig liver were used instead of yeast microsomal fraction, suggesting that this reaction is a ubiquitous one among the eucaryotes which are capable of synthesizing sterols . Based on these observations, the mechanisms of DeH2Sq and squalene formation are discussed.

Mol Biol (Mosk), 1982 Mar-Apr, 16(2), 363 - 8
{Isolation of intact supercoiled mitochondrial DNA from the yeast Saccharomyces cerevisiae}; Larionov VL et al.; Cells of Saccharomyces cerevisiae strain PG-60 were mechanically disrupted in the presence of ethidium bromide, and the total covalently closed circular DNA (ccDNA) was prepared by EthBr-CsCl equilibrium centrifugation of lysed 15 g pellet of the homogenate . Recentrifugation of ccDNA in equilibrium neutral CsCl density gradient resolved a discrete light peak (LccDNA) with the buoyant density equal to 1.684 g/cm3 to about 10% of total ccDNA . Electron microscopy has shown that LccDNA contained circular molecules with the contour length around 25 micrometer circles and their multimeres . Well characterized linear mtDNA was transcribed with E . coli RNA-polymerase and cRNA obtained was hybridized with LccDNA 2 mu DNA and total DNA from yeast petit mutant, lacking mtDNA . Hybridization experiments have shown that the transcript of mtDNA hybridized almost exclusively with LccDNA . We have concluded that 25 micrometer circles of the LccDNA represent intact molecules of yeast mitochondrial DNA.

Mol Cell Biol, 1982 Mar, 2(3), 221 - 32
Construction, replication, and chromatin structure of TRP1 RI circle, a multiple-copy synthetic plasmid derived from Saccharomyces cerevisiae chromosomal DNA; Zakian VA et al.; Transformation studies with Saccharomyces cerevisiae (bakers' yeast) have identified DNA sequences which permit extrachromosomal maintenance of recombinant DNA plasmids in transformed cells . It has been hypothesized that such sequences (called ARS for autonomously replicating sequence) serve as initiation sites for DNA replication in recombinant DNA plasmids and that they represent the normal sites for initiation of replication in yeast chromosomal DNA . We have constructed a novel plasmid called TRP1 R1 Circle which consists solely of 1,453 base pairs of yeast chromosomal DNA . TRP1 RI Circle contains both the TRP1 gene and a sequence called ARS1 . This plasmid is found in 100 to 200 copies per cell and is relatively stable during both mitotic and meiotic cell cycles . Replication of TRP1 RI Circle requires the products of the same genes (CDC28, CDC4, CDC7, and CDC8) required for replication of chromosomaL DNA . Like chromosomal DNA, its replication does not occur in cells arrested in the B1 phase of the cell cycle by incubation with the yeast pheromone alpha-factor . In addition, TRP1 RI Circle DNA is organized into nucleosomes whose size and spacing are indistinguishable from that of bulk yeast chromatin . These results indicate that TRP1 RI Circle has the replicative and structural properties expected for an origin of replication from yeast chromosomal DNA . Thus, this plasmid is a suitable model for further studies of yeast DNA replication in both cells and cell-free extracts.

Infect Immun, 1982 Mar, 35(3), 1157 - 61
Comparison of cytoplasmic extracts of eight Candida species and Saccharomyces cerevisiae; Greenfield RA et al.; Crossed-line immunoelectrophoresis of cytoplasmic extracts of eight Candida species and Saccharomyces cerevisiae demonstrated the presence of antigens reactive with a rabbit antiserum to a C . albicans extract in all species except C . glabrata . A previously defined major cytoplasmic antigen of C . albicans was also present in C . tropicalis and C . guilliermondii.

Biochim Biophys Acta, 1982 Feb 26, 696(2), 163 - 70
Purification and properties of two endonucleases specific for apurinic/apyrimidinic sites in DNA from Saccharomyces cerevisiae; Akhmedov AT et al.; Two distinct endonucleases from Saccharomyces cerevisiae, specific for apurinic/apyrimidinic sites (AP-endonucleases A and B), have been extensively purified and characterized . Both are free from unspecific and ultraviolet-specific endonucleases and exonucleases . The two enzymes are monomeric proteins of around 24000 daltons . Both are sensitive to ionic strength and most active in the presence of 150 and 100 mM NaCl for AP-endonucleases A and B, respectively . They are not absolutely dependent on divalent cations, since they are insensitive to EDTA, although AP-endonuclease A is activated by Ca2+ or Mg2+ and AP-endonuclease B by Mg2+ only . ATP inhibits the enzymes . AP-endonuclease A reacts optimally between pH 6 and 8, and AP-endonucleases B at pH 8 . AP-endonuclease A is more stable at 60 degree C (half-life of 17 min) than B (half-life of 4 min) . AP-endonuclease A is insensitive to N-ethylmaleimide or rho-chloromercuribenzoate . AP-endonuclease B is also insensitive to N-ethylmaleimide, but rho-chloromercuribenzoate inhibits its activity.

Nucleic Acids Res, 1982 Feb 11, 10(3), 889 - 902
Dihydrouridine-deficient tRNAs in Saccharomyces cerevisiae; Lo RY et al.; A mutation in Saccharomyces cerevisiae, designated mia, is responsible for the production of isoaccepting tRNA molecules with reduced extents of nucleoside modifications . The mia isoacceptors of tRNAPhe and one of the mutant isoacceptors of tRNATyr were highly purified for nucleoside composition analyses . The data indicate that the mutant isoacceptors are lacking some of the dihydrouridine moieties . This is consistent with our previous hypothesis that the mutant isoacceptors were accumulated due to a defect in a modification process {Lo, R.Y.C . and Bell, J.B . (1981) Current Genetics 3, 73-82) . Data from in vitro poly-U translation experiments also support the previous results, suggesting in vivo biological activity of these mutant tRNAs.

Nucleic Acids Res, 1982 Feb 11, 10(3), 1051 - 70
A novel species of double stranded RNA in mitochondria of Saccharomyces cerevisiae; Beilharz MW et al.; A double stranded RNA species has been detected in guanidine hydrochloride extracts of mitochondria from respiratory competent cells of Saccharomyces cerevisiae . This novel mitochondrial RNA, termed mtdsRNA, has been purified in a Cs2SO4 density gradient where it bands at a density of 1.58 g/ml . The mtdsRNA runs as a single slow moving band on agarose gels . Its double stranded RNA character was evidenced by its sensitivity to digestion by RNase III, but not by RNase H, or DNase I . Moreover the mtdsRNA hybridized to each separated strand of a petite mtDNA . It is concluded that mtdsRNA contains long transcripts derived from most regions of yeast mtDNA, because 1) its weight-average length as determined by electron microscopy was 4.5 micrometer (about 14 kb, or 20% of the wild type mtDNA genome), and 2) it hybridized to each of a series of eight petite mtDNA probes carrying sequences derived from widely different segments of mtDNA . It is proposed that prolonged transcription of both strands of yeast mtDNA can occur and that mtdsRNA arises from hybridization of these long complementary transcripts.

J Biol Chem, 1982 Feb 10, 257(3), 1392 - 7
Endochitinase, a mannan-associated enzyme from Saccharomyces cerevisiae; Correa JU et al.; A chitinase was extracted with digitonin from intact yeast cells and purified by adsorption-digestion on chitin . The purified enzyme liberates oligosaccharides of various sizes from chitin, thus behaving as an endochitinase . As found with other chitinases, the yeast enzyme is much more active on nascent chitin, i.e . the chitin formed in the same reaction mixture by the corresponding synthetase, than on preformed polysaccharide . The enzyme has a very low pH optimum, about pH 2.5, and is quite stable at pH 3 . Polyacrylamide gel electrophoresis of different preparations of purified chitinase revealed a variable number of protein bands, whose pattern often changed after storage of the enzyme . The distribution of activity in the gel matched that of the stainable material . Association of the chitinase with mannan is indicated by the following results: (a) coincidence in Coomassie blue-staining and periodate-Schiff-staining bands after disc gel electrophoresis; (b) adsorption of the activity on concanavalin A-Sepharose and elution with alpha-methylmanoside; (c) precipitation of the chitinase with appropriate antimannan sera . A carbohydrate content of approximately 18% was found in a trichloroacetic acid-precipitated sample of purified enzyme . Protein and mannan were not dissociated by boiling with sodium dodecyl sulfate, urea, and beta-mercaptoethanol . It could not, however, be conclusively established whether protein and carbohydrate are covalently linked, because the chitinase is resistant to endo-beta-N-acetyl-glucosaminidase.

Mol Cell Biol, 1982 Feb, 2(2), 171 - 8
Developmental regulation of a sporulation-specific enzyme activity in Saccharomyces cerevisiae; Clancy MJ et al.; An alpha-glucosidase activity (SAG) occurs in a/alpha Saccharomyces cerevisiae cells beginning at about 8 to 10 h after the initiation of sporulation . This enzyme is responsible for the rapid degradation of intracellular glycogen which follows the completion of meiosis in these cells . SAG differs from similar activities present in vegetative cells and appears to be a sporulation-specific enzyme . Cells arrested at various stages in sporulation (DNA replication, recombination, meiosis I, and meiosis II) were examined for SAG activity; the results show that SAG appearance depends on DNA synthesis and some recombination events but not on the meiotic divisions.

Mol Cell Biol, 1982 Feb, 2(2), 127 - 37
Identification of the genetic locus for the structural gene and a new regulatory gene for the synthesis of repressible alkaline phosphatase in Saccharomyces cerevisiae; Kaneko Y et al.; Two lines of evidence showed that the PHO8 gene encodes the structure of repressible, nonspecific alkaline phosphatase in Saccharomyces cerevisiae: (i) the enzyme produced by a temperature-sensitive pho8 mutant at the permissive temperature (25 degrees C) was more thermolabile than that of the wild-type strain, and (ii) the PHO8 gene showed a gene dosage effect on the enzyme activity . The pho8 locus has been mapped on chromosome IV, 8 centimorgans distal to rna3 . A new mutant carrying the pho9 gene was isolated which lacks repressible alkaline phosphatase, but has the normal phenotype for the synthesis of repressible acid phosphatase . The pho9 gene segregated independently of all known pho-regulatory genes and did not show the gene dosage effect on repressible alkaline phosphatase activity . The pho9/pho9 diploid hardly sporulated and showed no commitment to intragenic recombination when it was inoculated on sporulation medium . Hence the pho9 mutant has a phenotype similar to the pep4 mutant, which was isolated as a pleiotropic mutant with reduced levels of proteinases A and B and carboxypeptidase Y . An allelism test indicated that pho9 and pep4 are allelic.

Mol Cell Biol, 1982 Feb, 2(2), 117 - 26
Synthesis of specific identified, phosphorylated, heat shock, and heat stroke proteins through the cell cycle of Saccharomyces cerevisiae; Ludwig JR 2nd et al.; The methods of centrifugal elutriation, two-dimensional gel electrophoresis, and dual isotopic labeling were applied to the study and identification of a number of purified yeast proteins . The location of polypeptide spots corresponding to specific proteins was determined on two-dimensional gels . A dual-label method was used to determine the rates of synthesis through the cell cycle of the identified proteins as well as to confirm the results of previous studies from our laboratory on unidentified proteins . The identified proteins, and the more generally defined phosphorylated, heat shock, and heat stroke proteins were found to follow the general pattern of exponential increase in rate of synthesis through the cell cycle . In addition, colorimetric enzyme activity assays were used to examine the catabolic enzyme alpha-glucosidase (EC 3.2.1.20) . Both the activity and synthesis of alpha-glucosidase were found to be nonperiodic with respect to the cell cycle . These data contrast with earlier reports of periodicity, which employed induction and selection synchrony to study enzyme expression through the yeast cell cycle.

Eur J Cell Biol, 1982 Feb, 26(2), 219 - 27
Isolation and localization of plasma membrane-bound invertase in yeast (Saccharomyces cerevisiae); Maurer A et al.; Latex spheres of 60 nm diameter (synthesized by aqueous emulsion copolymerization of methacrylate derivatives according to {22}) were coated with bovine serum albumin (BSA) and concanavalin A . By virtue of their size and their high density (1.32-1.35 g/ml) they are well suited as scanning electron microscopy markers and as affinity density perturbation reagents . Yeast protoplasts could be labeled with these spheres and the amount of binding depended upon incubation time and temperature . Isolated and solubilized yeast plasma membranes were incubated with these spheres and by density gradient centrifugation the membrane glycoproteins could be separated from the other proteins by the method of affinity density perturbation . Since the yeast plasma membrane glycoproteins exhibit invertase activity {1, 19} the activity of the different fractions was either detected on gels by staining for invertase activity or measured in vitro and quantified; a 6 to 7fold purification of the enzyme was achieved . Protoplasts labeled with antibodies directed against these glycoproteins exhibited a distribution of ferritin marker molecules that was very similar to that of the intramembranous particles . Antibodies against extracellular invertase cross reacted with the plasma membrane of glycoproteins and showed the same distribution of markers as the antibodies against the glycoproteins . It can therefore be concluded that the yeast plasma membrane glycoproteins exhibit invertase activity and that they are associated with the intramembranous particles.

Eur J Biochem, 1982 Feb, 122(2), 369 - 74
Identity problems concerning subunits of the membrane factor of the mitochondrial ATPase of Saccharomyces cerevisiae; Somlo M et al.; As confirmed by fingerprint analysis, high-resolution acrylamide gel electrophoresis identifies subunits of the mitochondrial ATPase complex directly at the level of Coomassie-blue-stained yeast mitochondria . Using this technique, the following results were obtained . 1 . Three classes of subunits have been found in the ATPase complex . (a) The classical F1 peptides of cytoribosomal origin (57, 53 and 30.5 kDa) . (b) Two peptides of mitoribosomal origin with high cycloheximide-resistant label: one of 7.8 kDa (the OLII gene product), the second of 20 kDa (the OLI2 gene product) . Neither of these peptides is readily stained by Coomassie blue in the purified ATPase complex, (c) Four Coomassie-blue-stained peptides (27.5, 25, 23.5 and 22.5 kDa), the last one being subunit 6, which (like class b) functionally qualify as subunits of the membrane factor of the ATPase complex . As shown by labeling with 35SO24- and 3H-labeled amino acids, their biosynthesis is only very slightly cycloheximide-resistant, although it is submitted to coordinate regulation distinct from that of the class F1 peptides . 2 . Consequently it was shown that the OLI2 gene product, a 20-kDa peptide with high cycloheximide-resistant label, which was generally taken to be 'subunit 6' of the ATPase, is not in fact identical to this peptide.

Cell, 1982 Feb, 28(2), 355 - 64
Analysis of the junction between ribosomal RNA genes and single-copy chromosomal sequences in the yeast Saccharomyces cerevisiae; Zamb TJ et al.; The yeast Saccharomyces cerevisiae has a single tandem array of 100 ribosomal RNA (rRNA) genes . We have cloned and characterized a junction between the centromere-distal end of this array and the adjacent single-copy chromosomal sequences . We have shown that the junction occurs within the nontranscribed region of the repeat . By mapping the junction, we have found that the 35S rRNA precursor is transcribed toward the centromere while the 5S rRNA is transcribed away from the centromere . We have also shown that different yeast strains can have different single-copy sequence at the junction.

J Bacteriol, 1982 Feb, 149(2), 542 - 7
The herpes simplex virus thymidine kinase gene is not transcribed in Saccharomyces cerevisiae; Kiss GB et al.; The herpes simplex virus thymidine kinase gene has been cloned into a chimeric yeast plasmid cloning vehicle and transformed into appropriate yeast strains . Plasmids carrying the herpes simplex virus thymidine kinase gene can be propagated as autonomously replicating plasmids, but no RNA specific to the thymidine kinase coding sequence was detected.

J Biol Chem, 1982 Jan 25, 257(2), 870 - 4
Genetic evidence for a role of hexokinase isozyme PII in carbon catabolite repression in Saccharomyces cerevisiae; Entian KD et al.; A mutant of Saccharomyces cerevisiae that was selected for resistance to carbon catabolite repression also had reduced hexokinase activity . Hexokinase isoenzymes were purified from mutant and wild type cells . The specific glucokinase and hexokinase isozyme PI were present at normal levels in mutant and wild type, but no hexokinase isozyme PII activity was detected in the mutant . Staining for enzyme activity after electrophoresis of crude extracts also indicated that hexokinase PII was absent in the mutant . Mutant and wild type segregants gained by tetrad analysis were investigated electrophoretically . Staining for enzyme activity confirmed that catalytically inactive hexokinase PII and the defect in carbon catabolite repression always co-segregated . The results support the hypothesis that hexokinase PII might mediate carbon catabolite repression.

J Biol Chem, 1982 Jan 25, 257(2), 781 - 7
Repair of mitochondrial DNA in Saccharomyces cerevisiae . Induction of cytoplasmic petite mutants in a nuclear mutant exhibiting thermosensitive mitochondrial deoxyribonuclease activity; Foury F; Four nuclear thermosensitive mutants have been obtained in which induction of up 100% cytoplasmic petite mutants (rho-) is observed upon cell incubation at 36 degrees C . For a given incubation time at 36 degrees C, the percentage of rho- is increased by preliminary gamma-ray irradiation . Under these conditions, the induction of rho- is a linear function of the irradiation dose . The retention of genetic information by rho- and of mitochondrial DNA synthesis in vivo and in vitro exclude that the mutants are deficient in the replication of mitochondrial DNA . The degradation of mitochondrial DNA labeled with {3H}dTTP in isolated mitochondria, has been monitored at 26 degrees C and at 36 degrees C after addition of 0.5% Triton X-100 in the presence or in the absence of ethidium bromide . In assays carried out at 26 degrees C, the degradation of mitochondrial DNA is similar in the parental strain and in the mutant gamma s rho 2 . However, at 36 degrees C, the degradation of mitochondrial DNA is slower in the mutant . We have shown that a mitochondrial membrane deoxyribonuclease acting on double-stranded DNA at acid pH is thermosensitive in the mutant . Analysis of the meiotic segregants of a tetrad issued from the cross of the mutant with an isogenic parental strain shows co-segregation of rho- induction and of nuclease thermosensitivity in a 2:2 Mendelian pattern . These results suggest that a mitochondrial deoxyribonuclease is involved in the repair of damages caused to mitochondrial DNA by elevated temperature and by x-rays.

Biochim Biophys Acta, 1982 Jan 20, 679(1), 28 - 34
Affinity chromatography purification of cytochrome c oxidase and b-c1 complex from beef heart mitochondria . Use of thiol-sepharose-bound Saccharomyces cerevisiae cytochrome c; Bill K et al.; A method for simultaneous purification of cytochrome c reductase and cytochrome c oxidase using a cytochrome c affinity column is presented . Cytochrome c from Saccharomyces cerevisiae was linked to an activated thiol-Sepharose gel via its Cys-102 residue located far from the lysine residues on the front side of the molecule, responsible for the interaction with the reductase and oxidase . In previously reported affinity chromatography techniques these lysine residues most probably reacted with the column . Cytochrome c oxidase and reductase from bovine heart mitochondria bind specifically to the affinity column and can be recovered separately at different ionic strength in the elution buffer . The enzymes are highly pure and active.

Mol Cell Biol, 1982 Jan, 2(1), 21 - 9
Physiological characterization of Saccharomyces cerevisiae mutants supersensitive to G1 arrest by a factor and alpha factor pheromones; Chan RK et al.; Saccharomyces cerevisiae MATa cells carrying mutations in either sst1 or sst2 are supersensitive to the G1 arrest induced by alpha factor pheromone . When sst1 mutants were mixed with normal SST+ cells, the entire population recovered together from alpha factor arrest, suggesting that SST+ cells helped sst1 mutants to recover . Complementation tests and linkage analysis showed that sst1 and bar1, a mutation which eliminates the ability of MATa cells to act as a "barrier" to the diffusion of alpha factor, were lesions in the same genes . These findings suggest that sst1 mutants, are defective in recovery from alpha factor arrest because they are unable to degrade the pheromone . In contrast, recovery of sst2 mutants was not potentiated by the presence of SST+ cells in mixing experiments . When either normal MATa cells or mutant cells carrying defects in sst1 or sst2 were exposed to alpha factor for 1 h and then washed free of the pheromone, the sst2 cells subsequently remained arrested in the absence of alpha factor for a much longer time than SST+ or sst1 cells . These observations suggest that the defect in sst2 mutants is intrinsic to the cell and is involved in the mechanism of alpha factor action at some step after the initial interaction of the pheromone with the cell . The presence of an sst2 mutation appears to cause a growth debility, since repeated serial subculture of haploid sst2-1 strains led to the accumulation of faster-growing revertants that were pheromone resistant and were mating defective ("sterile").

Mol Cell Biol, 1982 Jan, 2(1), 11 - 20
Isolation and genetic analysis of Saccharomyces cerevisiae mutants supersensitive to G1 arrest by a factor and alpha factor pheromones; Chan RK et al.; Eight independently isolated mutants which are supersensitive (Sst-) to the G1 arrest induced by the tridecapeptide pheromone alpha factor were identified by screening mutagenized Saccharomyces cerevisiae MATa cells on solid medium for increased growth inhibition by alpha factor . These mutants carried lesions in two complementation groups, sst1 and sst2 . Mutations at the sst1 locus were mating type specific: MATa sst1 cells were supersensitive to alpha factor, but MAT alpha sst1 cells were not supersensitive to a factor . In contrast, mutations at the sst2 locus conferred supersensitivity to the pheromones of the opposite mating type on both MATa and MAT alpha cells . Even in the absence of added alpha pheromone, about 10% of the cells in exponentially growing cultures of MATa strains carrying any of three different alleles of sst2 (including the ochre mutation sst2-4) had the aberrant morphology ("shmoo" shape) that normally develops only after MATa cells are exposed to alpha factor . This "self-shmooing" phenotype was genetically linked to the sst2 mutations, although the leakiest allele isolated (sst2-3) did not display this characteristic . Normal MATa/MAT alpha diploids do not respond to pheromones; diploids homozygous for an sst2 mutation (MATa/MAT alpha sst2-1/sst2-1) were still insensitive to alpha factor . The sst1 gene was mapped to within 6.9 centimorgans of his6 on chromosome IX . The sst2 gene was unlinked to sst1, was not centromere linked, and was shown to be neither linked to nor centromere distal to MAT on the right arm of chromosome III.

Mol Cell Biol, 1982 Jan, 2(1), 1 - 10
Synthesis of repressible acid phosphatase in Saccharomyces cerevisiae under conditions of enzyme instability; Bostian KA et al.; The synthesis of repressible acid phosphatase in Saccharomyces cerevisiae was examined under conditions of blocked derepression as described by Toh-e et al . (Mol . Gen . Genet . 162:139-149, 1978) . Based on a genetic and biochemical analysis of the phenomenon these authors proposed a new regulatory model for acid phosphatase expression involving a simultaneous interaction of regulatory factors in the control of structural gene transcription . We demonstrate here that under growth conditions that fail to produce acid phosphatase the enzyme is readily inactivated . Furthermore, we demonstrate under these conditions the production of acid phosphatase mRNA which is active both in vitro and in vivo in the synthesis of enzyme . This eliminates any step prior to translation of acid phosphatase polypeptide as an explanation for the phenomenon . We interpret our results for the block in appearance of acid phosphatase as a result of both deaccelerated growth and cellular biosynthesis during derepression, accompanied by an enhanced instability of the enzyme.

Mol Gen Genet, 1982, 186(2), 295 - 7
Enrichment for auxotrophic mutants in Saccharomyces cerevisiae using the cell wall inhibitor, echinocandin B; McCammon MT et al.; Echinocandin B has been shown to inhibit fungal cell wall synthesis . This report describes the use of echinocandin B to enrich for nutritional auxotrophs in a mutagenized strain of Saccharomyces cerevisiae . Up to 20-fold enrichment levels for total auxotrophs were achieved after a single round of treatment with echinocandin; this level of enrichment is the highest of all procedures for which a specific mutant strain is not required.

Mol Gen Genet, 1982, 186(2), 253 - 8
Genes involved in the control of nuclear fusion during the sexual cycle of Saccharomyces cerevisiae; Polaina J et al.; Mutants of Saccharomyces cerevisiae defective for nuclear fusion have been isolated . Their mutations have been characterized by meiotic analysis, dominance-recessivity and complementation . Twelve of the mutations are allelic to the previously described kar 1-1; five affect a second gene designated KAR 2 and three affect a third gene designated KAR 3 . There is evidence suggesting that other two mutants are affected in a gene different from the three mentioned . Mutations in KAR 1 and KAR 2 genes are recessive and do not cause obvious effects other than the failure of the karyogamy . Mutations in KAR 3 are semidominant and do cause pleiotropic effects affecting both a mitosis and meiosis.

Mol Gen Genet, 1982, 186(1), 40 - 3
Suppression of maltose-negative phenotype by a specific nuclear gene (PMU1) in the petite cells of the yeast Saccharomyces cerevisiae; Khan NA; A small percentage of the primary petites isolated from strain 1403-7A-P1, constitutive for maltase synthesis, simultaneously lost the ability to utilize maltose and alpha-methylglucoside . Further studies showed that these primary petites were not stable with respect to maltose utilization . Approximately 30% of the secondary petites when isolated from the primary petites after vegetative growth were found to papillate on maltose plates . Tetrad analysis data revealed that a nuclear gene has reverted in these papillae, which is responsible for suppression of the maltose negative phenotype in primary petites . We have designated this nuclear gene as the PMU1 gene (petite maltose utilizer) . The functional form of the PMU1 gene is required in addition to the MAL4 gene for both constitutive maltase synthesis and maltose utilization in cytoplasmic petite cells derived from strain 1403-7A-P1.

Carcinogenesis, 1982, 3(4), 439 - 44
Role of DNA repair in ethyl methanesulfonate-induced mutagenesis in Saccharomyces cerevisiae; Prakash L et al.; Ethyl methanesulfonate (EMS)-induced reversion of two sites which revert preferentially by GC to AT transitions, cycl-131 and cycl-115, has been examined in readiation sensitive, rad, mutants of yeast belonging to the rad52 epistasis group . The rad50, rad51, rad52, rad54 and rad56 mutants showed reducted reversion of both tester sites when stationary phase diploid cells were treated with EMS . No correlation was found between EMS-induced reversion and EMS-induced homologous mitotic intragenic recombination . Survival of rad6 rad52 double mutants following EMS treatment indicates that there is one epistasis group for the repair of EMS-induced lethal damage in yeast . A model involving misrepair mutagenesis of specific lesions is proposed to account for the experimental results.

Mol Gen Genet, 1982, 185(2), 207 - 10
Changes in production of the mating-type-specific glycoproteins, agglutination substances in association with mating type interconversion in homothallic strains of the yeast, Saccharomyces cerevisiae; Nakagawa Y et al.; Sexual activity in homothallic strains of Saccharomyces cerevisiae was investigated . We succeeded in culturing homothallic haploid cells without conjugation, by lowering the pH value of the culture medium . In spore cultures of a homothallic strain both a and alpha pheromones were detected . Agglutination substance of a and alpha mating types were detected in homothallic haploid cells from spore cultures in early logarithmic phase regardless of mating type information at the HML and HMR loci, but either a or alpha agglutination substance was detected predominantly in homothallic haploid cells from spore culture in late logarithmic phase, depending on mating type information at the HML and HMR loci.

Proc Natl Acad Sci U S A, 1982 Jan, 79(2), 253 - 6
Chromatin conformational changes accompany transcriptional activation of a glucose-repressed gene in Saccharomyces cerevisiae; Sledziewski A et al.; In the present study we have analyzed the kinetics of DNase I digestion of the two alcohol dehydrogenase (ADH) genes within yeast nuclei . We have found that in yeast grown on glucose the constitutively transcribed ADCI gene is much more sensitive to DNase I digestion than is the repressible ADR2 gene . In yeast grown on ethanol, both genes are transcribed and both exhibit the same sensitivity to DNase I attack . We have also found and mapped DNase I hypersensitive sites near the 5' ends of constitutive and repressible ADH genes . These sites are well correlated with the position at which transcription is initiated.

Mutat Res, 1982 Jan-Feb, 100(1-4), 163 - 8
The activity of 4CMB, 4HMB and BC in Saccharomyces cerevisiae JD1; Wilcox P et al.; 3 structurally related compounds, 4-chloromethylbiphenyl (4CMB), 4-hydroxymethylbiphenyl (4HMB), and benzyl chloride (BC) were assayed for their ability to induce mitotic gene conversion in stationary phase cultures of the yeast, Saccharomyces cerevisiae JD1 . This strain allows gene conversion to be scored at 2 independent loci, trp 5 and his 4 . The results reported in this paper indicate that both 4CMB and BC are genetically active in yeast, producing dose-related increases in mitotic gene conversion at both the loci tested; 4HMB showed no such activity . At high survival levels 4CMB and BC showed comparable activity . However, as toxicity increased BC showed much more potent convertogenic activity, whereas with 4CMB a reduction in induced gene conversion was observed . The presence of a microsomal activation system derived from the livers of Aroclor-induced male rats did not significantly affect the activity of any of the compounds.

Mutat Res, 1982 Jan-Feb, 100(1-4), 157 - 62
The induction of mitotic gene conversion in the yeast, Saccharomyces cerevisiae JD1 by 4-chloromethylbiphenyl (4CMB), benzyl chloride (BC) and 4-hydroxymethylbiphenyl (4HMB); Brooks TM et al.; The induction of mitotic gene conversion by 4CMB, BC and 4HMB was studied in both log-phase and stationary-phase cultures of the yeast, Saccharomyces cerevisiae JD1 . Assays were performed both in the presence and in the absence of S9 microsomal fraction obtained from a liver homogenate from rats pretreated with Aroclor 1254 . Exposure of both stationary-phase and log-phase cultures to 4CMB and BC resulted in an increase in mitotic gene conversion, both in the presence and in the absence of a microsomal activation system; the magnitude of response was greater in stationary-phase cultures . 4HMB did not increase the gene conversion frequency in log-phase or stationary-phase cultures.

Eur J Biochem, 1982 Jan, 121(3), 643 - 7
Pleiotropic deficiency in nitrogen-uptake systems and derepression of nitrogen-catabolic enzymes in npr-1 mutants of Saccharomyces cerevisiae; Grenson M et al.; The npr-1 mutation has two types of effects in Saccharomyces cerevisiae . On one hand, several nitrogen-catabolic enzymes are derepressed in the presence of ammonia, glutamine, or asparagine, which provoke their repression in a wild-type strain . On the other hand, the activity of several ammonia-sensitive permeases is decreased (from 50-100% depending on the permease considered) in npr-1 cells, independently of the nitrogen source used for growth . Our results favour the idea that the primary effect of the npr-1 mutation is on the permeases, and that the derepression of the enzymes is a consequence of the reduced uptake rate of the repressing nitrogen compounds . Hence, the product of the npr-1 gene appears to be directly involved in the development of the activity of a set of permeases which transport nitrogen compounds and which are regulated by nitrogen effectors.

Microbios, 1982, 35(141-142), 161 - 8
Peptide fragments of the his4 trifunctional protein from Saccharomyces cerevisiae; Bigelis R; Labile altered forms of the his4 protein from Saccharomyces cerevisiae were studied in extracts of mutant strains or in extracts subjected to proteolysis . Fragments of the his4 protein were detected by immunoautoradiography in high salt extracts of his4 deletion strains having histidinol dehydrogenase activity but lacking two of the other enzyme activities associated with the trifunctional his4 protein . A form of the protein altered by proteolysis was also detected in low salt crude extracts by immunodiffusion and shown to possess histidinol dehydrogenase function by activity staining of the precipitin lines . These immunological data indicated that histidinol dehydrogenase can function independently of the other two activities in extracts of deletion strains or in extracts where the wild type his4 protein has undergone proteolytic cleavage . The data also indicated that high concentration in (NH4)2SO4 had a substantial stabilizing effect on the his4 protein and on fragments of the his4 protein from mutant strains.

Can J Genet Cytol, 1982, 24(5), 493 - 503
Allelism of pleiotropic drug resistance in Saccharomyces cerevisiae; Saunders GW et al.; Allelism of pleiotropic drug resistant (pdr) mutants was evaluated by complementation tests, linkage to chromosome-VII centromere markers and response to a partial suppressor (sur) . Complementation tests were confounded by incomplete dominance and somatic segregation . Phenotypic suppression by sur was observed for all mutant and wild type alleles and thus could not be used to distinguish alleles . Five different alleles were tentatively identified by their close linkage to leul; 88 tetrads from three factor crosses produced the following linkages--leul (4.7) pdrl (17.0) trp5 . Resistance of DRI 9/T7, a {cir o} strain of French origin, was not inherited as an allele of pdr but was controlled by a different pleiotropic centromere linked gene . An evaluation of published data suggest that antl, AMYl, till, cyh3, BOR2, and axe1 may be alleles of pdr . Thus pdr appears to be an allele that influences permeability to many inhibitors.

Microbios, 1982, 35(140), 99 - 110
Studies on calcium efflux in the yeast Saccharomyces cerevisiae; Eilam Y; The properties of the 45Ca efflux systems in Saccharomyces cerevisiae were investigated in yeast cells grown overnight in medium containing 45Ca . Efflux was measured in medium containing glucose and Tris-Hepes buffer adjusted to the required pH . In the absence of permeable cations in the medium, at pH 5.2, 20% of the cellular Ca was extruded from the cells during the first 2 h . There was no further decrease in the amount of cellular Ca during an additional 24 h of incubation . The initial rate of Ca efflux was markedly reduced with the increase in the pH of the medium . On the other hand the efflux during the second phase (2-24 h) increased with the increase in medium-pH up to pH 7.5 . It is suggested that the initial rapid phase of Ca efflux, in the absence of permeable cations, represents transport across the plasma membrane and is mediated via a Ca2+/H+ antiport . The second phase represents the release of Ca sequestered in some cellular organelles, probably the vacuoles, and is mediated via a different mechanism . Addition of Ca or Mg to the medium markedly stimulated the rate of Ca efflux from both cellular compartments . At the same time a predominant influx of divalent cations was observed . This exchange between intracellular Ca and extracellular divalent cations was not affected by the pH of the medium between pH 5.2 and 7.0 . Both processes, Ca efflux and Ca-Mg exchange, required cellular energy; they were almost completely inhibited in the absence of glucose and the presence of antimycin A, a respiratory inhibitor.

Mol Gen Genet, 1982, 188(3), 384 - 91
Ribosome structure, maturation of ribosomal RNA and drug sensitivity in temperature-sensitive mutants of Saccharomyces cerevisiae; Gritz LR et al.; Selected strains of Saccharomyces cerevisiae were mutagenized with nitrosoguanidine and temperature-sensitive mutants isolated . These mutants were screened by two-dimensional gel-electrophoresis for the presence of ribosomal proteins with altered mobility relative to parental preparations . Electrophoretic changes were detected in three mutants designated ts205, ts212 and ts417, with the alterations apparently the same in the three cases . All three mutants were more sensitive than were their parents to the antibiotics G418, hygromycin B and MDMP . Mutant ts212 has an abnormal distribution of native ribosomal subunits and appears to be defective in its assembly of the smaller subparticle.

Mol Gen Genet, 1982, 187(1), 54 - 60
Meiotic cytology of Saccharomyces cerevisiae in protoplast lysates; Goetsch L et al.; This report describes cytological features of meiosis in Saccharomyces cerevisiae prepared for electron microscopy by lysis of protoplasts or nuclei on an aqueous surface . Whereas the chromatin of cells lysed before or after meiotic prophase was widely dispersed, pachytene bivalents appeared as discrete, elongate masses of compact chromatin . These bivalents were of nearly uniform thickness; they ranged in length from about 0.6 micrograms to 4.0 micrograms, with a median of 1.6-1.8 micrograms . Enzymatic digestion of chromosomal DNA removed the chromatin to reveal the underlying synaptonemal complex . The lysis of partially purified nuclei was less disruptive and thereby revealed the regular association of the telomeres with fragments of the nuclear envelope . In tetraploid cells, pachytene lysates contained quadrivalents characterized by the close apposition of chromatin masses of similar length . One or more points of intimate association appear to represent sites of exchange between pairing partners . The departure of the diploid cells from pachytene was accompanied by the renewed association of spindle microtubules with the chromosomes shortly before the diplotene chromosomes decondensed . Later, the successive meiotic divisions were identified by the appearance of a single spindle for meiosis I and of two spindles for meiosis II.

Mol Gen Genet, 1982, 187(1), 47 - 53
Reversible pachytene arrest of Saccharomyces cerevisiae at elevated temperature; Byers B et al.; The temperature sensitivity of sporulation in a well-characterized yeast strain lacking any known temperature sensitive genes has been investigated . Cytological observations by electron microscopy demonstrate that cells incubated in sporulation medium at a temperature inhibitory to sporulation became arrested in meiotic prophase . The stage of arrest was identified as pachytene by the presence of duplicated (but unseparated) spindle pole bodies and synaptonemal complex . Transfer of the arrested culture to lower temperature permitted resumption of meiosis and sporulation; transfer to vegetative medium resulted in reversion to mitotic division . Genetic analysis of cells that had reverted to mitosis revealed that commitment to intragenic recombination had occurred by the time of arrest . Prolonged incubation at the elevated temperature resulted in the enhancement of intragenic recombination above normal levels, suggesting that some aspect of recombination continued to occur during the pachytene arrest . Evidence is presented that DNA replication, although depressed overall in the arrested cultures, had occurred to completion in many arrested cells.

Mol Gen Genet, 1982, 187(1), 42 - 6
New temperature-sensitive mutants of Saccharomyces cerevisiae affecting DNA replication; Dumas LB et al.; We have isolated new mutants of the yeast Saccharomyces cerevisiae that are defective in mitotic DNA synthesis . This was accomplished by directly screening 11000 newly isolated temperature-sensitive yeast clones for DNA synthesis defects . Ninety-seven different mutant strains were identified . Approximately half had the fast-stop DNA synthesis phenotype; synthesis ceased quickly after shifting an asynchronous population of cells to the restrictive temperature . The other half had an intermediate-rate phenotype; synthesis continued at a reduced rate for at least 3 h at the restrictive temperature . All of the DNA synthesis mutants continued protein synthesis at the restrictive temperature . Genetic complementation analysis of temperature-sensitive segregants of these strains defined 60 apparently new complementation groups . Thirty-five of these were associated with the fast-stop phenotype, 25 with the intermediate-rate phenotype . The fast-stop groups are likely to include many genes whose products play direct roles in mitotic S phase DNA synthesis . Some of the intermediate-rate groups may be associated with S phase as well . This mutant collection should be very useful in the identification and isolation of gene products necessary for yeast DNA synthesis, in the isolation of the genes themselves, and in further analysis of the DNA replication process in vivo.

Mol Gen Genet, 1982, 188(2), 261 - 5
Mutations affecting the activity and the regulation of the general amino-acid permease of Saccharomyces cerevisiae . Localisation of the cis-acting dominant pgr regulatory mutation in the structural gene of this permease; Grenson M et al.; Mutants lacking the general amino acid permease activity fall into two classes of complementation . Mutations at the GAP1 locus abolish the general amino acid permease activity specifically, while those in the NPR1 locus simultaneously affect several other ammonia-sensitive uptake systems . The NPR1 locus as well as the GAP1 locus code for proteins, as shown by the identification of nonsense mutations in both these genes . Frameshift mutations in the GAP1 locus, and conditional, thermosensitive, mutations in the NPR1 locus were also obtained . No intragenic complementation was detected among 33000 crosses between gap1- mutant strains . Mutations at three unlinked loci, namely MUT2, MUT4, and PGR, release one of the two controls which prevent expression of the GAP1 gene in ammonia-grown yeast cells . The pgr regulatory mutation is located in the GAP1 locus near one end of this region, the fine structure of which has been determined by X-ray-induced mitotic recombination . On the basis of the properties of the mutants it is likely that the PGR region determines a receptor site for the negative control mediated by the products of the MUT2 and MUT4 genes . The data presented here are compatible with this negative control operating either at the transcriptional or at a post-transcriptional level of the GAP1 gene expression . The present work initiates the study of the regulation of the general amino acid permease at the molecular level.

Mol Gen Genet, 1982, 188(2), 235 - 9
Defective excision of pyrimidine dimers and interstrand DNA crosslinks in rad7 and rad23 mutants of Saccharomyces cerevisiae; Miller RD et al.; Excision of pyrimidine dimers and interstrand DNA crosslinks was examined in the deletion mutants rad7-delta 1, rad23-delta 1, and rad7-delta 1 rad23-delta 1 . These mutants remove pyrimidine dimers and crosslinks much less efficiently than the RAD+ strains; only 30-60% of pyrimidine dimers and 25-40% of crosslinks are removed even after prolonged incubation . The rad7 and rad23 mutations may represent defects in protein factors which increase the efficiency of the nicking enzyme complex or make chromatin more accessible to the nicking activity.

Zentralbl Mikrobiol, 1982, 137(5), 421 - 32
Fine cytochemical localization of polyphosphates in the yeast Saccharomyces cerevisiae; Vorisek J et al.; A late exponential culture, cultivated in the absence of phosphates, and a similar culture supplied with phosphate (phosphate overcompensation conditions) were prepared from an industrial strain of Saccharomyces cerevisiae . For the cytochemical staining, the cellular phosphates were transformed into polymeric metal-phosphate complexes by Ca2+ and Mg2+ ions, added to the fixative . The fixative contained 3% glutaraldehyde, buffered by 100 mM Tris-HCl to pH 6.0, plus 100 mM MgCl2 and 100 mM CaCl2 . Staining with lead acetate was followed by OsO4 post-fixation . In cells cultivated in the absence of phosphates lead deposits were found in vacuoles only . In the late exponential culture the staining was observed on the surface of the plasmalemma, on the membranes of the endoplasmic reticulum, in mitochondria, in the cell nucleus, and in vacuoles . As a rule, extensive polyphosphate deposits (metachromatic granules) were found in vacuoles . Two hours after phosphate overcompensation, a high quantity of polyphosphate as found also in the cell wall, e.g., in the isthmus of budding cells (scar ring), in the secondary septa of mother and daughter cells, and in the growth apex . When divalent cations were omitted from the fixative, the staining of polyphosphates was limited to the cell wall and large vacuolar granules . The results of cytochemical staining were compared with the biochemical analysis of polyphosphate content in the cultures under study.

Mikrobiologiia, 1982, 51(5), 761 - 4
{Concurrent effect of ultraviolet rays of various wave lengths on Saccharomyces cerevisiae survival}; Ivanova EV et al.; While studying the combined action of different UV wavelengths on Saccharomyces cerevisiae (strains XII and Cl-9), it has been found that preliminary irradiation with UV at 334 nm caused photoprotection of the cells against the lethal action of UV at 254 and 313 nm . Postradiation irradiation of strain XII incapable of photoreactivation with UV at 334 nm increased the lethal action of UV at 254 and 313 nm . The mechanisms of the both effects are based on serotonin synthesis induced by the light at 334 nm, as was shown using p-chlorophenylalanine, a specific inhibitor of serotonin synthesis . In strain CI-9 capable of photoreactivation, the postradiation effect of the light at 334 nm depends on the interaction of two different photobiological reactions induced by it, namely, photoreactivation and induced serotonin synthesis.

Mol Gen Genet, 1982, 186(4), 501 - 6
Mitochondrial and nuclear myxothiazol resistance in Saccharomyces cerevisiae; Thierbach G et al.; Mitochondrial and nuclear mutants resistant to myxothiazol were isolated and characterized . The mitochondrial mutants could be assigned to two loci, myx1 and myx2, by allelism tests . The two loci map in the box region, the split gene coding for apocytochrome b . Locus myx1 maps in the first exon (box4/5) whereas myx2 maps in the last exon (box6) . The nuclear mutants could be divided into three groups: two groups of recessive mutations and one of dominant mutations . Respiration of isolated mitochondria from mitochondrial mutants is resistant to myxothiazol . These studies support the conclusion that myxothiazol is an inhibitor of the respiratory chain of yeast mitochondria . The site of action of myxothiazol is mitochondrial cytochrome b.

Mol Gen Genet, 1982, 186(4), 478 - 81
cdc7-1 a temperature sensitive cell-cycle mutant which interferes with induced mutagenesis in Saccharomyces cerevisiae; Njagi GD et al.; The mutant cdc7-1 is shown here to block UV induced reversion of six different auxotrophic mutations and forward mutations at several genes concerned with adenine biosynthesis in Saccharomyces cerevisiae . Chemical mutagenesis is also drastically reduced . In its effect on mutagenesis cdc7-1 resembles rad6-1 . However, in contrast to rad6-1, cdc7-1 does not affect sporulation or mitotic recombination neither is it sensitive to the antifolate drug trimethoprim . It appears to fall in the same epistatic group as rad6-1 . Possible explanations for its action are briefly considered.

Mol Gen Genet, 1982, 186(3), 445 - 8
A further two mutants defective in initiation of the S phase in the yeast Saccharomyces cerevisiae; Johnston LH et al.; Two new mutants dbf3 and 4, are specifically defective in DNA synthesis . When synchronous cultures of dbf4 were transferred from the permissive to restrictive temperature before the start of the S phase, no DNA synthesis occurred . However when switched after the beginning of DNa replication, the cells completed that round of synthesis . Dbf4 therefore resembles cdc7, a mutant believed to be required for initiation of DNA synthesis, and indeed dbf4 acts at a point in the cell cycle close to cdd7, namely between cdd4 and the final requirement for protein synthesis before the S phase . Like dbf4, cultures of dbf3 transferred to restrictive conditions before the start of S showed little DNA synthesis . However a small burst did occur at a time roughly corresponding to when normal initiation would be expected . Cultures switched after this time completed the ongoing round of DNA synthesis.

Mol Gen Genet, 1982, 186(3), 439 - 44
The isolation of new DNA synthesis mutants in the yeast Saccharomyces cerevisiae; Johnston LH et al.; Sixty-eight new conditional cell cycle mutants have been isolated on the basis of their terminal cellular morphology ('dumbbells') . Fifteen mutants falling into nine complementation groups, were grossly defective in DNA replication and have been assigned the provisional gene symbol dbf (for dumbbell former) . Dbf1 and 2 stop DNA synthesis immediately on transfer to 37 degrees C and are presumably defective in enzymes required for polymerization . Neither, however, possess a thermolabile DNA polymerase A or B . Dbf3 and 4 show a pattern of synthesis consistent with their being deficient in initiation of DNA synthesis . This is confirmed in the accompanying paper . The remaining mutants are deficient in the synthesis of RNA as well as DNA . Indeed the four members of one complementation group are allelic with rna3, one of the group of mutants originally isolated as defective in RNA synthesis, and which do not exhibit a cell cycle phenotype . A re-examination of this group of mutants, however, showed the bulk of them also to be defective in DNA synthesis . Furthermore, in preliminary experiments rna3 and our four new alleles of it, together with rna6 and dbf5 and 6, showed enhanced spontaneous mutation frequency.

EMBO J, 1982, 1(9), 1125 - 31
Molecular cloning, DNA structure, and RNA analysis of the arginase gene in Saccharomyces cerevisiae . A study of cis-dominant regulatory mutations; Jauniaux JC et al.; The Saccharomyces cerevisiae gene cargA + or CAR1 , encoding arginase has been cloned by recovering function in transformed yeast cells . It was used to analyse RNA and chromosomal DNA from six strains bearing cis-dominant regulatory mutations leading to constitutive arginase synthesis . The DNA from the four cargA + O- strains in which constitutive arginase synthesis was independent of the mating-type functions showed no detectable differences with the wild- typye . The cargA + O- mutations were, therefore, small alterations, possibly single base substitutions . On the other hand, the cargA + Oh-1 and cargA + Oh-2 mutations, leading to a constitutive and mating-type dependent arginase synthesis, were identified as insertions . Their size and restriction pattern strongly suggested that they were induced by the Ty1 yeast transposable element . This was confirmed by cloning and analysis of the cargA + Oh-1 mutant gene . The concentration of arginase RNA was significantly increased in the mutants, indicating that the regulation of arginase synthesis was exerted, at least in part, at the level of RNA synthesis or stability . In the cargA + Oh-2 strain the Ty1 element was located at a distance of approximately 600 base pairs from the insertion present in the cargA + Oh-1 strain . This result suggests either a surprisingly large arginase regulatory region or an indirect influence of the Ty1 element on gene expression over long distances.

Mol Gen Genet, 1982, 188(1), 51 - 9
The cytochrome oxidase subunit I split gene in Saccharomyces cerevisiae: genetic and physical studies of the mtDNA segment encompassing the 'cytochrome b-homologous' intron; Netter P et al.; We have constructed a refined genetic and physical map of 38 oxi3 mutations . With the help of the rho- clones derived from 'short' and 'long' genes, pairwise crosses between mutants, estimations of their reversion frequencies and analyses of mitochondrially synthesized proteins, we have characterized and localized several mutants in the exon A4 and in the intron aI4 . We present genetic and physical evidence that in the 'long' gene the exon A5 is split into at least three quite distinct exons, A5-1, A5-2 and A5-3 where numerous mutations are localized . We suggest that a novel 56 Kd polypeptide, which accumulates in some cis-dominant oxi3- mutants results from the translation of the upstream exons and the downstream aI4 intron.

Acta Microbiol Pol, 1982, 31(2), 119 - 28
cdc and prt Mutants of Saccharomyces cerevisiae with increased sensitivity to diepoxybutane and ultraviolet; Baranowska H et al.; The sensitivity to UV and DEB of nine temperature-sensitive mutants was investigated . All prt and four cdc mutants (cdc4, cdc7, cdc8, cdc28) showed a higher level of sensitivity to UV than the wild type strain . Three mutants cdc7, cdc9 and prt1 are more sensitive than the wild type to DEB, but only two cdc7 and prt1 are sensitive to UV and DEB . Treatment with restrictive temperature increases the sensitivity after UV treatment of cdc9, cdc21, prt1, prt3 and after DEB treatment of cdc28 and all prt mutants.

EMBO J, 1982, 1(9), 1133 - 9
Expression of the ROAM mutations in Saccharomyces cerevisiae: involvement of trans-acting regulatory elements and relation with the Ty1 transcription; Dubois E et al.; The regulatory mutations in Saccharomyces cerevisiae designated cargA + Oh, cargB + Oh, and durOh are alterations in the control regions of the respective structural genes . The alteration causing the cargA + Oh mutation has been shown to be an insertion of a Ty1 element in the 5' noncoding region of the CAR1 ( cargA ) locus . All three mutations cause overproduction of their corresponding gene products and belong to the ROAM family of mutations (Regulated Overproducing Allele responding to Mating signals) in yeast . The amount of overproduction in ROAM mutants is determined, at least in part, by signals that control mating functions in yeast . We report the identification of two genetic loci that regulate Oh mutant gene expression but that do not affect mating ability . These loci are defined by the recessive roc mutations ( ROAM mutation control) that reduce the amount of overproduction caused by the cargA + Oh, cargB + Oh, and durOh mutations . RNAs homologous to CAR1 ( cargA ), DUR1 ,2 and Ty1 DNA probes were analyzed by the Northern hybridization technique . In comparison with wild-type strains, cargA + Oh and durOh mutant strains grown on ammonia medium contain increased amounts of CAR1 and DUR1 ,2 RNA . This RNA overproduction is diminished in MATa/MAT alpha diploid strains as well as in haploid strains that also carry the ste7 mutation which prevents mating or that carry either of the roc1 or roc2 mutant alleles . The amount of RNA homologous to Ty1 DNA is also reduced in ste7 , roc1 , and roc2 mutant strains . This reduction is not observed in a strain with the ste5 mutation, which prevents mating but has no effect on overproduction of ROAM mutant gene products.(ABSTRACT TRUNCATED AT 250 WORDS)

Microbios, 1982, 34(136), 71 - 88
On the uptake of nystatin by saccharomyces cerevisiae . 5 The release of amino acids; Beezer AE et al.; The release of amino acids from Saccharomyces cerevisiae under the action of nystatin has been studied . The kinetic results do not show any dependence upon molecular size but do support earlier results suggesting that interactions depend upon slow diffusion processes through the cell wall . The effects of temperature, sterol concentration and the presence of calcium ion have also been investigated.

Int J Biochem, 1982, 14(2), 111 - 8
Enzyme activities during early ascosporulation in Saccharomyces cerevisiae; Ota A; 1 . Several enzyme activities were examined during the initial sporulating phase in Saccharomyces cerevisiae . 2 . Catalase activity increased obviously after transfer to sporulation medium . 3 . Catalase is probably considered to play an essential role in sporulation . 4 . Both activities of inorganic pyrophosphatase and glycerol-2-phosphatase decreased . 5 . Conditions necessary for sporulation were suggested.

J Biol Chem, 1981 Dec 25, 256(24), 13074 - 8
The gcr (glycolysis regulation) mutation of Saccharomyces cerevisiae; Clifton D et al.; gcr is a mutation considerably decreasing the assayed amounts of most glycolysis enzymes in Saccharomyces cerevisiae (Clifton, D., Weinstock, S . B., and Fraenkel, D . G . (1978) Genetics 88, 1-11) . We show here that although in the wild type strain the amounts of these enzymes do not greatly differ between cells from different media, in the gcr mutant strain most of the enzyme amounts are 5% or less, relative to wild type, from cells grown without sugars, but 20-50% from cells grown with sugars . Lower relative values were found for phosphoglycerate mutase and enolase . A corresponding alteration in the mutant in the intensities of several major protein bands could even be seen in stained gels after electrophoresis of crude extracts: the profiles were otherwise normal . Results of titration of phosphoglycerate kinase with antibody accorded with activity . Transfer of cells between the two types of media did not lead to a more rapid adjustment of enzyme amounts than expected from the steady state levels . gcr is not allelic to GPM (the gene for phosphoglycerate mutase) or to RNA1 (which affects transport of RNA from the nucleus) . Translation of total RNA in a rabbit reticulocyte lysate gave a pattern of polypeptides similar to the in vivo one . Thus, gcr is likely to affect somehow mRNA synthesis or lifetime for a discrete number of proteins.

J Biol Chem, 1981 Dec 25, 256(24), 13048 - 54
An assessment of the specificity of sterol uptake and esterification in Saccharomyces cerevisiae; Taylor FR et al.; By growing a sterol-requiring strain of Saccharomyces cerevisiae in the presence of pairs of sterols differing by a single structural change, the in vivo specificity of sterol uptake and esterification was measured . Uptake specificity was demonstrated for the delta 5-, delta 7-, and delta 22- bonds as well as the 24 beta-methyl . Sterol uptake was shown to depend on the metabolic state of the cell, and the apparent Km of uptake for ergosterol (11.1 microM) was lower than that of cholesterol (66.7 microM) . This difference in apparent Km can explain the preferential utilization of ergosterol . The selectivity for esterification showed that sterols lacking the delta 7- or delta 22- bond or the 24 beta-methyl were preferentially esterified . However, sterols lacking the delta 5-bond were not preferentially esterified . This specificity of uptake and esterification did not change significantly with alterations in the fatty acid source . These results suggest that both uptake and esterification are used to control the types of sterols in the free sterol fraction, resulting in the enrichment of ergosterol-like sterols in cellular membranes . An additional finding was that cells supplemented with sterols which have a delta 5,7-diene (7-dehydrocholesterol and ergosterol) had much reduced levels of steryl ester . This may be attributable to inhibition by a breakdown product(s) of these sterols.

Biochim Biophys Acta, 1981 Dec 21, 649(3), 550 - 6
Temperature effects on kinetic properties of plasma membrane ATPase from the yeast Saccharomyces cerevisiae; Ahlers J; The reaction of plasma membrane ATPase from yeast with Mg2+ and Mg X ATP was studied in a temperature range of 10-30 degrees C . The random mechanism of activation by Mg2+ and the pseudocompetitive inhibition at higher concentrations was not altered when the temperature was varied, nor were the kinetic constants representing substrate binding . However, at low temperature, the affinity of the enzyme for Mg2+ is greatly reduced . The Arrhenius plot of log V vs . 1/T shows straight lines with an inflection point at 24 degrees C, which disappears in the presence of detergent . Calorimetric studies of the plasma membranes show a transition point at the same temperature . From these findings we suppose that Mg2+ is bound at a regulatory site of the ATPase, which is influenced by surrounding phospholipids.

Nucleic Acids Res, 1981 Dec 11, 9(23), 6339 - 50
The actin gene in yeast Saccharomyces cerevisiae: 5' and 3' end mapping, flanking and putative regulatory sequences; Gallwitz D et al.; The 5' and 3' flanking regions of the yeast actin gene have been sequenced and the ends of the actin mRNA were determined by the single-strand nuclease mapping procedure . The mRNA starts with a pyrimidine residue 141 (or 140) nucleotides upstream from the initiation codon . The actin gene lacks a typical "TATA" box 30 base pairs upstream from the mRNA start site but it contains a region homologous to the canonical sequence 5'-GGCTCAATCT-3' which is found in several eukaryotic genes 70 to 80 bp upstream from the mRNA cap site . Judging from the S1 nuclease mapping, there are two populations of actin mRNA terminating 98 and 107 nucleotides downstream from the stop codon . The 3' termini are preceded by three AATAAA sequences found in most eukaryotic polyadenylated mRNAs.

J Biol Chem, 1981 Dec 10, 256(23), 12456 - 62
Thymidylate synthetase from Saccharomyces cerevisiae . Purification and enzymic properties; Bisson LF et al.; Thymidylate synthetase of Saccharomyces cerevisiae was purified over 20,000-fold to apparent homogeneity by a procedure involving two new affinity methods and several precautions for avoiding proteolysis . Molecular weight of the native enzyme was about 65,000, as determined by gel filtration and velocity sedimentation . Electrophoresis in polyacrylamide gels containing sodium dodecyl sulfate yielded a single band of molecular weight 30,000, suggesting that thymidylate synthetase is a dimer of very similar or identical subunits . The purified enzyme exhibited normal Michaelis-Menten kinetics toward both substrates, with apparent Km values for dUMP and for (--)-5,10-methylene-tetrahydropteroylglutamate of 5 microM and 70 microM, respectively . When the pentaglutamyl form of the cofactor was used, its apparent Km was lower (7 microM), but Vmax was unaltered . Reaction kinetics and product inhibition studies were most consistent with an ordered mechanism, wherein dUMP is the first substrate to bind and 7,8-dihydropteroylglutamate is the first product released . Halogenated analogs of the nucleotide substrate were competitive inhibitors of the yeast enzyme, with apparent Ki values for 5-fluoro-dUMP of 5 nM and for 5-Br-dUMP of 10 microM . Analogs of the cofactor were also competitive inhibitors, with apparent Ki values for both methotrexate and aminopterin of about 20 microM . Cibacron blue, a dye used as the ligand in an affinity adsorbent for one of the purification steps, was a potent competitive inhibitor with respect to either substrate, yielding apparent Ki values of 4 nM for the nucleotide binding site and 40 nM for the cofactor binding site.

Gene, 1981 Dec, 16(1-3), 335 - 41
Cloning of the HIS5 gene of Saccharomyces cerevisiae by yeast transformation; Harashima S et al.; A yeast DNA fragment complementing a yeast his5 mutation was cloned on a shuttle vector, YRp7, by transformation of the yeast host with plasmid DNA prepared from a gene bank constructed in the Escherichia coli host . That the DNA fragment contains the yeast HIS5 gene was confirmed by the integration of the cloned plasmid into the his5 region of the yeast chromosome . The cloned yeast HIS5 gene weakly complemented the E . coli hisC mutation and gave rise to a slow-growing E . coli transformant without addition of histidine . From the slow-growing culture, rapidly growing variants of E . coli were easily obtained by mutations either in the bacterial chromosome or in the plasmid.

Med Biol, 1981 Dec, 59(5-6), 272 - 8
Mutants of Saccharomyces cerevisiae deficient in polyamine biosynthesis: studies on the regulation of ornithine decarboxylase; Tabor CW; We have isolated the following mutants in the polyamine biosynthetic pathway in yeast: (i) spe10 mutants, which have no ornithine decarboxylase activity and therefore cannot make putrescine; (ii) spe2 mutants, which have no adenosylmethionine decarboxylase and therefore cannot make spermidine or spermine; (iii) spe3 mutants, which have no putrescine aminopropyltransferase and therefore cannot make spermidine and spermine, and (iv) spe4 and spe40 mutants (suppressors of spe10 mutations), which have no spermidine aminopropyltransferase and therefore cannot make spermine . These mutants show that (i) yeast has an absolute requirement for these amines for growth (ii) putrescine in the absence of spermidine and spermine supports growth at one-sixth the wild type rate; (iii) intracellular spermine controls the ornithine decarboxylase activity and thus mutants that cannot make spermine are derepressed for ornithine decarboxylase; (iv) Saccharomyces cerevisiae can make putrescine only by one pathway, i.e., ornithine decarboxylase; (v) spermidine and spermine are synthesized by different aminopropyltransferases in yeast; and (vi) spermidine and/or spermine are absolutely required for both sporulation and maintenance of the double-stranded RNA "killer" plasmid . We have purified ornithine decarboxylase to homogeneity and shown that loss of ornithine decarboxylase activity resulting from growth with added spermidine and spermine is the result of post-translational modification.

Mutat Res, 1981 Dec, 90(4), 411 - 23
Induction of petite formation in Saccharomyces cerevisiae by experimental antitumour agents . Structure--activity relationships for 9-anilinoacridines; Ferguson LR et al.; A series of 9-anilinoacridine derivatives has been compared in terms of DNA binding, ability to inhibit the growth of L1210 murine leukaemia cells in vitro, and ability to induce the formation of respiration-deficient (petite) mutations in Saccharomyces cerevisiae . The acridine ring in the derivatives was either unsubstituted, or substituted with amino groups at the 3 and/or 6 positions . The 9-anilino group was para-substituted with a variety of substituents . 3-Aminoacridine, 3,6-diaminoacridine (proflavine) and ethidium were included for comparison . The results show that: (i) at least one acridinyl amino group is necessary for conferring petite inducing activity in the yeast system; (ii) for 3-amino-9-anilinoacridine congeners, small anilino substituents provide compounds which resemble proflavine in their ability to produce petite mutants, whereas large substituents abolish activity; (iii) the 3,6-diamino-substituted anilinoacridines resemble ethidium rather than proflavine in being highly efficient inducers of petites; (iv) the requirements for optimal activity in L1210 leukaemia cell cultures are different to those for petite formation in yeast . It is concluded that the size of the anilino substituent, as well as its contribution to DNA binding, is critical in conferring biological activity in each system.

Eur J Biochem, 1981 Dec, 121(1), 113 - 8
Improved assay and mechanism of the reaction catalyzed by the chitin synthase from Saccharomyces cerevisiae; Fahnrich M et al.; 1 . An improved filtration method is introduced to perform kinetic investigations on the chitin synthase reaction . This method is especially suited for the assay of a large number of samples necessary in enzyme kinetic studies . 2 . From initial rate data the possibility could be excluded that the two-substrate reactions occurs by a random or a ping-pong mechanism . 3 . Investigations of product inhibition exclude a rapid random mechanism but favour an ordered mechanism with UDP-N-acetylglucosamine as the first substrate . 4 . This result was confirmed by isotope exchange studies.

Gene, 1981 Dec, 16(1-3), 325 - 9
Variation in frequency of transformation by plasmid YRp7 in Saccharomyces cerevisiae; Johnston J et al.; Hybrid plasmid YRp7 (Escherichia coli plasmid pBR322 carrying a 1.4-kb yeast fragment containing the TRP1 gene) transforms two related haploid yeast strains (trp1) to Trp+ at frequencies per microgram DNA varying by two orders of magnitude . The diploid cross of these two strains is transformed at a frequency less than that of the low-frequency parent, indicating that high-frequency transformation is inherited in a recessive fashion . Segregant strains of two tetrads of this cross showed intermediate levels of transformability, suggesting the polygenic inheritance of transformation frequency . Levels of variation are changed extensively when frequencies are expressed as transformants per regenerated spheroplast colony, although the inheritance of higher frequency remains clearly recessive . Storage of spheroplast-DNA preparations at 4 degrees C increases the yield of transformants with some strains.

Gene, 1981 Dec, 16(1-3), 133 - 9
Sequence variation in the LEU2 region of the saccharomyces cerevisiae genome; Dobson MJ et al.; The LEU2 regions present on yeast plasmid vectors come from two sources, a series of strains derived from S288c and strain M127 . The LEU2 region from the S288c series contains a Tyl-17 element with its associated delta sequences and a small repetitive RNA gene while the LEU2 region from M127 which is present on pJDB248, lacks the Tyl-17 element, but carries a delta sequence and a small RNA gene . The various LEU2 plasmids currently in use vary with respect to these sequences depending on which restriction fragment from the region is present on the recombinant molecule . In addition, strain M127 contains three LEU2 homologous sequences that are represented by different EcoRI fragments and which segregate independently at meiosis . Therefore, there are at least four forms of the centromere-distal EcoRI fragment of the LEU2 locus in the Saccharomyces cerevisiae gene pool; these are 7.1 kb, 1.9 kb, 1.48 kb and 1.15 kb long.

Eur J Biochem, 1981 Dec, 120(3), 541 - 6
Purification and properties of an endonuclease from the mitochondrion of Saccharomyces cerevisiae; Rosamond J; An endonuclease, which is found only in the mitochondrion of the yeast Saccharomyces cerevisiae, has been purified . The protein has a sedimentation coefficient of 6.3 S, equivalent to a molecular weight of 105,000 . The enzyme is active at pH 7.6, when it degrades single-stranded DNA about 10-times faster than double-stranded DNA, but at pH 5.4 only double-stranded DNA is degraded . In both cases the enzyme acts endonucleolytically, breaking a single phosphodiester bond at a random location within the DNA substrate . Mn2+ or Mg2+ are required for activity; Ca2+ and Zn2+ are ineffective cofactors . Enzyme activity at pH 7.6 is severely inhibited by low concentrations of NaCl or KCl, while activity at pH 5.4 is unaffected by salt . Ethidium bromide inhibits both the DNase activity at pH 5.4 and the activity with single-stranded DNA at pH 7.6, but has no effect on the DNase activity with double-stranded DNA at pH 7.6.

J Bacteriol, 1981 Dec, 148(3), 919 - 25
Adenosine 3',5'-phosphate phosphodiesterase and pheromone response in the yeast Saccharomyces cerevisiae; Liao HH et al.; Theophylline, aminophylline, and isobutylmethylxanthine, compounds reported to be inhibitors of adenosine 3',5'-phosphate (cAMP) phosphodiesterase, prevented the alpha-factor-induced cell cycle arrest of Saccharomyces cerevisiae a cells . To determine whether the in vivo effect of these methylxanthines on yeast pheromone response was related to their known biochemical mode of action, two assays for cAMP phosphodiesterase based on affinity of the product of the reaction (5'-AMP) for boronate groups were developed and were used to monitor the activity of the low Km cAMP phosphodiesterase present in yeast extracts . It was found that the relative efficacy of the methylxanthines as inhibitors of this enzyme in vitro was correlated with the degree to which they antagonized alpha-factor action in vivo . These results were consistent with our previous proposal that pheromone action involves a lowering of cAMP level in the target cell.

Biochim Biophys Acta, 1981 Nov 20, 649(1), 83 - 8
Uptake and accumulation of Mn2+ and Sr2+ in Saccharomyces cerevisiae; Nieuwenhuis BJ et al.; Initial uptake of Mn2+ and Sr2+ in the yeast Saccharomyces cerevisiae was studied in order to investigate the selectivity of the divalent cation uptake system and the possible involvement of the plasma-membrane ATPase in this uptake . The initial uptake rates of the two ions were not significantly different . This ruled out a direct role of the plasma-membrane ATPase, since this ATPase is specific for Mn2+ compared to Sr2+ . After 1 h uptake, Mn2+ had accumulated 10-times more than Sr2+ . Influx of Mn2+ and Sr2+ remained unchanged during that time, however . The differences in accumulation level found for Mn2+ and Sr2+ could be ascribed to a greater efflux of Sr2+ as compared with Mn2+ . Probably this greater efflux of Sr2+ was only apparent, since differential extraction of the yeast cells revealed that Mn2+ is more compartmentalised than Sr2+, giving rise to a lower relative cytoplasmic Mn2+ concentration.

J Biol Chem, 1981 Nov 10, 256(21), 10859 - 63
Properties of H+-translocating adenosine triphosphatase in vacuolar membranes of SAccharomyces cerevisiae; Kakinuma Y et al.; The properties of Mg2+-ATPase in the vacuole of Saccharomyces cerevisiae were studied, using purified intact vacuoles and right-side-out vacuolar membrane vesicles prepared by the method of Y . Ohsumi and Y . Anraku ((1981) J . Biol . Chem . 256, 2079) . The enzyme requires Mg2+ ion but not Ca2+ in . Cu2+ and Zn2+ ions inhibit the activity . The optimal pH is at pH 7.0 . The enzyme hydrolyzes ATP, GTP, UTP, and CTP in this order and the Km value for ATP was determined as 0.2 mM . It does not hydrolyze ADP, adenosyl-5'-yl imidodiphosphate, or p-nitrophenyl phosphate . ADP does not inhibit hydrolysis of ATP by the enzyme . The activities of intact vacuoles and of vacuolar membrane vesicles were stimulated 3- and 1.5-fold, respectively, by the protonophore uncoupler 3,5-di-tert-butyl-4-hydroxybenzilidenemalononitrile and the K+/H+ antiporter ionophore nigericin . Sodium azide at a concentration exerting an uncoupler effect also stimulated the activity . The activity was sensitive to the ATPase inhibitor N,N'-dicyclohexylcarbodiimide, but not to sodium vanadate . The ATP-dependent formation of an electrochemical potential difference of protons, measured by the flow-dialysis method, was determined as 180 mV, with contribution of 1.7 pH units, interior acid, and of a membrane potential of 75 mV . It is concluded that the Mg2+-ATPase of vacuoles is a new marker enzyme for these organelles and is a N,N'-dicyclohexylcarbodiimide-sensitive, H+-translocating ATPase whose catalytic site is exposed to the cytoplasm.

Mutat Res, 1981 Nov, 84(1), 17 - 27
Induction of genetic changes in Saccharomyces cerevisiae by partial drying in air of constant relative humidity and by UV; Hieda K; It was investigated whether there was a critical degree of dryness for induction of genetic changes by drying . Saccharomyces cerevisiae cells were dried in air of 0, 33, 53 and 76% relative humidity (RH) . The frequencies of mitotic recombination at ade2, of gene conversion at leu1, and of gene mutation at can1 were measured in X2447, XS1473 and S288C strains, respectively . After the cells had been dried at 0% RH for 4 h the frequencies of the genetic changes at ade2, leu1 and can1 were, respectively, 56, 7 and 3.5 times higher than each spontaneous frequency . Induction rates, defined as the frequencies of the induced genetic changes per unit time (1 h) of drying, were greatly decreased with increase in RH . Partial drying in air of 76% RH up to 4 and 8 h induced no genetic change at ade2 and leu1, respectively . It was concluded, therefore, that drying at a certain RH between 53 and 76% gave the critical degree of dryness of cells for the induction of the genetic changes . The water contents of cells (g water per g dry material) were 12% at 53% RH and 21% at 76% RH, whereas the water content of native cells was 212% . Removal of a large amount of cellular water had no effect on the induction of the genetic changes . UV sensitivity of partially dried cells of X2447 for the induction of the genetic change at ade2 drastically increased with decrease in RH between 76 and 53% . The drastic change in the UV sensitivity suggested that photochemical reactivity of DNA of chromosome XV, in which the ade2 locus is located, changed between 76 and 53% RH . It seems that the genetic changes were induced only in the low RH region where DNA in vivo had a different photochemical reactivity.

Cell, 1981 Nov, 27(1 Pt 2), 15 - 23
The sequence of the DNAs coding for the mating-type loci of Saccharomyces cerevisiae; Astell CR et al.; The complete sequences of the yeast a mating-type locus, MATa, and of the silent alpha cassette, HML alpha, have been determined . A segment of 642 nucleotides is unique to MATa, and a corresponding segment of 747 nucleotides is unique to MAT alpha . The major mRNAs (a1, a2, alpha 1 and alpha 2) encoded by MATa and MAT alpha have been aligned with the DNA sequence . The a1 mRNA is encoded entirely within the a-specific DNA sequence . The a2 mRNA, which is transcribed divergently from a1 mRNA, is encoded in a region common to both Mata and Mat alpha . The alpha 1 and alpha 2 mRNAs are also transcribed divergently and have their 5' starts about 240 nucleotides apart within the alpha-specific sequence . The amino acid sequences of the MAT proteins have been predicted from the DNA sequences . An unanticipated conclusion is that the a1 protein, containing 148 amino acids, results from readthrough of a UGA at codon 45 . Polymorphic forms of the homologous outer segments of the HML alpha, MAT alpha, MATa and HMRa sequences suggest that the boundaries of the segments involved in mating-type switching are immediately adjacent to the a-specific and alpha-specific sequences.

J Bacteriol, 1981 Nov, 148(2), 659 - 69
Proteolytically induced changes in the molecular form of the carbamyl phosphate synthetase-uracil-aspartate transcarbamylase complex coded for by the URA2 locus in Saccharomyces cerevisiae; Denis-Duphil M et al.; When a uracil-auxotrophic yeast strain is grown under uracil-limiting conditions, the aspartate transcarbamylase activity found in crude extracts shows a variation in sensitivity to feedback inhibition by uridine 5'-triphosphate . In this study we correlated this variation with changes in the molecular form of the carbamyl phosphate synthetase-uracil-aspartate transcarbamylase complex . Carbamyl phosphate synthetase-uracil (molecular weight, 240,000) and uridine 5'-triphosphate-insensitive aspartate transcarbamylase (molecular weight, 140,000) were present separately in extracts from cells collected in the early exponential phase; this was in contrast to the presence of a single high-molecular-weight form (molecular weight, about 900,000) bearing both activities in extracts from stationary-phase cells . The lack of sensitivity to uridine 5'-triphosphate by aspartate transcarbamylase was delayed by adding uridine 5'-triphosphate before cell disruption and was prevented completely by adding phenylmethylsulfonyl fluoride . Thus, this event was attributed to a transient serine protease activity detected only in early exponential-phase cell extracts . However, even in the presence of phenylmethylsulfonyl fluoride, a sucrose density gradient analysis in the absence of uridine 5'-triphosphate revealed a change in the aggregation state of the complex which might have occurred in vivo . None of these events was observed in extracts from cells that lacked protease B activity (strain HP232-2B).

J Bacteriol, 1981 Nov, 148(2), 618 - 23
Incision and postincision steps of pyrimidine dimer removal in excision-defective mutants of Saccharomyces cerevisiae; Wilcox DR et al.; cdc9, a temperature-sensitive mutant defective in polynucleotide deoxyribonucleic acid (DNA) ligase activity, accumulates low-molecular-weight DNA fragments (as measured by sedimentation of DNA in alkaline sucrose gradients) at the nonpermissive temperature after irradiation with ultraviolet light . This phenotype of cdc9 is a sensitive indicator of successful incision during excision repair of dimers . In strains containing excision-defective mutations in any of nine genes in combination with the cdc9 mutation, the absence of low-molecular-weight DNA at the nonpermissive temperature after ultraviolet treatment suggests that these mutants are incision defective, whereas the presence of low-molecular-weight DNA indicates that the mutants are defective in a step after incision . With rad1, rad2, rad3, rad4, and rad10 mutants, the molecular weight of the DNA remained unchanged after ultraviolet irradiation and incubation at the restrictive temperature, despite the presence of the cdc9 mutation; these mutants are therefore incision defective . Low-molecular-weight DNA was observed in rad14 cdc9 and rad16 cdc9 strains . With the rad16 strain, the accumulation of low-molecular-weight DNA correlated with the amount of excision taking place, whereas in the rad14 mutant strain, no evidence of dimer removal was obtained . Therefore, rad14 is likely to be defective in a step after incision.

Can J Microbiol, 1981 Nov, 27(11), 1140 - 9
Solubilization of microsomal-associated phosphatidylserine synthase and phosphatidylinositol synthase from Saccharomyces cerevisiae; Carman GM et al.; Membrane-associated cytidine 5'-diphospho-1,2-diacyl-sn-glycerol (CDP-diacylglycerol):L-serine O-phosphatidyltransferase (phosphatidylserine synthase, EC2.7.8.8.) and CDP-diacylglycerol:myo-inositol phosphatidyltransferase (phosphatidylinositol synthase, EC 2.7.8.11) were solubilized from the microsomal fraction of Saccharomyces cerevisiae . A variety of detergents were examined for their ability to release phosphatidylserine synthase and phosphatidylinositol synthase activities from the microsome fraction . Both enzymes were solubilized from the microsome fraction with Renex 690 in yield over 80% with increase to specific activity of 1.6-fold . Both solubilized enzymatic activities were dependent on manganese ions and Triton X-100 for maximum activity . The pH optimum for each reaction was 8.0 . The apparent Km values for CDP-diacylglycerol and serine for the phosphatidylserine synthase reaction were 0.1 and 0.25 mM, respectively . The apparent Km values for CDP-diacylglycerol and inositol for the phosphatidylinositol synthase reaction were 70 microM and 0.1 mM, respectively . Thioreactive agents inhibited both enzymatic activities . Both solubilized enzymatic activities were thermally inactivated at temperatures above 30 degrees C.

J Biol Chem, 1981 Oct 25, 256(20), 10420 - 5
Effect of yeast killer toxin on sensitive cells of Saccharomyces cerevisiae; de la Pena P et al.; Killer toxin from Saccharomyces cerevisiae inhibited the pumping of protons into the medium by metabolically active sensitive cells . Such inhibition coincided with that of the uptake of potassium ions which are thought to be accumulated by yeast cells in order to neutralize the membrane potential created because of the extrusion of protons . The consumption of glucose, however, was identical in killer-treated and untreated cells . These alterations can be explained by the ability of the toxin to reduce the chemical proton gradient across the plasma membrane as measured by the accumulation of the weak permeable {14C}propionic acid . With this method, an internal pH of 6.42 was calculated from normal cells (the external pH was 4.6) while that of toxin-treated cells was decreased as a function of time . The proton concentration gradient was reduced from 66- to 17-fold . It is shown that the toxin-induced alteration of the proton gradient is due to an enhanced proton permeability of the yeast plasma membrane upon binding of the toxin . It is suggested that killer toxin acts as a macromolecular proton conductor similar in some respects to the known proton conductors 2,4-dinitrophenol and carbonyl cyanide m-chlorophenylhydrazone, since all the described effects are also observed with these substances.

J Biol Chem, 1981 Oct 10, 256(19), 9999 - 10004
Protein synthesis in yeast . I . Purification and properties of elongation factor 3 from Saccharomyces cerevisiae; Dasmahapatra B et al.; Elongation factor 3 from the yeast Saccharomyces cerevisiae was purified over 230-fold from a high speed supernatant fraction . The homogeneity of the protein was shown by gel filtration and sedimentation equilibrium analysis of the native protein and by sodium dodecyl sulfate gel electrophoresis of the denatured protein . The molecular weight of the protein was estimated to be 125,000 by the above-mentioned methods . The protein consists of a single polypeptide chain . Amino acid analysis revealed no unusual features . Antibody raised against the purified factor showed a single cross-reacting band with the characteristic hexagonal pattern in an Ouchterlony double diffusion plate . The immune serum had no reactivity against the other two elongation factors (EF) . The polymerization reaction was inhibited by the anti-EF3 . Addition of excess EF3 could overcome this effect . Factor 3 is absolutely required by the yeast ribosomes for polyphenylalanine synthesis . Ribosomes from other eukaryotes do not require this protein . The function of the third factor in polyphenylalanine synthesis cannot be defined at this time . The protein showed ribosome-dependent GTPase and ATPase activities . Studies of partial reactions showed that EF3 was not required for Phe-tRNA binding to ribosomes, peptide bond formation, or translocation . Nucleotide exchange by EF1 was not stimulated by EF3.

Nucleic Acids Res, 1981 Oct 10, 9(19), 5021 - 36
Ribosomal protein genes rp 39(10 - 78), rp 39(11 - 40), rp 51, and rp 52 are not contiguous to other ribosomal protein genes in the Saccharomyces cerevisiae genome; Woolford JL Jr et al.; A library of recombinant phage containing EcoR1 fragments of Saccharomyces cerevisiae DNA has been constructed . This library was screened with four different recombinant plasmids, each containing a different yeast ribosomal protein gene, in order to isolate chromosomal fragments extending in both directions from these genes . These chromosomal fragments were assayed for the presence of additional ribosomal protein genes by hybridization selection and cell free translation, and none were found . These four regions are not closely linked to each other, since DNA from one domain does not cross hybridize with DNA from any of the other three, except for the sequences within the homologous ribosomal protein 39 gene pair . Northern blots demonstrate that although the concentrations of ribosomal protein mRNAs are diminished significantly in a strain containing the ts mutation rna2, transcripts from genes in these flanking segments are relatively unaffected.

J Biol Chem, 1981 Oct 10, 256(19), 10176 - 83
The genes for fifteen ribosomal proteins of Saccharomyces cerevisiae; Fried HM et al.; We have isolated recombinant lambda phage carrying the genes for 14 of the ribosomal proteins of the yeast Saccharomyces cerevisiae . Analysis of these and of the plasmid carrying the gene tcm1, which codes for the ribosomal protein responsible for resistance to trichodermin, demonstrates that in general the genes for ribosomal proteins are unlinked . One exceptional recombinant carries the genes for two ribosomal proteins within a 2-kilobase region . DNA fragments bearing individual ribosomal protein genes were used to probe restriction digests of the yeast genome to determine whether any of the genes were duplicated . Only 3 of 12 of the genes are present unequivocally as a single copy . Similar fragments were used to probe blots of mRNA separated on denaturing agarose gels to determine the size of the mRNA for each protein . In each case, the mRNA is near the minimum size necessary to code for its protein . In certain temperature-sensitive mutants which fail to synthesize functional mRNA for ribosomal protein, Rosbash et al . (Rosbash, M., Harris, P . K . W., Woolford, J., and Teem, J . L . (1981) Cell, 24, 679-686) have demonstrated the accumulation of a larger RNA molecule, homologous to a ribosomal protein gene, that appears to be a transcript which retains an intervening sequence . We find that for 8 of the 11 ribosomal protein genes examined, a larger molecule accumulates in such a mutant strain, suggesting that in general transcripts of ribosomal protein genes may have introns.

Mol Cell Biol, 1981 Oct, 1(10), 958 - 60
rme1 Mutation of Saccharomyces cerevisiae: map position and bypass of mating type locus control of sporulation; Rine J et al.; Sporulation in Saccharomyces cerevisiae normally occurs only in MATa/MAT alpha diploids . We show that mutations in RME1 bypassed the requirements for both a and alpha mating type information in sporulation and therefore allowed MATa/MATa and MAT alpha/MAT alpha diploids to sporulate . RME1 was located on chromosome VII, between LEU1 and ADE6.

Mol Cell Biol, 1981 Oct, 1(10), 910 - 8
Nature and timing of some sporulation-specific protein changes in Saccharomyces cerevisiae; Wright JF et al.; During meiosis and spore formulation in Saccharomyces cerevisiae, changes that occur in a/alpha diploids, but not in isogenic nonsporulating a/a diploids, have been detected in cellular polypeptides . These were found by the technique of prelabeling growing cells with 35SO4(2-) and suspending them in sulfur-free sporulation medium . Under the conditions used, about 400 polypeptides were detected by two-dimensional gel electrophoresis, and 45 were altered during sporulation; of these, 21 changes were specific to a/alpha strains . These alterations were mainly due to the appearance of new polypeptides or to marked increases in the concentrations of a few polypeptides produced during vegetative growth . They could have been due either to modifications of existing polypeptides present in growing cells or to de novo synthesis of new gene products . They occurred at characteristic times during sporulation; whereas the majority of changes took place early (within the first 6 h in sporulation conditions), there were several changes characterizing the later stages of sporulation . Ten of the 35SO4(2-)-labeled polypeptides were also labeled with 32P in the presence of {32P}orthophosphate; of these, three were previously found to be sporulation specific . One of these was phosphorylated at all stages of sporulation and was labeled when {32P}orthophosphate was added either during growth of the culture of 1 h after transfer to sporulation medium . Another was labeled in the same way by adding 32P at either time, so that by 7 h in sporulation medium it was phosphorylated, but was dephosphorylated by 24 h . The third sporulation-specific peptide was labeled in extracts prepared at 7 h in sporulation medium (but not at 24 h) when {32P}-orthophosphate was added during presporulation growth, but not when {32P}-orthophosphate was added 1 h after transfer of the culture to sporulation medium . This polypeptide appeared early during sporulation; it is probably phosphorylated as it appears and is dephosphorylated at some time between 7 h and 24 h of sporulation.

Mol Cell Biol, 1981 Oct, 1(10), 891 - 901
Recombinationless meiosis in Saccharomyces cerevisiae; Malone RE et al.; We have utilized the single equational meiotic division conferred by the spo13-1 mutation of Saccharomyces cerevisiae (S . Klapholtz and R . E . Esposito, Genetics 96:589-611, 1980) as a technique to study the genetic control of meiotic recombination and to analyze the meiotic effects of several radiation-sensitive mutations (rad6-1, rad50-1, and rad52-1) which have been reported to reduce meiotic recombination (Game et al., Genetics 94:51-68, 1980); Prakash et al., Genetics 94:31-50, 1980) . The spo13-1 mutation eliminates the meiosis I reductional segregation, but does not significantly affect other meiotic events (including recombination) . Because of the unique meiosis it confers, the spo13-1 mutation provides an opportunity to recover viable meiotic products in a Rec- background . In contrast to the single rad50-1 mutant, we found that the double rad50-1 spo13-1 mutant produced viable ascospores after meiosis and sporulation . These spores were nonrecombinant: meiotic crossing-over was reduced at least 150-fold, and no increase in meiotic gene conversion was observed over mitotic background levels . The rad50-1 mutation did not, however, confer a Rec- phenotype in mitosis; rather, it increased both spontaneous crossing-over and gene conversion . The spore inviability conferred by the single rad6-1 and rad52-1 mutations was not eliminated by the presence of the spo13-1 mutation . Thus, only the rad50 gene has been unambiguously identified by analysis of viable meiotic ascospores as a component of the meiotic recombination system.

Genetics, 1981 Oct, 99(2), 197 - 209
Selective abortion of two nonsister nuclei in a developing ascus of the hfd-1 mutant in Saccharomyces cerevisiae; Okamoto S et al.; A recessive mutation, hfd1-1, in strain SOS4 of Saccharomyces cerevisiae leads the mutant cells to produce predominantly two-spored asci . Light microscopical examination of Giemsa-stained cells revealed no significant differences in the meiotic figures between mutant and wild-type strains . However, only two of the four meiotic products in a developing ascus matured to ascospores in SOS4 . Dyad analysis was carried out on an hfd1-1 mutant strain heterozygous for three markers, asp5, gal1, and arg4, which are closely linked to their centromeres, and for his4, which is loosely linked to its centromere . The two-spored asci produced by the hfd1-1 mutant segregated dominant (+) and recessive (-) alleles of each marker in a 1:1 ratio; they generally contained one + and one - spore for any given marker . The occurrence of rare dyads with two + or two - spores can be explained quantitatively by recombination between the marker and its centromere . From the results of these cytological and genetical analyses, we infer that, in the mutant strain, one genome set is partitioned to each of the four second-meiotic division poles, but only two nonsister genomes are incorporated into mature spores . Thus, the hfd1-1 mutation in SOS4 blocks incorporation of two nonsister nuclei into mature ascospores, but does not block enclosure of the remaining two nonsister nuclei.

Eur J Biochem, 1981 Oct, 119(3), 613 - 8
Spectral properties of cytochrome b-561 and cytochrome b-565 in mucidin-resistant mutants of Saccharomyces cerevisiae; Subik J et al.; The oxidation of NADH in submitochondrial particles isolated from MUC1, MUC2 and MUC3 mucidin-resistant mutants of Saccharomyces cerevisiae is specifically resistant to mucidin . Extra reduction of cytochrome b-565 induced by mucidin is demonstrated in all tested mucidin-resistant mutants . Red shift of cytochrome b-561 is induced by mucidin in two independent MUC3 mutants . In MUC1 and MUC2 mutants, the red shift is not induced by mucidin, while that promoted by antimycin A and 2-n-heptyl-4-hydroxyquinoline N-oxide are normal . It is concluded that the extra reduction of cytochrome b-565 and the red shift of cytochrome b-561 elicited by mucidin can be largely dissociated from the overall inhibition of the electron flow by distinct mucidin-resistant mutations in different exons of the split mitochondrial gene of cytochrome b.

J Bacteriol, 1981 Oct, 148(1), 248 - 56
Interactions between mutations for sensitivity to psoralen photoaddition (pso) and to radiation (rad) in Saccharomyces cerevisiae; Henriques JA et al.; The mode of interaction in haploid Saccharomyces cerevisiae of two pso mutations with each other and with rad mutations affected in their excision-resynthesis (rad3), error-prone (rad6), and deoxyribonucleic acid double-strand break (rad52) repair pathways was determined for various double mutant combinations . Survival data for 8-methoxypsoralen photoaddition, 254-nm ultraviolet light and gamma rays are presented . For 8-methoxypsoralen photoaddition, which induces both deoxyribonucleic acid interstrand cross-links and monoadditions, the pso1 mutation is epistatic to the rad6, rad52, and pso2 mutations, whereas it is synergistic to rad3 . The pso2 mutation, which is specifically sensitive to photoaddition of psoralens, is epistatic to rad3 and demonstrates a nonepistatic interaction with rad6 and rad52 . rad3 and rad6, as well as rad 6 and rad52, show synergistic interactions with each other, whereas rad 3 is epistatic to rad52 . Consequently, it is proposed that PSO1 and RAD3 genes govern steps in the independent pathways . The PSO1 activity leading to an intermediate which is repaired via the three incidence pathways controlled by RAD6, RAD52, and PSO2 genes . Since pso1 interacts synergistically with rad3 and rad52 and epistatically with rad6 after UV radiation, the PSO1 gene appears to belong to the RAD6 group . For gamma ray sensitivity, pso1 is epistatic to rad6 and rad52, which suggests that this gene controls a step which is common to the two other independent pathways.

Biochemistry, 1981 Sep 15, 20(19), 5611 - 6
Identification of 3-methoxy-4-hydroxy-5-hexaprenylbenzoic acid as a new intermediate in ubiquinone biosynthesis by Saccharomyces cerevisiae; Goewert RR et al.; A ubiquinone-deficient mutant strain of Saccharomyces cerevisiae, 26H, was found to accumulate a previously unidentified intermediate in ubiquinone biosynthesis when grown in the presence of p-hydroxy{7-14C}- or -{u-14C}benzoic acid . This intermediate was isolated from the lipid extracts of a 100-L culture of 26H and purified by various chromatographic procedures to yield 20 mg of product . Analysis by means of NMR, IR, UV, and mass spectrometry revealed the structure of this new intermediate to be 3-methoxy-4-hydroxy-5-hexaprenylbenzoic acid (3-MHHB) . In vitro experiments with isolated yeast and rat mitochondria showed that 3-MHHB could be converted to ubiquinone-6 . These findings indicate that 3-O-methylation precedes decarboxylation of the prenylated protocatechuic acid intermediate in the biosynthesis of ubiquinone in eukaryotes.

Mikrobiologiia, 1981 Sep-Oct, 50(5), 891 - 7
{Protein yield in the rehydration of dehydrated Saccharomyces cerevisiae yeast cells}; Popova MV et al.; The release of proteins was studied during rehydration of Saccharomyces cerevisiae cells dehydrated using two techniques which were characterized by different survival rate . A correlation was established between the loss of proteins and the survival rate of cells after dehydration and rehydration . Proteins with a molecular mass from 6000 to 70,000 daltons (low molecular weight proteins predominating) were found in the rehydration medium by the method of disc-electrophoresis in polyacrylamide gel in the presence of sodium dodecylsulfate . Moreover, the rehydration medium exhibited the activity of certain phosphohydrolases and dehydrogenases; this finding suggests that proteins with a higher molecular weight are present in the medium.

Z Naturforsch {C}, 1981 Sep-Oct, 36(9-10), 798 - 803
The effect of clotrimazole and triadimefon on 3-hydroxy-3-methyl-glutaryl-CoA-reductase-{EC 1.1.1.34}-activity in Saccharomyces cerevisiae; Berg D et al.; Clotrimazole and triadimefon are known as potent inhibitors of ergosterol synthesis in pathogenic yeast and fungi, respectively . As their mode of action generally the inhibition of sterol desmethylation reactions is accepted . We report about a second effect, a "feed-back" inhibition of 3-hydroxy-3-methyl-glutaryl (HMG)-CoA-reductase by accumulation of ergosterol precursors . Addition of lanosterol to intact cells leads to an inhibition of HMG-CoA-reductase as well, but not to fungistatic effects . From the reported data the influences of clotrimazole and triadimefon have to be considered as an inhibition of desmethylation reactions involved in ergosterol synthesis of yeast and fungi with a concomitant decreased production of mevalonate.

Proc Natl Acad Sci U S A, 1981 Sep, 78(9), 5778 - 82
Mitotic chromosome loss in a radiation-sensitive strain of the yeast Saccharomyces cerevisiae; Mortimer RK et al.; Cells of Saccharomyces cerevisiae with mutations in the RAD52 gene have previously been shown to be defective in meiotic and mitotic recombination, in sporulation, and in repair of radiation-induced damage to DNA . In this study we show that diploid cells homozygous for rad52 lose chromosomes at high frequencies and that these frequencies of loss can be increased dramatically by exposure of these cells to x-rays . Genetic analyses of survivors of x-ray treatment demonstrate that chromosome loss events result in the conversion of diploid cells to cells with near-haploid chromosome numbers.

Z Naturforsch {C}, 1981 Sep-Oct, 36(9-10), 813 - 9
Inhibition cyclic guanosine 3':5'-monophosphate of the soluble DNA polymerase activity, and of partially purified DNA polymerase A (DNA polymerase I) from the yeast Saccharomyces cerevisiae; Eckstein H; DNA polymerase activity from extracts of growing yeast cells is inhibited by cGMP . Experiments with partially purified yeast DNA polymerases show, that cGMP inhibits DNA polymerase A (DNA polymerase I from Chang), which is the main component of the soluble DNA polymerase activity in yeast extracts, by competing for the enzyme with the primer-template DNA . Since the enzyme is not only inhibited by 3', 5'-cGMP, but also by 3',5'-cAMP, the 3'--:5'-phosphodiester seems to be crucial for the competition between cGMP and primer . This would be inconsistent with the concept of a 3'-OH primer binding site in the enzyme . The existence of such a site in the yeast DNA polymerase A is indicated from studies with various purine nucleoside monophosphates . When various DNA polymerases are compared, inhibition by cGMP seems to be restricted to those enzymes, which are involved in DNA replication, DNA polymerases with an associated nuclease activity are not inhibited, DNA polymerase B from yeast is even activated by cGMP . Though some relations between the cGMP effect and the presumed function of the enzymes in the living cell are apparent, the biological meaning of the observations in general remains open.

Gene, 1981 Sep, 14(4), 263 - 78
Molecular cloning of the SUF2 frameshift suppressor gene from Saccharomyces cerevisiae; Cummins CM et al.; A genetic approach to the molecular cloning of frameshift suppressor genes from yeast is described . These suppressors act by suppressing +1 G:C base-pair insertion mutations in glycine or proline codons . The cloning regimen involves an indirect screen for yeast transformants which harbor a functional suppressor gene inserted into the autonomously replicating "shuttle" vector YEp13, followed by transfer of the hybrid plasmid from yeast into Escherichia coli . Using this procedure a 10.7-kb DNA fragment carrying the SUF2 frameshift suppressor gene has been isolated . This suppressor acts specifically on +1 G:C insertions in proline codons . When inserted into an integrative vehicle and reintroduced into yeast by transformation, this fragment integrates by homologous recombination in the region of the SUF2 locus on chromosome III . A large proportion of the fragment overlaps with another cloned DNA segment which carries the closely linked CDC10 gene . The SUF2 fragment carries at least two tRNA genes . The SUF2 gene and one of the tRNA genes are located on a 0.85-kb restriction fragment within the 10.7-kb segment . A method is also described for the isolation of DNA fragments carrying alternative alleles of the SUF2 locus . Using this procedure, the wild-type suf2+ allele has been cloned.

Biochem J, 1981 Aug 15, 198(2), 281 - 7
Glucose-induced inactivation of mitochondrial enzymes in the yeast Saccharomyces cerevisiae; Takeda M; 1 . Addition of glucose induced an inactivation of mitochondrial enzymes in the yeast Saccharomyces cerevisiae containing normal mitochondrial particles . 2 . The glucose-induced inactivation of mitochondrial enzymes was inhibited by the presence of cycloheximide . 3 . Pepstatin also inhibited the inactivation, but phenylmethanesulphonyl fluoride accelerated the inactivation . 4 . The specific activities of fructose 1,6-bisphosphatase and cytoplasmic malate dehydrogenase were decreased on the exposure to glucose, as well as those of the mitochondrial enzymes . However, the glucose-induced inactivation of cytoplasmic enzymes was not inhibited by the presence of pepstatin . 5 . The specific activities of hexokinase and phosphofructokinase, which are cytoplasmic enzymes were increased by the addition of glucose, and this effect was not affected by pepstatin . 6 . Addition of glucose resulted in an increase in the synthesis of proteins of the mitochondria and the cytosol, and simultaneously in degradation of these mitochondrial and cytoplasmic proteins.

Biochemistry, 1981 Aug 4, 20(16), 4584 - 90
Purification and characterization of hypoxanthine-guanine phosphoribosyltransferase from Saccharomyces cerevisiae; Nussbaum RL et al.; Hypoxanthine-guanine phosphoribosyltransferase (HPRT) was purified 12 000-fold to homogeneity from yeast by a three-step procedure including acid precipitation, anion-exchange chromatography, and guanosine 5' -monophosphate affinity chromatography . The enzyme is a dimer consisting of two, probably identical, subunits of Mr 29 500 . The enzyme recognized hypoxanthine and guanine, but not adenine or xanthine, as substrates . An antiserum against both native and denatured enzyme has been raised and shown to be specific for the enzyme . The antiserum has no affinity for Chinese hamster or human HPRT but does recognize subunits of yeast HPRT as well as some cyanogen bromide fragments of the enzyme.

Genetics, 1981 Aug, 98(4), 691 - 711
Agents that cause a high frequency of genetic change from {psi+} to {psi-} in Saccharomyces cerevisiae; Tuite MF et al.; The {psi} factor of yeast is cytoplasmically inherited . Singh, Helms and Sherman (1979) reported that high concentrations of KCl and of ethylene glycol induce the genetic change from {psi+} to {psi-} . In this study, the following agents have been shown to induce the same genetic change: guanidine hydrochloride at 1 mM, dimethyl sulfoxide at 2.5% v/v and ethanol or methanol at 10% v/v . It is likely that a number of other agents also cause the change, namely 2 M glycerol, M succinate, M glutamate and M MgCl2 . Most of these agents induce the change at very high frequencies; with some, the frequency is 100% . Although the observed phenotypic change can also occur as a result of chromosomal gene mutation, no changes of this type were identified . Some of the agents also cause mutation from {rho+} to {rho-} and from killer to sensitive.

J Cell Sci, 1981 Aug, 50, 361 - 76
Variability in individual cell cycles of Saccharomyces cerevisiae; Lord PG et al.; The kinetics of cell proliferation of Saccharomyces cerevisiae were studied at 4 growth rates using time-lapse cinephotomicrography . Cells were grown on media with a high refractive index to reveal greater intracellular detail under the phase-contrast microscope . The morphological cell-cycle events scored were: bud emergence, nuclear migration, nuclear division, onset of cytokinesis and cell separation . Cell size was measured at cell separation and at bud emergence . The daughter-cycle time was always longer than the parent-cycle time mainly due to the large difference in the lengths of the unbudded phases . Parent cells had a shorter budded period than daughter cells . The large variance in daughter-cycle times was accounted for by the large variance in the lengths of the unbudded phase of daughter cells . The duration and variability of the periods in the cyclc from nuclear migration onwards were equivalent for parent and daughter cells . Daughter cells were always smaller than parent cells at division . There was wide variation in cell size at both division and bud emergence . The results indicated that a modified deterministic model could best explain cell proliferation kinetics in yeast . The data were used to evaluate 2 different models . The 'sloppy size control' model of Wheals (1981 a) was more consistent with the data than the 'tandem' model of Shilo, Shilo & Simchen (1976) . The distribution of unbudded periods of daughter cells suggested that there was an additional incompressible period not present in parent cells.

Can J Microbiol, 1981 Aug, 27(8), 847 - 49
Radioimmunoassay for yeast killer toxin from Saccharomyces cerevisiae; Siddiqui FA et al.; A radioimmunoassay was developed for the K1 killer toxin from strain T158C/S14a of Saccharomyces cerevisiae . 125I-labeled toxin was made to a specific activity of 100 microCi/mg of protein (1 microCi = 37 kBq) . Antibody to purified toxin was prepared in rabbits using toxin cross-linked to itself . These antibodies, partially purified by 50% ammonium sulfate precipitation and Sepharose CL-6B column chromatography, produced one precipitation band with killer toxin and bound 125I-labeled toxin in a radioimmunoassay . The antibody preparation also bound with the toxins from another K1 killer, A364A, and three chromosomal superkiller mutants derived from it.

J Bacteriol, 1981 Aug, 147(2), 705 - 8
Molecular mechanisms of pyrimidine dimer excision in Saccharomyces cerevisiae: excision of dimers in cell extracts; Reynolds RJ et al.; Cell-free extracts prepared from rad1-19, rd2-2, rad3-1, rad4-3, rad7-1, rad10-1, rd14-1, rad16-1, and cyc1-1 (rad7) mutants of Saccharomyces cerevisiae all catalyze the preferential excision of thymine-containing pyrimidine dimers from ultraviolet-irradiated DNA specifically incised with M . luteus ultraviolet deoxyribonucleic acid incising activity.

J Bacteriol, 1981 Aug, 147(2), 517 - 25
Spontaneous mitotic recombination in mms8-1, an allele of the CDC9 gene of Saccharomyces cerevisiae; Montelone BA et al.; The methyl methane sulfonate (MMS)-sensitive mutation mms8-1 increases the rate of spontaneous mitotic intragenic recombination at five heteroallelic loci on three chromosomes . Complementation, segregation, and mapping studies indicate that mms8-1 is allelic to cdc9, known to be defective in deoxyribonucleic acid ligase . Both mms8-1 and cdc9 mutants are lethal in combination with the recombination-defective mutant rad52-1 . Genetic analysis of spontaneous red/white sectors in an ade2-1/ade2-1 ade5/+ mms8-1/mms8-1 strain shows nonreciprocal recombinational events involving long chromosome segments . We also observe greater than expected rates of simultaneous recombination at loci on different chromosomes in both wild-type and mms8-1 mutants.

Proc Natl Acad Sci U S A, 1981 Aug, 78(8), 4743 - 7
A mitochondrial locus is necessary for the synthesis of mitochondrial tRNA in the yeast Saccharomyces cerevisiae; Martin NC et al.; We have used a cloned yeast mitochondrial tRNAUCNSer gene as a probe to detect RNA species that are transcripts from this gene in wild-type Saccharomyces cerevisiae and in petite deletion mutants . In RNA from wild-type cells, the tRNA is the most prominent transcript of the gene . In RNA from deletion mutants that retain this gene but have lost other regions of mtDNA, high molecular weight transcripts containing the tRNAUCNSer sequences accumulate but tRNAUCNSer is not made . tRNAUCNSer synthesis can be restored in these mutants when they are mated to other deletion mutants that retain a different portion of the mitochondrial genome . Protein synthesis is not necessary for the restoration, and the restoration is not due to a nuclear effect or to an effect of mating alone, because strains without mtDNA are not able to restore tRNA synthesis . These results definitively demonstrate the existence of a yeast mitochondrial locus that is necessary for tRNA synthesis and, because the restoration of tRNAUCNSer synthesis appears to result from intergenic complementation, not recombination, indicate that this locus acts in trans.

Biochim Biophys Acta, 1981 Jul 20, 645(2), 226 - 8
Photoinactivation of the thiamine transport system in Saccharomyces cerevisiae with 4-azido-2-nitrobenzoylthiamine; Sempuku K et al.; A newly synthesized photoreactive thiamine derivative, 4-azido-2-nitrobenzoylthiamine was found to be a competitive inhibitor of the thiamine transport system in Saccharomyces cerevisiae, exhibiting an apparent Ki of 36 nM . When exposed to visible light, 4-azido-2-nitrobenzoylthiamine irreversibly inactivated the thiamine transport . 4-Azido-2-nitrobenzoylthiamine-dependent photoinactivation of thiamine transport was partially protected by thiamine, but not by the nitrene-trapping reagent p-aminobenzoate . On the other hand, the irradiation of the yeast cells in the presence of 4-azido-2-nitrobenzoylthiamine did not significantly lead to inactivation of the biotin transport system . The results suggest that 4-azido-2-nitrobenzoylthiamine is a specific irreversible inhibitor of the thiamine transport system in Saccharomyces cerevisiae.

Mol Cell Biol, 1981 Jul, 1(7), 584 - 93
Biological role of the general control of amino acid biosynthesis in Saccharomyces cerevisiae; Niederberger P et al.; The biological role of the "general control of amino acid biosynthesis" has been investigated by analyzing growth and enzyme levels in wild-type, bradytrophic, and nonderepressing mutant strains of Saccharomyces cerevisiae . Amino acid limitation was achieved by using either bradytrophic mutations or external amino acid imbalance . In the wild-type strain noncoordinate derepression of enzymes subject to the general control has been found . Derepressing factors were in the order of 2 to 4 in bradytrophic mutant strains grown under limiting conditions and only in the order of 1.5 to 2 under the influence of external amino acid imbalance . Nonderepressing mutations led to slower growth rates under conditions of amino acid limitation, and no derepression of enzymes under the general control was observed . The amino acid pools were found to be very similar in the wild type and in nonderepressing mutant strains under all conditions tested . Our results indicate that the general control affects all branched amino acid biosynthetic pathways, namely, those of the aromatic amino acids and the aspartate family, the pathways for the basic amino acids lysine, histidine, and arginine, and also the pathways of serine and valine biosyntheses.

Farmaco {Sci}, 1981 Jul, 36(7), 492 - 505
Mutagenic activity of three monofunctional and three bifunctional furocoumarins in yeast (Saccharomyces cerevisiae); Averbeck D et al.; The mutagenic activity of photoadditions induced by 4,5'-dimethylangelicin (4,5'-DMA), 3-carbethoxypsoralen (3-CPs), angelicin, 8-methoxypsoralen (8-MOP), 5-methoxypsoralen (5-MOP) and 4,5',8-trimethylpsoralen (TMP), was studied in haploid yeast cells at the nuclear and cytoplasmic levels . With regard to the induction of cell killing 4,5'-DMA and 3-CPs were about 5 to 6 times more active than 8-MOP probably due to an efficient repair of 4,5'-DMA or 3'CPs induced monoadditions in DNA . 4,5'-DMA was about 5-fold less active than angelicin, reflecting the different photoreactivities of the compounds towards DNA . In accord with its monofunctional activity, 4,5'-DMA was less efficient on the induction of nuclear reversions but more efficient on the induction of cytoplasmic "petite" mutants per viable cell than 8-MOP and the other bifunctional compounds . In contrast to 3-CPs and angelicin, the reversion induction by 4,5'-DMA followed a 2 hit kinetics . For the induction of CanR forward mutants per viable cell 4,5'-DMA was more efficient than 3-CPs and approached the activity of 8-MOP; angelicin, 5-MOP and TMP were slightly more efficient than 8-MOP, whereas 3-CPs was clearly less efficient . Thus, the monofunctional furocoumarins exert different genotoxic effects when compared on a survival basis . Cell killing effects in the presence and in the absence of oxygen were also evaluated.

Eur J Biochem, 1981 Jul, 117(2), 333 - 9
Modification of the spectral properties of cytochrome b in mutants of Saccharomyces cerevisiae resistant to 3-(3,4-dichlorophenyl)-1,1-dimethylurea . Mapping at two distinct genetic loci of the split mitochondrial gene of cytochrome b; Briquet M et al.; The effects of five inhibitors of the cytochrome bc1 complex: 3-(3,4-dichlorophenyl)-1,1-dimethylurea (diuron), 2-n-heptyl-4-hydroxyquinoline-N-oxide (HpHOQnO), antimycin A, funiculosin and mucidin were measured in submitochondrial particles of strains of the yeast Saccharomyces cerevisiae belonging to two classes of diuron-resistant mutants Diu 1 and Diu 2 which are modified in different exons of the split mitochondrial gene of cytochrome b . 1 . The oxidation of NADH and of cytochrome b-561 exhibits a similar resistance to diuron and HpHOQnO in Diu 1 and Diu 2 mutants . 2 . No extra reduction of cytochrome b-561 and cytochrome b-565 is observed in the presence of diuron and HpHOQnO . 3 . Both Diu 1 and Diu 2 mutants exhibit the red shift of cytochrome b-561 induced by concentrations of HpHOQno 2 -- 3-times higher than those required in the parental strains . 4 . The spectral and respiratory effects of antimycin A, funiculosin and mucidin and generally similar in the diuron-resistant mutants and in their parental strains . However a cross-resistance between diuron and antimycin A is indicated in one Diu 2 mutant . 5 . From the combined genetic and biochemical data it is concluded that the interaction of diuron and HpHOQnO with cytochrome b is mediated by at least two specific amino acids located apart in the central region of the apocytochrome b peptide coded by mitochondrial DNA . These two amino acids control tightly the extra reduction of cytochromes b-565 and b-561 as well as the flow of electrons through the bc1 complex . However the binding of HpHOQnO required for the expression of the red shift of cytochrome b-561 is only slightly affected by the diu-1 and diu-2 mutations.

Biochim Biophys Acta, 1981 Jun 26, 654(1), 149 - 55
Further characterization of recessive suppression in yeast . Isolation of the low-temperature sensitive mutant of Saccharomyces cerevisiae defective in the assembly of 60 S ribosomal subunit; Surguchov AP et al.; It has been shown that recessive suppressor mutations in the yeast Saccharomyces cerevisiae may cause sensitivity towards low temperatures (very slow growth or lack of growth at 10 degrees C) . One of the sup 1 low temperature sensitive (Lts-) mutants, 26-125A-P-2156, was studied in detail . After a prolonged period of incubation (70 h) under restrictive conditions the protein synthesis apparatus in the mutant cells was irreversibly damaged . In addition, Lts- cells incubated under restrictive conditions synthesize unequal amounts of ribosomal subunits, the level of 60 S subunit being reduced . It has been suggested that the recessive suppression is mediated by a mutation in the gene coding for 60 S subunit component, probably a ribosomal protein . The mutation leads simultaneously to a defect in the assembly of 60 S subunit and to low-temperature sensitive growth of the mutant.

Biochim Biophys Acta, 1981 Jun 12, 636(1), 104 - 12
Biogenesis of mitochondria . Defective assembly of the proteolipid into the mitochondrial adenosine triphosphatase complex in an oli2 mit- mutant of Saccharomyces cerevisiae; Stephenson G et al.; A single mutation in the oli2 region of the mitochondrial DNA causes a charge alteration in a mitochondrially translated subunit of the mitochondrial ATPase (subunit 6; apparent Mr 20 000; apparent pI 6.9 and 7.1) . This alteration leads to the defective assembly of the proteolipid subunit into the enzyme complex . The mutant, which is able to grow only very slowly by oxidative metabolism at 28 degrees C offers new possibilities for studying the assembly of the membrane sector (F0) into the mitochondrial ATPase complex and the role of subunit 6 in this process.

J Biochem (Tokyo), 1981 Jun, 89(6), 1667 - 73
Purification and properties of sterol-ester hydrolase from Saccharomyces cerevisiae; Taketani S et al.; Sterol-ester hydrolase {EC 3.1.1.13} from Saccharomyces cerevisiae grown aerobically was solubilized with 1% Tween 20 and purified about 700-fold by the protamine sulfate treatment, DEAE-cellulose-, Sepharose 6B- and DEAE-cellulose column chromatographies . The molecular weight of the enzyme was estimated to be 70,000 by Sepharose 6B gel filtration . The enzyme activity showed two peaks of pH optimum at 4.4 and 6.8 . Triton X-100 stimulated the activity as its low concentrations at both pH regions, but decreased the activity at its high concentrations at pH 6.8 . The presence of Tween 20 or Tween 80 also stimulated the activity . These results were different from those in the previous report showing no stimulation of the crude enzyme by these detergents . The stimulation of the activity by phosphatidylcholine or low concentrations of lysophosphatidylcholine was similar to that by Triton X-100, and taurocholate was less effective than Triton X-100 . The enzyme activity was inhibited by divalent cations such as Hg2+ and Cu2+.

Mutat Res, 1981 Jun, 82(1), 95 - 100
Comparison of mutagenic and recombinogenic effects of some adenine analogues in Saccharomyces cerevisiae D7; Sorenson WG et al.; 2-Aminopurine, 2-amino-N6-hydroxyadenine and N6-hydroxyaminopurine were compared in suspension test with growing and non-growing cells for their mutagenic and recombinogenic (reciprocal and nonreciprocal) activities in Saccharomyces cerevisiae strain D7 . Ethyl methanesulfonate was used as a positive control . No increases above spontaneous frequencies were observed when non-growing cells were treated with the base analogues although EMS induced concentration-dependent responses at all 3 genetic end-points . When growing cells were treated, HAP was recombinogenic and mutagenic and AHA was mutagenic, but only weakly recombinogenic . HAP induced comparable numbers of revertants at much lower concentrations than AHA . 2AP failed to induce any detectable response even at concentrations as high as 2400 microgram/ml.

J Cell Biol, 1981 Jun, 89(3), 395 - 405
Roles of the CDC24 gene product in cellular morphogenesis during the Saccharomyces cerevisiae cell cycle; Sloat BF et al.; Temperature-sensitive yeast mutants defective in gene CDC24 continued to grow (i.e., increase in cell mass and cell volume) at restrictive temperature (36 degrees C) but were unable to form buds . Staining with the fluorescent dye Calcofluor showed that the mutants were also unable to form normal bud scars (the discrete chitin rings formed in the cell wall at budding sites) at 36 degrees C; instead, large amounts of chitin were deposited randomly over the surfaces of the growing unbudded cells . Labeling of cell-wall mannan with fluorescein isothiocyanate-conjugated concanavalin A suggested that mannan incorporation was also delocalized in mutant cells grown at 36 degrees C . Although the mutants have well-defined execution points just before bud emergence, inactivation of the CDC24 gene product in budded cells led both to selective growth of mother cells rather than of buds and to delocalized chitin deposition, indicating that the CDC24 gene product functions in the normal localization of growth in budded as well as in unbudded cells . Growth of the mutant strains at temperatures less than 36 degrees C revealed allele-specific differences in behavior . Two strains produced buds of abnormal shape during growth at 33 degrees C . Moreover, these same strains displayed abnormal localization of budding sites when growth at 24 degrees C (the normal permissive temperature for the mutants); in each case, the abnormal pattern of budding sites segregated with the temperature sensitivity in crosses . Thus, the CDC24 gene product seems to be involved in selection of the budding site, formation of the chitin ring at that site, the subsequent localization of new cell wall growth to the budding site and the growing bud, and the balance between tip growth and uniform growth of the bud that leads to the normal cell shape.

Mol Cell Biol, 1981 Jun, 1(6), 522 - 34
Homothallic mating type switching generates lethal chromosome breaks in rad52 strains of Saccharomyces cerevisiae; Weiffenbach B et al.; In homothallic cells of Saccharomyces cerevisiae, a or alpha mating type information at the mating type locus (MAT) is replaced by the transposition of the opposite mating type allele from HML alpha or HMRa . The rad52-1 mutation, which reduces mitotic and abolishes meiotic recombination, also affects homothallic switching (Malone and Esposito, Proc . Natl . Acad . Sci . U.S.A . 77:503-507, 1980) . We have found that both HO rad52 MATa and HO rad52 MAT alpha cells die . This lethality is suppressed by mutations that substantially reduce but do not eliminate homothallic conversions . These mutations map at or near the MAT locus (MAT alpha inc, MATa-inc, MATa stk1) or are unlinked to MAT (HO-1 and swi1) . These results suggest that the switching event itself is involved in the lethality . With the exception of swi1, HO rad52 strains carrying one of the above mutations cannot convert mating type at all . MAT alpha rad52 HO swi1 strains apparently can switch MAT alpha to MATa . However, when we analyzed these a maters, we found that few, if any, of them were bona fide MATa cells . These a-like cells were instead either deleted for part of chromosome III distal to and including MAT or had lost the entire third chromosome . Approximately 30% of the time, an a-like cell could be repaired to a normal MATa genotype if the cell was mated to a RAD52 MAT alpha-inc strain . The effects of rad52 were also studied in mata/MAT alpha-inc rad52/rad52 ho/HO diploids . When this diploid attempted to switch mata to MATa, an unstable broken chromosome was generated in nearly every cell . These studies suggest that homothallic switching involves the formation of a double-stranded deoxyribonucleic acid break or a structure which is labile in rad52 cells and results in a broken chromosome . We propose that the production of a double-stranded deoxyribonucleic acid break is the lethal event in rad52 HO cells.

Biochim Biophys Acta, 1981 May 29, 668(3), 333 - 8
Possible functional roles of carboxyl and histidine residues in a soluble thiamine-binding protein of Saccharomyces cerevisiae; Nishimura H et al.; The reaction of a soluble thiamine-binding protein of Saccharomyces cerevisiae with water-soluble carbodiimide, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide, at pH 4.5, results in a remarkable loss of its binding activity with thiamine . Thiamine above 0.1 mM substantially protects the protein against this inactivation . In addition to 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide, the thiamine-binding protein is also inactivated by diethylpyrocarbonate . The inactivation is time-dependent and follows second-order kinetics . Restoration of the binding activity by incubation of inactivated protein with hydroxylamine was observed . thiamine and pyrithiamine are effective to prevent the inactivation . From these results it is strongly suggested that both the carboxyl and the histidine residues in the protein are involved in the binding site for thiamine . It is proposed that the binding involves interactions between charged groups on the protein with the quaternary nitrogen of the thiazolium moiety and with the basic ring nitrogen of the pyrimidine moiety in thiamine molecule.

Biochim Biophys Acta, 1981 May 29, 653(3), 416 - 22
mit-Mutations in the structural gene of subunit III of cytochrome oxidase in Saccharomyces cerevisiae; Stephenson G et al.; Two-dimensional electrophoretic analysis of the mitochondrial translation products of four mit-mutants indicate that subunit III of cytochrome oxidase is the only mitochondrial translation product affected by mutations in the oxi2 region of the mtDNA . Mitochondria of two of these mutants synthesize new products which coprecipitate with an anticytochrome oxidase antiserum and produce proteolytic digests similar to those of subunit III of the enzyme complex . These data strongly support the suggestion that the oxi2 region of the yeast mtDNA contains the structural gene of subunit III of cytochrome oxidase.

J Biol Chem, 1981 May 25, 256(10), 5299 - 603
Partial purification from Saccharomyces cerevisiae of a soluble glucosidase which removes the terminal glucose from the oligosaccharide Glc3Man9GlcNAc2; Kilker RD Jr et al.; Glucosidase activities capable of removing the three glucose residues from Glc3Man9GlcNAc2 oligosaccharide were detected in a cell-free preparation of Saccharomyces cerevisiae X-2180 . The glucosidase which cleaves the glucose residue at the nonreducing terminus (Glc3Man9GlcNAc2 oligosaccharide glucosidase) was equally distributed between the particulate and the supernatant fractions obtained after centrifugation of the yeast homogenate at 27,000 X g for 30 min . The membrane-bound activity was stimulated by Triton X-100, whereas the supernatant activity was not affected . The soluble Glc3Man9GlcNAc2 oligosaccharide glucosidase was partially purified from the supernatant by ammonium sulfate fractionation followed by DEAE-Sephadex chromatography . It was clearly separated from alpha-glucosidase, which acts onp-nitrophenyl-alpha-D-glucopyranoside, but still contained beta-glucosidase and alpha-mannosidase acting on p-nitrophenyl-beta-D-glucopyranoside and alpha-D-mannopyranoside, respectively . The Glc3Man9GlcNAc2 oligosaccharide glucosidase had a pH optimum of 6.8, and showed no requirement for divalent cations . The enzyme was very active with glucose-labeled Glc3Man9GlcNAc2, was slightly active with Glc2Man9GlcNAc2, and showed no activity with Glc1Man9GlcNAc2 . These properties suggest that this enzyme is involved in the first step of processing of oligosaccharides after transfer from dolichyl pyrophosphate to proteins.

J Biol Chem, 1981 May 25, 256(10), 5041 - 5
Effect of methylation on the stability of cytochrome c of Saccharomyces cerevisiae in vivo; Farooqui J et al.; The in vivo stability of methylated and unmethylated cytochrome c in Saccharomyces cerevisiae was studied by pulse-labeling the hemoproteins with {methyl-3H}-methionine and/or {2-14C}methionine and following the fate of these proteins under anaerobiosis and in the presence of cycloleucine . These two conditions will respectively block further cytochrome c synthesis and inhibit methylation by lowering the cellular S-adenosyl-L-methionine pool and, thus, permit an unambiguous interpretation of the data . The results showed that the rate of degradation of unmethylated cytochrome c was constant throughout the chase period, while methylated cytochrome c degradation was seen only in the later part of cold chase . At the end of the chase period (40 h), the extent of degradation of the unmethylated species was three times higher than the methylated species . This indicated that the methylation of cytochrome c has a protective effects against the intracellular proteolytic enzyme attack on itself . Furthermore, this protective effect was considerably reduced in the petite mutant, which lacks high affinity cytochrome c binding sites, functional cytochrome c reductase, and oxidase, and possesses a less integrated and organized mitochondrial membrane . These results led us to the conclusion that the mechanism of methylated cytochrome c stabilization is best explained by a higher efficacy of binding to the mitochondria.

Biochim Biophys Acta, 1981 May 20, 643(3), 583 - 92
Processes involved in the creation of buffering capacity and in substrate-induced proton extrusion in the yeast Saccharomyces cerevisiae; Sigler K et al.; The high pH-maintaining capacity of yeast suspension after glucose-induced acidification, measured as its ability to neutralize added alkali, was found to be due mainly to actively extruded acidity (H+) . The buffering action of passively excreted metabolites (CO2, organic acids) and cell surface polyelectrolytes contributed only 15--40% to the overall pH-maintaining capacity which was 10 mmol NaOH/l per pH unit between pH 3 and 4 and 3.5 nmol NaOH/l per pH unit between pH 4 and 7 . The buffering capacity of yeast cell-free extract was still higher (up to 4.5-times) than that of glucose-supplied cell suspension; addition of glucose to the extract thus produced considerable titratable acidity but negligible net acidity . The glucose-induced acidification of yeast suspension was stimulated by univalent cations in the sequence K+ greater than Rb+ much greater than Li+ congruent to Cs+ congruent to Na+ . The processes participating in the acidification and probably also in the creation of extracellular buffering capacity include excretion of CO2 and organic acids, net extrusion of H+ and K+ (in K+-free media; in K+-containing media this is preceded by an initial rapid K+ uptake), and movements of some anions (phosphate, chlorides) . The overall process appears to be electrically silent.

Biochim Biophys Acta, 1981 May 20, 643(3), 572 - 82
Factors governing substrate-induced generation and extrusion of protons in the yeast Saccharomyces cerevisiae; Sigler K et al.; Experiments with respiration deficient (rho-), ADP/ATP transport deficient (op1) and double (op1 rho-) mutants, with glycolytic and tricarboxylic acid cycle substrates showed that the substrate-induced acidification of yeast suspensions is closely associated with glycolysis . The glucose/proton stoichiometry is 2.5 : 1 to 4 : 1 depending on glucose concentration . The kinetics of the process are complex, the acidification curve having a very fast initial component and two slower exponential components . The first component suggests an initial proton efflux from endogenous sources, triggered by exogenous substrates . The acidification process exhibits two Km values at about 1 and 15 mM D-glucose, indicating two distinct saturable pathways of proton extrusion . The total extent of acidification and thus the final pHout reaches a saturation value with increasing glucose concentration and suspension density . Both the total extent and the rate of acidification are subject to control by extracellular pH which reflects the tendency of the cells to build a fixed {H+}out/{H+}in ratio . When the control is lifted, both quantities are considerably increased . A crucial role in the substrate-induced acidification is thus played by active membrane processes and their control mechanisms.

Biochem J, 1981 May 15, 196(2), 531 - 6
Changes in membrane proteins associated with inhibition of the general amino acid permease of yeast (Saccharomyces cerevisiae); Woodward JR et al.; The general amino acid permease ('Gap') system of the wild-type yeast (Saccharomyces cerevisiae) strain Y185 is inhibited by the uptake and accumulation of its substrate amino acids . Surprisingly, this inhibition persists even after 'pools' of amino acids, accumulated initially, have returned to normal sizes . Recovery from this inhibition depends on a supply of energy and involves the synthesis of a membrane protein component of the Gap system.

Proc Natl Acad Sci U S A, 1981 May, 78(5), 3030 - 3
Nature of the G1 phase of the yeast Saccharomyces cerevisiae; Singer RA et al.; Under conditions that protract the S phase for Saccharomyces cerevisiae without affecting steady-state rates of cell growth or proliferation, there were striking decreases in the length of the G1 period . These decreases were localized in the period between mitosis and the start event that initiates a new cell cycle . We conclude that this major fraction of the G1 period has no functional role in the DNA-division sequence of cell cycle events.

Proc Natl Acad Sci U S A, 1981 May, 78(5), 2693 - 7
13C NMR study of transamination during acetate utilization by Saccharomyces cerevisiae; den Hollander JA et al.; 13C NMR was used to follow the metabolism of {2- 13C}acetate and {1- 13C}acetate in aerobic suspensions of Saccharomyces cerevisiae . In the experiment with {2- 13C}acetate, the 13C label appeared first in glutamate C4 and subsequently in glutamate C2 and C3 . After exhaustion of the acetate, the glutamate signals diminished and the aspartate C2 and C3 peaks increased . During a subsequent chase experiment with unlabeled acetate, the aspartate peaks decreased and the glutamate C2 and C3 peaks increased in intensity . These observations are interpreted in terms of an interplay between the glutamic-oxalacetic transaminase and Krebs cycle activity . This interpretation was confirmed by an experiment with the transaminase inhibitor 2-amino oxyacetate . During all of these experiments, we observed the formation of trehalose . The NMR gives a direct measurement of the label distribution and from that information it followed that the flows through the glyoxylate and the Krebs cycles are comparable . The intermediates citrate, succinate, fumarate, malate, phosphoenolpyruvate, 3-phosphoglycerate, and glucose 6-phosphate were identified in a 13C NMR spectrum of a perchloric acid extract taken during the metabolism of {2- 13C}acetate . Enrichment of the glutamate C5 position shows the existence of a futile cycle in which phosphoenolpyruvate, formed from oxaloacetate, returns to the Krebs cycle through pyruvate and acetyl CoA

Eur J Biochem, 1981 May, 116(1), 1 - 6
Induction of choline transport and its role in the stimulation of the incorporation of choline into phosphatidylcholine by polyamines in a polyamine auxotroph of Saccharomyces cerevisiae; Hosaka K et al.; 1 . A mutant of Saccharomyces cerevisiae, defective in ornithine decarboxylase, was isolated . A prolonged culture of the mutant in a polyamine-free medium resulted in a great decrease in the polyamine content and in cessation of growth . The addition of polyamines to the culture induced the growth after a lag period of 5--6.5 h . The growth rate in the presence of polyamine was comparable to that of the wild-type strain . The effectiveness of polyamines was as follows: spermidine greater than putrescine approximately equal to spermine . 2 . Phosphatidylcholine-synthesizing activity during the lag phase of growth was determined by measuring the rate of incorporation of {14C}choline into phosphatidylcholine . The incorporation rate was markedly increased with time by polyamine prior to the initiation of cell division . Polyamines were effective in the following order: spermidine greater than putrescine approximately equal to spermine . Experiments with methylglyoxal bis(guanylhydrazone), an inhibitor of S-adenosylmethionine decarboxylase, showed that putrescine stimulates cell growth and choline incorporation into phosphatidylcholine after it has been converted into spermidine in the cell . 3 . The induction of the choline transport system was shown to be responsible for the increase in the rate of incorporation of {14C}choline into phosphatidylcholine effected by polyamines . A low concentration of cycloheximide completely prevented the induction of choline transport by polyamines . The levels of the CDP-choline pathway enzymes such as choline kinase, cholinephosphate cytidyltransferase and cholinephosphotransferase were not significantly changed.

J Bacteriol, 1981 May, 146(2), 775 - 83
Metabolism of myo-inositol during sporulation of myo-inositol-requiring Saccharomyces cerevisiae; Schroeder R et al.; We investigated the sporulation properties of a series of diploid Saccharomyces cerevisiae strains homozygous for inositol auxotrophic markers . The strains required different amounts of inositol for the completion of sporulation . Shift experiments revealed two phases of inositol requirement during sporulation which coincided with the two phases of lipid synthesis found by earlier workers . Phase I was at the beginning and during premeiotic deoxyribonucleic acid synthesis; phase II immediately preceded the appearance of mature asci . Of the inositol taken up by sporulating cells, 90% was incorporated into inositol phospholipids . By two-dimensional thin-layer chromatography, eight compounds were resolved, one of which was sporulation specific . The majority of the inositol phospholipids were, however, identical to those found in vegetatively growing cells . In the absence of inositol, the cells did not sporulate but, after a certain time, were unable to return to vegetative growth . These nonsporulating cells did, however, incorporate acetate into lipids and double their deoxyribonucleic acid content in the premeiotic phase . We believe that it is this lack of coordination of biosynthetic events which causes inositol-less death on sporulation media without inositol.

J Bacteriol, 1981 May, 146(2), 684 - 91
a/alpha-specific effect on the mms3 mutation on ultraviolet mutagenesis in Saccharomyces cerevisiae; Martin P et al.; A new gene involved in error-prone repair of ultraviolet (UV) damage has been identified in Saccharomyces cerevisiae by the mms3-1 mutation . UV-induced reversion is reduced in diploids that are homozygous for mms3-1, only if they are also heterozygous (MATa/MAT alpha) at the mating type locus . The mms3-1 mutation has no effect on UV-induced reversion either in haploids or MATa/MATa or MAT alpha/MAT alpha diploids . The mutation confers sensitivity to UV and methyl methane sulfonate in both haploids and diploids . Even though mutation induction by UV is restored to wild-type levels in MATa/MATa mms3-1/mms3-1 or MAT alpha/MAT alpha mms3-1/mms3-1 diploids, such strains still retain sensitivity to the lethal effects of UV . Survival after UV irradiation in mms3-1 rad double mutant combinations indicates that mms3-1 is epistatic to rad6-1 whereas non-epistatic interactions are observed with rad3 and rad52 mutants . When present in the homozygous state in MATa/MAT alpha his1-1/his1-315 heteroallelic diploids, mms3-1 was found to lower UV-induced mitotic recombination.

Biochem J, 1981 May 1, 195(2), 407 - 17
Apurinic endonuclease from Saccharomyces cerevisiae; Thielmann HW et al.; An endonuclease cleaving depurinated and alkylated double-stranded DNA has been purified 500-fold from Saccharomyces cerevisiae, strain MB 1052 . The enzyme has an Mr of 31 000 +/- 2000, a sedimentation value of 3.2S and a diffusion coefficient of 9.5 X 10-7 cm2/s . The enzyme was active only at apurinic/apyridiminic sites, regardless of whether they were produced by heating the DNA at acidic pH or by alkylation with the ultimate carcinogen methyl methanesulphonate . Native DNA was not acted upon . U.v.-irradiated DNA and DNA treated with the ultimate carcinogen N-acetoxy-2-acetylaminofluorene were cleaved to an extent related to the extent of apurinic/apyridiminic sites . Enzymic activity was not dependent upon Mg2+, but was stimulated approx . 3-fold by 4mM-Mg2+ . The enzyme did not bind to DEAE-cellulose or CM-cellulose at KCl concentrations greater than 160 mM . The endonuclease was obtained free of exonuclease and 3-methyladenine-DNA glycosylase activity in five chromatographic steps.

Proc Natl Acad Sci U S A, 1981 Apr, 78(4), 2125 - 9
31P NMR studies of intracellular pH and phosphate metabolism during cell division cycle of Saccharomyces cerevisiae; Gillies RJ et al.; We have analyzed changes in intracellular pH and phosphate metabolism during the cell cycle of Saccharomyces cerevisiae (NCYC 239) by using high-resolution 31P NMR spectroscopy . High-density yeast cultures (2 x 10(8) cells per ml) were arrested prior to "start" by sequential glucose deprivation, after which they synchronously replicated DNA and divided after a final glucose feeding . Oxygenation of arrested cultures in the absence of glucose led to increased levels of sugar phosphates and ATP and an increase in intracellular pH . However, these conditions did not initiate cell cycle progression, indicating that energization is not used as an intracellular signal for initiation of the cell division cycle and that the cells need exogenous carbon sources for growth . Glucose refeeding initiated an alkaline intracellular pH transient only in the synchronous cultures, showing that increased intracellular pH accompanies the traversal of start . Changes in phosphate flow and utilization also were observed in the synchronous cultures . In particular, there was increased consumption of external phosphate during DNA synthesis . When external phosphate levels were low, the cells consumed their internal polyphosphate stores . This shows that, under these conditions, polyphosphate acts as a phosphate supply.

J Virol, 1981 Apr, 38(1), 263 - 71
Replication of double-stranded RNA of the virus-like particles in Saccharomyces cerevisiae; Newman AM et al.; The mode of replication of the L double-stranded RNA (dsRNA) present in virus-like particles in Saccharomyces cerevisiae was examined by density transfer experiments . After transfer to light medium, significant amounts of fully heavy dsRNA persisted over a number of cell doublings . In addition, very little material of hybrid density was ever formed, and the accumulation of fully light material began as early as 0.5 doubling after transfer to light medium . Our results are compatible with a conservative mode of replication or with a semiconservative mode of replication carried out by a small portion of the total dsRNA population . In additional experiments the synthesis of dsRNA relative to the cell cycle was studied . This was done by determining the ratio of short-term to long-term radioactive label in size-separated cell fractions of a prelabeled exponential culture . The ratio of short-term to long-term label remained constant for all fractions, implying that dsRNA is synthesized throughout the cell cycle, increasing through the cell cycle at an exponential rate.

Mol Cell Biol, 1981 Apr, 1(4), 381 - 6
Physical evidence for a Saccharomyces cerevisiae transposable element which carries the his4C gene; de Bruijn F et al.; A Saccharomyces cerevisiae transposable element which carries the his4C structural gene and which is capable of transposition, excision, and mutator activity is described . Physical evidence is presented for transposition of the his4C deoxyribonucleic acid sequences to a new location in the genome and for precise excision of these transposed deoxyribonucleic acid sequences in spontaneous his4C- segregants.

Nucleic Acids Res, 1981 Mar 25, 9(6), 1351 - 64
A putative precursor for the small ribosomal RNA from mitochondria of Saccharomyces cerevisiae; Osinga KA et al.; We have characterized a putative precursor RNA (15.5S) for the 15S ribosomal RNA in mitochondria of Saccharomyces cerevisiae . Hybrids were formed with mitochondrial RNA and mtDNA fragments terminally labelled at restriction sites located within the gene coding for 15S ribosomal RNA and treated with S1 nuclease (Berk, A.J . and Sharp, J.A . (1977) 12, 721-732) . Sites of resistant hybrids were measured by agarose gel electrophoresis and end points of RNAs determined . The 15.5S RNA is approximately 80 nucleotides longer than the 15S ribosomal RNA, with the extra sequences being located at the 5'-end . Both 15S ribosomal RNA and 15.5S RNA are fully localised within a 2000 base pair HapII fragment . This putative precursor and the mature 15S ribosomal RNA are also found in petite mutants which retain the 15S ribosomal RNA gene . The petite mutant with the smallest genetic complexity has its end point of deletion (junction) just outside the HapII site located in the 5' flank of the 15S ribosomal RNA genes as determined by S1 nuclease analysis . This leaves a DNA stretch approximately 300 base pairs long where an initiation signal for mitochondrial transcription may be present.

J Biol Chem, 1981 Mar 10, 256(5), 2561 - 6
alpha-Factor-mediatd modification of a 32P-labeled protein by MATa cells of Saccharomyces cerevisiae; Finkelstein DB et al.; Addition of the polypeptide mating pheromone alpha-factor to haploid MATa cells of Saccharomyces cerevisiae results in the modification of a 32P-labeled protein (P17) with an apparent Mr of 17,000 to a form having an apparent Mr of 17,500 (P17) . 32P associated with both P17 and P17 exhibits an unusually rapid rate of turnover . The conversion of P17 to P17 precedes the appearance of morphologically abnormal cells and, in contrast to other responses elicited by this pheromone, this change in apparent molecular weight does not require protein synthesis . Upon removal of alpha-factor, the P17/P17 ratio returns to pretreatment levels.

J Biol Chem, 1981 Mar 10, 256(5), 2079 - 82
Active transport of basic amino acids driven by a proton motive force in vacuolar membrane vesicles of Saccharomyces cerevisiae; Ohsumi Y et al.; The mechanism of transport of basic amino acids into vacuoles of cells of the yeast Saccharomyces cerevisiae was investigated in vitro . Right-side-out vacuolar membrane vesicles were prepared from purified vacuoles . Arginine was taken up effectively by the vesicles only in the presence of ATP, not in the presence of ADP or AMP-adenosyl-5'-yl imidodiphosphate . It was exchangeable and was released completely by a protonophore, 3,5-di-tert-butyl-4-hydroxybenzilidenemalononitrile (SF6847) . The transport required Mg2+ ion but was inhibited by Cu2+, Ca2+, or Zn2+ ions . The transport activity was sensitive to the ATPase inhibitor N,N'-dicyclohexylcarbodiimide (DCCD), but not to oligomycin or sodium vanadate . SF6847 or nigericin blocked arginine uptake completely, but valinomycin had no effect . ATP-dependent formation of a delta pH across the membrane vesicles was shown by quenching of 9-aminoacridine fluorescence . These results indicate that DCCD-sensitive, Mg2+-ATPase of vacuolar membranes is essential as an energy-donating system for the active transport, and that an electrochemical potential difference of protons is a driving force of this basic amino acid transport . Arginine transport showed saturation kinetics with a Km value of 0.6 mM and the mechanism was well explained by an H+/arginine antiport.

Genetics, 1981 Mar-Apr, 97(3-4), 551 - 62
An endomitotic effect of a cell cycle mutation of Saccharomyces cerevisiae; Schild D et al.; A recessive temperature-sensitive mutation of Saccharomyces cerevisiae has been isolated and shown to cause an increase in ploidy in both haploids and diploids . Genetic analysis revealed that the strain carrying the mutation was an aa diploid, although MNNG mutagenesis had been done on an a haploid strain . When the mutant strain was crossed with an alpha alpha diploid and the resultant tetraploid sporulated, some of the meiotic progeny of this tetraploid were themselves tetraploid, as shown by both genetic analysis and DNA measurements, instead of diploid as expected of tetraploid meiosis . The ability of these tetraploids to continue to produce tetraploid meiotic progeny was followed for four generations . Homothallism was excluded as a cause of the increase in ploidy; visual pedigree analysis of spore clones to about the 32-cell stage failed to reveal any zygotes, and haploids that diploidized retained their mating type . An extra round of meiotic DNA synthesis was also considered and excluded . It was found that tetraploidization was independent of sporulation temperature, but was dependent on the temperature of germination and the growth of the spores . Increase in ploidy occurred when the spores were germinated and grown at 30 degrees, but did not occur at 23 degrees . Two cycles of sporulation and growth at 23 degrees resulted in haploids, which were shown to diploidize within 24 hr when grown at 30 degrees . Visual observation of the haploid cells incubated at 36 degrees revealed a cell-division-cycle phenotype characteristic of mutations that affect nuclear division; complementation analysis demonstrated that the mutation, cdc31-2, is allelic to cdc31-1, a mutation isolated by Hartwell et al . (1973) and characterized as causing a temperature-sensitive arrest during late nuclear division . The segregation of cdc31-2 in heterozygous diploids was 2:2 and characteristic of a noncentromere-linked gene.

An Acad Bras Cienc, 1981 Mar, 53(1), 165 - 72
Biochemical genetics of trehalose metabolism in Saccharomyces cerevisiae; Panek AD et al.; Different strains of Saccharomyces cerevisiae exhibit alternative patterns of trehalose accumulation during growth on a glucose medium . The active pattern is designated as TAC(+) phenotype and the alternative, low-activity pattern as tac(-) phenotype . The tac(-) phenotype is expressed only during growth, since tac(-) strains actively accumulate trehalose during incubation in a glucose medium lacking a nitrogen source . The tac(-) phenotype appears to be determined by a single, recessive gene tac1 . The quantitative expression of the dominant, alternative allele TAC1 is subject to wide variation . A highly active pattern of trehalose accumulation requires TAC1 and an amplification factor, TAM, which consists of one or more dominant gene(s) . TAM does not appear to alter significantly the expression of tac1 . Highly amplified TAC(+) strains may contain a labile factor not present in a TAC(+) strain which accumulates intermediate levels of trehalose.

Biokhimiia, 1981 Mar, 46(3), 444 - 52
{Isolation and properties of maltase from Saccharomyces cerevisiae-II}; Glemzha AA et al.; Maltase from Saccharomyces cerevisiae-II was purified by ion-exchange chromatography on DEAE-Sephadex A-50 and isoelectric focusing . The purification procedure resulted in two enzyme isoforms with pI of 5.35 and 5.3 and identical specific activities . The molecular weights of the isoforms as determined by SDS polyacrylamide gel electrophoresis and gel filtration through Sephadex G-100 are 60 000 and 55 000, respectively . Both isoforms were electrophoretically polydisperse . The maltase isoforms are glycoproteins containing 1.5-2% of glucosamine and 5-8% (isoform A) and 2-3% (isoform B) of neutral sugars . Using paper chromatography and glucose oxidase, it was shown that glucose is an indispensable constituent of neutral sugars in both isoforms.

Lipids, 1981 Mar, 16(3), 195 - 8
Metabolism of sterols by anaerobic Saccharomyces cerevisiae; Sekula BC et al.; Anaerobically grown Saccharomyces cerevisiae retained the ability to transfer a C1-group to the C-24 position of a delta 24(25)-sterol and to reduce the delta 25(28)-bond of a 24-methylenesterol . Both desmosterol and 24-methylenecholesterol yielded 24 beta-methylcholesterol . However, when the substituent at C-24 was enlarged to a 24-ethylidene group (fucosterol), reduction of the delta 24(28)-bond did not occur . In no cases was a delta 7- or a delta 22-bond introduced . Because the delta 24(28)-bond was reduced in the absence of the delta 22-bond, the delta 22-bond is not an obligatory requirement for reduction.

J Bacteriol, 1981 Mar, 145(3), 1421 - 4
Curing of the 2 mu DNA plasmid from Saccharomyces cerevisiae; Toh-e A et al.; The 2 mu DNA plasmid is often eliminated from yeast cells when they are transformed with the 2 mu DNA-LEU2-pMB9 composite plasmid pJDB219 . Since pJDB219 is subsequently lost with high frequency, derivatives lacking all 2 mu DNA can be prepared from any strain.

J Bacteriol, 1981 Mar, 145(3), 1359 - 64
Subcellular compartmentation in control of converging pathways for proline and arginine metabolism in Saccharomyces cerevisiae; Brandriss MC et al.; Enzymes of proline biosynthesis and proline degradation which act on the same compound, delta 1-pyrroline-5-carboxylate, are physically separated in yeast cells . The enzyme responsible for the final step in proline biosynthesis, pyrroline-5-carboxylate reductase, converts pyrroline-5-carboxylate to proline and is located in the cytoplasm . The last enzyme in the proline degradative pathway, pyrroline-5-carboxylate dehydrogenase, converts pyrroline-5-carboxylate to glutamate and is found in the particulate fraction of the cell, presumably in the mitochondrion . By subcellular compartmentation, yeast cells avoid futile cycling between proline and pyrroline-5-carboxylate.

J Bacteriol, 1981 Mar, 145(3), 1325 - 33
Corresponding changes in kynurenine hydroxylase activity, membrane fluidity, and sterol composition in Saccharomyces cerevisiae mitochondria; McLean-Bowen CA et al.; The effect of sterol composition on the properties of the mitochondrial membrane of Saccharomyces cerevisiae was investigated . The physical state of mitochondrial membranes from wild-type strains and sterol mutants was compared, using a fluorescence polarization technique with 1,6-diphenyl-1,3-5-hexatriene . Changes in the rate of depolarization of the probe molecule as a function of temperature suggest the occurrence of a phase transition in the mitochondrial membranes isolated from the sterol mutants but not in the membranes isolated from the wild types . Arrhenius kinetics of the mitochondrial membrane-bound enzyme L-kynurenine-3-hydroxylase exhibited changes in activation energy at temperatures similar to those observed in the fluorescence polarization study . The ratio of mitochondrial sterol to phospholipid and the phospholipid fatty acid composition of the organisms were characterized.

Eur J Biochem, 1981 Mar, 114(3), 609 - 13
Effect of phosphorylation on the affinity of acidic proteins from Saccharomyces cerevisiae for the ribosomes; Sanchez-Madrid F et al.; Electrofocusing of the acidic proteins extracted from Saccharomyces cerevisiae ribosomes shows the presence of eight bands in the gels, which upon treatment with alkaline phosphatase are reduced to three . Two of them, proteins L44 and L45, correspond to the proteins equivalent to the bacterial L7 and L12 and the third, protein Ax, behaves like a supernatant factor . In the ribosome, proteins L44 and L45 are found unphosphorylated and monophosphorylated while protein Ax is detected mostly in a modified state, showing from one to three phosphate groups per molecule . In the cytoplasm where protein Ax is abundant and proteins L44 and L45 are present in small quantities, the three proteins are unphosphorylated . Protein Ax, having one or two phosphate groups, can be removed from the ribosomes in conditions that release the initiation factors, while the triphosphorylated molecules are tightly bound to the particles . The data indicate a relationship between the degree of phosphorylation of protein Az and its affinity for the ribosome.

Mol Cell Biol, 1981 Mar, 1(3), 245 - 53
Internuclear transfer of genetic information in kar1-1/KAR1 heterokaryons in Saccharomyces cerevisiae; Dutcher SK; Heterokaryons of Saccharomyces cerevisiae have been constructed utilizing the kar1-1 mutation, which prevents nuclear fusion during conjugation (J . Conde and G . Fink, Proc . Natl . Acad . Sci . U.S.A . 73:3651-3655, 1976) . Each heterokaryon contained two haploid nuclei that were marked on several chromosomes . They segregated haploid progeny (cytoductants), most of which have the nuclear genotype of one or the other of the heterokaryon parents, but they occasionally segregated progeny having a recombinant genotype (exceptional cytoductants) . Exceptional cytoductants receive the majority of their genome from one parent (the recipient) and a minority from the other (the donor) . Transfer of two markers from the donor nucleus to the recipient is rarely coincident for markers located on different chromosomes but is nearly always coincident for those markers located on the same chromosome, suggesting that whole chromosomes are transferred from the donor nucleus to the recipient . In crosses of kar1-1 X KAR1 parents, either nucleus may act as a recipient or donor with equal probability . Recipient nuclei acquired 9 of the 10 chromosomes examined, with frequencies which were inversely correlated with the size of the chromosome . When a chromosome is acquired by the recipient nucleus, it either replaces its homolog or exists in a disomic condition . Haploid progeny emanating from kar1 X KAR1 crosses are frequently inviable . I tested whether this inviability might be the result of chromosome loss by donor nuclei . Viability of progeny from kar1 X KAR1 heterokaryons was improved when the parental nuclei were diploid to an extent consistent with the hypothesis, and diploid progeny which had become monosomic were recovered from these heterokaryons . The following sequence of events accounts for chromosome transfer in kar1 X KAR1 heterokaryons . After cell fusion, each nucleus in the heterokaryon has a probability of about 0.38 of losing one or more chromosomes . A nucleus sustaining such a loss can become a donor in a chromosome transfer event . If the other nucleus does not sustain a mortal chromosome loss, it can become a recipient in a transfer event . The chance of acquiring a chromosome lost by the donor is greater for smaller chromosomes than for larger ones and is about 0.05 for the average chromosome.

Mol Cell Biol, 1981 Mar, 1(3), 228 - 36
Physical analysis of the CYC1-sup4 interval in Saccharomyces cerevisiae; Shalit P et al.; CYC1 and sup4 are part of a tightly linked cluster of genes on chromosome X in the yeast Saccharomyces cerevisiae . Using as probes previously cloned fragments containing the CYC1 and sup4 genes, we have identified and cloned the deoxyribonucleic acid (DNA) present between these genes in one strain of yeast . We find that the CYC1 and sup4 genes are approximately 21 kilobases apart . In the same strain, the meiotic map distance is approximately 3.7 centimorgans, for a ratio of 5.6 kilobases per centimorgan in this interval . The physical mapping has allowed unambiguous determination of the orientation of CYC1 and sup4 relative to each other, the centromere, and a nearby transfer ribonucleic acid (tRNA(2Ser)) gene . The spontaneous mutation cyc1-1 inactivates the CYC1 gene as well as the neighboring loci OSM1 and RAD7 . We have determined that a cyc1-1-bearing strain lacks approximately 13 kilobases of single-copy DNA from the CYC1-sup4 region, including all of the CYC1 coding information . There is a sequence homologous to the middle-repetitive element Ty1 at or near the breakpoint of the cyc1-1 deletion . We discuss the possibility that Ty elements play a role in the formation of such large, spontaneous deletions, which occur frequently in this region of chromosome X in certain yeast strains.

Genetics, 1981 Mar-Apr, 97(3-4), 531 - 49
Mating-type differentiation by transposition of controlling elements in Saccharomyces cerevisiae; Oshima T et al.; The nonfunctional mutation of the homothallic gene HML alpha, designated hml alpha, produced two mutant alleles, hml alpha-1 and hml alpha-2 . Both mutant clones were mixed cultures consisting of a mating-type cells and nonmating haploid cells . The frequencies of the two cell types were different, and a few diploid cells able to sporulate were found in the hml alpha-2 mutant . Conversions of an a mating-type cell to nonmater, and vice versa, were observed in both mutants . The conversion of an a mating phenotype to nonmating is postulated to occur by alteration of the a mating type to the sterile mating-type allele in the hml alpha-1 mutant . In tetrad dissection of prototrophic diploids that were obtained by rare mating of hml alpha-1 mutants with a heterothallic strain having the MATa ho HMRa HMLa genotype, many mating-deficient haploid segregants were found, while alpha mating-type segregants were observed in a similar diploid using an hml alpha-2 mutant . The mating-type-deficient haploid segregants were supposed to have the sterile alpha mating-type allele because the nonmating genetic trait always segregated with the mating-type locus . Sporogenous diploid cells obtained in the hml alpha-2 mutant clone had the MATa/MAT alpha HO/HO HMRa/HMRa hml alpha-2/hml alpha-2 genotype . These observations suggested that the hml alpha-1 allele produces a transposable element that gives rise to the sterile alpha mating type by transposition into the mating-type locus, and that the hml alpha-2 allele produces an element that provides alpha mating-type information, but is defective in the structure for transposition.

Biochim Biophys Acta, 1981 Feb 13, 657(2), 482 - 94
A particulate form of alkaline phosphatase in the yeast, Saccharomyces cerevisiae; Mitchell JK et al.; A new form of alkaline phosphatase (orthophosphoric-monoester phosphohydrolase (alkaline optimum), EC 3.1.3.1) has been identified in the yeast Saccharomyces cerevisiae . Utilizing either synthetic or natural substrates, the enzyme exhibited a broad pH activity curve with maximum activity between 8.5 and 9.0 . The enzyme was nonspecific with respect to substrate, attacking a variety of compounds containing phosphomonoester linkages, but has no detectable activity against polyphosphate, pyrophosphate or phosphodiester linkages . The enzyme exhibited an apparent Km of 0.25 mM with respect to p-nitrophenyl phosphate, 0.38 mM with respect to alpha-naphthyl phosphate, and 1.0 mM with respect to 5'AMP . The enzyme is regulated in a constitutive manner and its activity does not increase during phosphate starvation or sporulation, as does the repressible alkaline phosphatase . The enzyme is tightly bound to a particulate fraction of the cell, tentatively identified as the tonoplast membrane . It is not solubilized by treatment with high concentrations of NaCl, KH2PO4 or chaotropic agents . Triton X-100 (0.1%) solubilizes 12% of the particulate activity . This enzyme is differentiated from the other alkaline phosphatases found in yeast by its chromatographic elution DEAE-cellulose, kinetic parameters, heat stability and pH stability, as well as its particulate nature . This particulate alkaline phosphatase was found in every strain examined . It has a significantly lower specific activity in the phoH mutant and a higher activity in the acid phosphatase constitutive mutant A137.

J Biol Chem, 1981 Feb 10, 256(3), 1474 - 81
Purification of mitochondrial RNA polymerase from Saccharomyces cerevisiae; Levens D et al.; The RNA polymerase from the mitochondria of Saccharomyces cerevisiae has been extensively purified by Sepharose 4B, heparin Sepharose 4B phosphocellulose, and DEAE-Sephadex A-50 chromatography . The activity co-sediments with a 45,000-dalton polypeptide at 6.3 S in glycerol gradients . The activity is inhibited by antibodies to the 45,000-dalton polypeptide . The activity is not inhibited by rifampicin or alpha-amanitin . It requires Mg2+ and is inhibited by elevated ionic strength and Mn2+ . The most efficient template for the RNA polymerase is poly{d(AT)}, with mtDNA being the preferred natural template . The RNA polymerase transcribes mtDNA from the petite strain F11 in a nonrandom manner.

J Biol Chem, 1981 Feb 10, 256(3), 1466 - 72
Mitochondrial transcription complex from Saccharomyces cerevisiae; Levens D et al.; A DNA protein complex has been isolated from the mitochondria of Saccharomyces cerevisiae . The complex transcribes RNA complementary to mtDNA in a nonrandom manner . The RNA polymerase activity contained in the transcription complex is not dependent on the addition of exogenous template . The activity is rendered template-dependent by autolysis and can be further purified by heparin-Sepharose 4B chromatography . The activity is inhibited by heparin, Mn2+, and increasing ionic strength . The activity requires Mg2+ and ribonucleotides . The preferred template for the template dependent activity is poly{d(AT)} . The majority of the RNA synthesized by the transcription complex from endogenous DNA is complementary to the DNA strands directing the synthesis of the large and small ribosomal RNA . In yeast the 21 S and 14S rRNA genes are widely separated, therefore the transcription of these two regions but not of the intervening regions by the transcription complex suggests the existence of at least two transcriptional promoters on the yeast mitochondrial genome.

J Biol Chem, 1981 Feb 10, 256(3), 1063 - 6
Isolation of eukaryotic initiation factor 2 from yeast Saccharomyces cerevisiae; Baan RA et al.; Protein synthesis initiation factor eIF-2 has been isolated from the high speed supernatant fraction of the yeast Saccharomyces cerevisiae . This purification steps include ammonium sulfate fractionation, column chromatography on Sephacryl 300 and hydroxyapatite, and glycerol gradient sedimentation . Electrophoresis of the purified factor on sodium dodecyl sulfate polyacrylamide gels reveals three polypeptides with a total Mr of 127,000 . The molecular weights of the subunits are 31,000, 46,500, and 49,600 . The pI of each of these subunits is 4.5, 7.3, and 8.6, respectively . The stoichiometry of the subunits varies from 1:1:1 to 1:0.25:1 suggesting that the active factor may be composed of only two subunits with total Mr of 80,000 . The factor is part of a high molecular weight complex during the first steps of the purification . Prior to chromatography on Sephacryl, this complex is broken up in high salt . The activity of the factor is stabilized by inclusion of GDP in all buffers during the preparation and is stimulated by magnesium ion.

J Gen Microbiol, 1981 Feb, 122(Pt 2), 281 - 7
Genetic and biochemical properties of thialysine-resistant mutants of Saccharomyces cerevisiae; Zwolshen JH et al.; Three groups of lysine-excreting, thialysine-resistant mutants of Saccharomyces cerevisiae were derived from the wild-type strain (X2180) by mutagenic treatment and selected on the basis of a cross-feeding assay . Mutants MNNG2-9, MNNG2-27, MNNG2-39 and MNNG2-62 (group 1) exhibited a 2:2 segregation for thialysine resistance following mating with a wild-type strain and a lower than wild-type lysyl-tRNA synthetase activity; the thialysine-resistant phenotype was dominant in specific hybrids . Mutant MNNG2-2 (group II) was similar to group I mutants except that the thialysine-resistant phenotype was recessive in the hybrid . Mutant MNNG3-142 (group III) exhibited an irregular ratio of segregation of thialysine resistance and a significantly lower lysyl-tRNA synthetase activity; the thialysine-resistant phenotype was recessive in the hybrid . The growth of both group I and group III mutants was temperature-sensitive . The thialysine-resistant mutants exhibited pleiotropic properties including the increased production and excretion of lysine, thermosensitive growth and an impairment of lysyl-tRNA synthetase activity.

Mol Cell Biol, 1981 Feb, 1(2), 83 - 93
Isolation and characterization of dominant mutations resistant to carbon catabolite repression of galactokinase synthesis in Saccharomyces cerevisiae; Matsumoto K et al.; Seven dominant mutations showing greatly enhanced resistance to the glucose repression of galactokinase synthesis have been isolated from GAL81 mutants, which have the constitutive phenotype but are still strongly repressible by glucose for the synthesis of the Leloir enzymes . These glucose-resistant mutants were due to semidominant mutations at either of two loci, GAL82 and GAL83 . Both loci are unlinked to the GAL81- gal4, gal80, or gal7 X gal10 X gal1 locus or to each other . The GAL83 locus was mapped on chromosome V at a site between arg9 and cho1 . The GAL82 and GAL83 mutations produced partial resistance of galactokinase to glucose repression only when one or both of these mutations were combined with a GAL81 or a gal80 mutation . The GAL82 and GAL83 mutations are probably specific for expression of the Leloir pathway and related enzymes, because they do not affect the synthesis of alpha-D-glucosidase, invertase, or isocitrate lyase.

Mol Cell Biol, 1981 Feb, 1(2), 153 - 7
RAD6+ gene of Saccharomyces cerevisiae codes for two mutationally separable deoxyribonucleic acid repair functions; Tuite MF et al.; The response of two mutant alleles of the RAD6+ gene of Saccharomyces cerevisiae to the ochre translational suppressor SUQ5 was determined . Both the ultraviolet sensitivity phenotype and the deficiency in ultraviolet-induced mutagenesis phenotype of the rad6-1 allele were suppressed in a {psi+} background . For the rad6-3 allele, only the ultraviolet-sensitivity phenotype was suppressible in a {psi+} background . An SUQ5 rad6-3 {psi+} strain that was examined showed the normal rad6-3 deficiency in ultraviolet-induced mutagenesis . We propose that the RAD6+ gene is divided into two cistrons, RAD6A and RAD6B . RAD6A codes for an activity responsible for the error-prone repair of ultraviolet-induced lesions in deoxyribonucleic acid but is not involved in a cell's resistance to the lethal effects of ultraviolet light . RAD6B codes for an activity essential for error-free repair of potentially lethal mutagenic damage.

Experientia, 1981 Jan 15, 37(1), 39 - 40
Effect of growth factor deficiency on nystatin sensitivity in Saccharomyces cerevisiae; Jirku V et al.; The nystatin sensitivity, as well as the levels of total sterols and individual phospholipids of Saccharomyces cerevisiae are influenced by variations in the supply of growth factors.

Experientia . 1981 Jan 15;37(1):36.
Inhibitory effect of 5-bromo-6-azauracil on growth and cell division of Saccharomyces cerevisiae; Jirku V et al.; The most marked effect of 5-bromo-6-azauracil (BrAzU) on yeast cells is to cause cell lysis . The inhibition of the lytic process is delayed and this delay coincides in time with the capacity of preformed pyrimidines to reverse the effect of BrAzU.

Microbios, 1981, 31(125-126), 135 - 40
Regulation of catalase biosynthesis in Saccharomyces cerevisiae: factor repressing catalase biosynthesis; Martinez J et al.; A factor which represses the catalase biosynthesis in yeast has been demonstrated in Saccharomyces cerevisiae . This factor can be obtained from yeast cells having both low and normal catalase levels, and is unable to enter the intact cytoplasmic membrane . Moreover, the factor-containing cell extracts obtained either from acatalasemic mutants or normal strains grown in catalase repressive conditions showed higher activity than those obtained from normal strains after being cultured in permissive conditions.

Mol Biol (Mosk), 1981 Jan-Feb, 15(1), 124 - 31
{Induction of saccharomyces cerevisiae extrachromosomal-DNA}; Larionov VL et al.; In Saccharomyces yeast the significant portion of rRNA genes may be located as extrachromosomal copies . Extrachromosomal rDNA is represented by covalently-closed circular molecules with a major class contour length of 3.3 micrometer (the same as the length of one rDNA repeating unit) . Covalently-closed circular rDNA can only be isolated from the stationary phase yeast culture (more than 60 hours of growth) . Electron microscopy revealed several discrete length classes of circular molecules . Three of them are 3.3, 6.6 and 9.9 micrometers . Circular molecules were found with contour length less than 3.3 micrometers (1.1, 2.0 and 2.6 micrometers) . The proportion of these molecules is 8% from the total number.

Mol Gen Genet, 1981, 184(3), 471 - 8
Characterization of postreplication repair in Saccharomyces cerevisiae and effects of rad6, rad18, rev3 and rad52 mutations; Prakash L; Postreplication repair of nuclear DNA was examined in an excision defective haploid strain of yeast lacking mitochondrial DNA (rad1 rho 0) . The size of the DNA synthesized in cells exposed to various fluences of ultraviolet light (UV) corresponds approximately to the average interdimer distance in the parental DNA . Upon further incubation of cells following exposure to 2.5 J/m2, the DNA increases in size; by 4 h, it corresponds to DNA from uniformly labeled cells . The alkaline sucrose sedimentation pattern of DNA pulse labeled at various times after UV irradiation, for up to 4 h, does not change substantially, indicating that dimers continue to block DNA replication . A significant amount of postreplication repair requires de novo protein synthesis, as determined by its inhibition by cycloheximide . The rad6 mutant does not carry out postreplication repair, the rad18 and rad52 mutants show great inhibition while the rev3 mutation does not affect postreplication repair . Both recombinational and nonrecombinational repair mechanisms may function in postreplication repair and most of postreplication repair is error free.

Mol Gen Genet, 1981, 184(3), 410 - 5
Recombination and mutagenesis in rad6 mutants of Saccharomyces cerevisiae: evidence for multiple functions of the RAD6 gene; Montelone BA et al.; The rad6-1 and rad6-3 mutants are highly UV sensitive and show an increase in spontaneous and UV induced mitotic heteroallelic recombination in diploids . Both rad6 mutants are proficient in spontaneous and UV induced unequal sister chromatid recombination in the reiterated ribosomal DNA sequence and are deficient in UV induced mutagenesis . In contrast to the above effects where both mutants appear similar, rad6-1 mutants are deficient in sporulation and meiotic recombination whereas rad6-3 mutants are proficient . The differential effects of these mutations indicate that the RAD6 gene is multifunctional . The possible role of the RAD6 gene in error prone excision repair of UV damage during the G1 phase of the cell cycle in addition to its role in postreplication repair is discussed.

Mol Gen Genet, 1981, 183(3), 459 - 62
Recessive and dominant genes controlling inducible sexual agglutinability in Saccharomyces cerevisiae; Nakagawa Y et al.; We have characterized the two dominant genes, IND 1 and IND 2, responsible for inducible sexual agglutinability . The strains carrying these genes differ from the inducible strains carrying the recessive gene, saa 1 in the following points . The former strains produce agglutination substance at 22 degree, 28 degree, and 37 degree C only response to sex pheromone of the opposite mating type, but the latter strains produce the substance constitutively without the pheromone at 22 degree C, only in response to the pheromone at 28 degree C, and do not produce the substance, even in the presence of the pheromone, at 37 degree C . We suggest that strains carrying one of the dominant, inducible genes are wild type and have a pheromone-controlled regulatory system of sexual agglutinability.

Mol Gen Genet, 1981, 183(1), 85 - 92
Genetic and biochemical characterization of mutants of Saccharomyces cerevisiae blocked in six different steps of heme biosynthesis; Urban-Grimal D et al.; Heme-deficient mutants of Saccharomyces cerevisiae have been isolated from two isogenic strains with the use of an enrichment method based on photodynamic properties of Zn-protoporphyrin . They defined seven non-overlapping complementation groups . A mutant representative of each group was further analysed . Genetic analysis showed that each mutant carried a single nuclear recessive mutations . Biochemical studies showed that the observed accumulation and/or excretion of the different heme synthesis precursors by the mutant cells correlated well with the enzymatic deficiencies measured in acellular extracts . Six of the seven mutants were blocked in a different enzyme activity: 5-aminolevulinate synthase, porphobilinogen synthase, uroporphyrinogen I synthase, uroporphyrinogen decarboxylase, coproporphyrinogen III oxidase and ferrochelatase . The other mutant had the same phenotype as the mutant deficient in ferrochelatase activity . However, it possessed a normal ferrochelatase activity when measured in vitro, so this mutant was assumed to be deficient in protoporphyrinogen oxidase activity or in the transport and/or reduction of iron . The absence of PBG synthesis led to a total lack of uroporphyrinogen I synthase activity . The absence of heme, the end product, led to an important increase of coproporphyrinogen III oxidase activity, while the activity of 5-aminolevulinate synthase, the first enzyme of the pathway, was not changed . These results are discussed in terms of possible modes of regulation of heme synthesis pathway in yeast.

Z Allg Mikrobiol, 1981, 21(7), 489 - 97
A comparison of the toxic effects of 2-deoxy-D-glucose and 2-deoxy-2-fluoro-D-hexoses on Saccharomyces cerevisiae cells and protoplasts; Biely P et al.; The toxicity to the cells and protoplasts of Saccharomyces cerevisiae of the sugar analogues modified at carbon 2 increases in the order 2-deoxy-D-glucose (DG), 2-deoxy-2-fluoro-D-glucose (FG) and 2-deoxy-2-fluoro-D-mannose (FM) . The fluorohexoses, similarly as DG, behave generally as analogues of both glucose and mannose, depending on the hexose used as a carbon source in the medium . Relative inhibitions of glucan and mannan synthesis in protoplasts were found to be dependent more on glucose and mannose used as the growth support than on the type of the sugar analogue . Certain degree of structural relationship of fluorohexoses to the corresponding natural hexoses was reflected in their effects on growth of intact cells . Growth on glucose was inhibited most effectively by FM, growth on mannose by FG . The data obtained support the view that the sugar analogues interfere mainly with the glucose-mannose interconversion catalyzed by hexosephosphateisomerases . A comparison of the effects of fluorohexoses and DG on the synthesis of extracellular invertase an intracellular alpha-glucosidase and alkaline phosphatase in protoplasts pointed to the fact that all three sugar analogues tested also participate in metabolic control of enzyme synthesis.

J Gen Microbiol, 1981 Jan, 122(Pt 1), 79 - 87
Characterization of two mutant strains of Saccharomyces cerevisiae deficient in coproporphyrinogen III oxidase activity; Bilinski T et al.; Two new haem-deficient mutants of Saccharomyces cerevisiae were isolated on the basis of their catalase deficiency . Mutant H11 accumulated and excreted coproporphyrin III and was completely deficient in haem; the cell-free extract had no coproporphyrinogen oxidase activity . Mutant H12 accumulated uroporphyrin to coproporphyrin III and excreted coproporphyrin III, and contained a small amount of haem; the cell-free extract had a residual coproporphyrinogen oxidase activity . The two mutations were allelic and the mutant phenotypes were under the control of a single, recessive nuclear gene.

J Gen Microbiol, 1981 Jan, 122(Pt 1), 151 - 4
Duplication cycle in filamentous forms of Saccharomyces cerevisiae; Thompson PW et al.; Saccharomyces cerevisiae strain S288C/l was grown in a glucose-limited chemostat . At the fastest growth rates filamentous forms constituted a small percentage of the total cell number and were presumed to arise from the failure of cells to undergo cell separation . The phenomenon seemed to be distinct from chain formation, dimorphism and pseudomycelial growth and showed extensive analogies with the duplication cycle described for the filamentous fungi.

Microbios, 1981, 30(121-122), 139 - 51
On the uptake of nystatin by Saccharomyces cerevisiae . 1 Basic considerations; Beezer AE et al.; The uptake of nystatin by sensitive Saccharomyces cerevisiae cells has been studied as a function of temperature and nystatin concentration . Following analysis of the data, kinetic rate constants and a monolayer capacity are calculated . This shows that c 13 X more nystatin is taken up than can be supported by the cell surface . The process is accompanied by an activation energy of 41.6 kJ mol-1 . It is suggested that these data support a rapid uptake process onto the cell wall followed by a slower diffusion process through the wall to the membrane.

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Last modified: May 25, 2005