S. cerevisiae

J Biol Chem, 1984 Mar 25, 259(6), 3805 - 11
Carbohydrate structure of Saccharomyces cerevisiae mnn9 mannoprotein; Tsai PK et al.; The neutral oligosaccharides from Saccharomyces cerevisiae mnn1 mnn9, mnn2 mnn9, and mnn9 mutant mannoproteins, and from mnn1 and wild type carboxypeptidase Y, have been characterized . The major oligosaccharide from the mnn1 mnn9 mutant, Man10GlcNAc, has the structure (formula; see text) whereas the largest oligosaccharide from the mnn9 mutant, Man13GlcNAc, has the structure (formula; see text) the differences being due to the mnn1 mutation . The smaller mnn9 homologs had lesser amounts of terminal alpha 1----3-linked mannose and may be precursors of the mature oligosaccharide . The mnn2 mutation had no effect on the mnn9 oligosaccharide structures . Carboxypeptidase Y and mnn9 oligosaccharides were identical, which suggests that the mnn9 mutation eliminates the differences in carbohydrate structure that distinguish intra- from extracellular mannoproteins . One mnn1 mnn9 oligosaccharide, Man11GlcNAc, retained the terminal alpha 1----2-linked mannose of the lipid-linked core precursor, which suggests that processing to give the larger oligosaccharides can occur without removal of this unit . A smaller mnn1 mnn9 oligosaccharide, Man9GlcNAc, was a mixture of isomers that must, in part, have arisen by action of an alpha 1----2-mannosidase.

Eur J Biochem, 1984 Mar 15, 139(3), 651 - 5
Isolation and characterization of a mutant from Saccharomyces cerevisiae lacking fructose 1,6-bisphosphatase; Gancedo C et al.; Mutants lacking fructose 1,6-bisphosphatase activity have been isolated from Saccharomyces cerevisiae and genetically purified . Mutants were unable to grow on gluconeogenic carbon sources . Revertants that grew on glycerol have regained the fructose 1,6-bisphosphatase activity, showing that there is no bypass in yeast for this enzymatic step . No significant differences were found in the growth of the mutants and the parental strains in other carbon sources . Other mutants lacking fructose 1,6-bisphosphatase activity but pleiotropically affected in the derepression of several enzymes sensitive to catabolite repression were also isolated . All the mutants isolated were of nuclear origin and defined three complementation groups.

J Cell Sci, 1984 Mar, 66, 21 - 38
Fluorescence microscopic studies of mitochondrial nucleoids during meiosis and sporulation in the yeast, Saccharomyces cerevisiae; Miyakawa I et al.; Configurational changes of mitochondria and mitochondrial nucleoids (mt-nucleoids) during meiosis and sporulation in the yeast, Saccharomyces cerevisiae, were examined using the mitochondrial membrane-binding fluorescent dye, dimethyl aminostyrylmethylpyridiniumiodine (DASPMI) and the DNA-binding fluorescent dye, 4',6-diamidino-2-phenylindole (DAPI) . In zygotes just after mating, mt-nucleoids were observed as many small discrete light spots in the cytoplasm . During meiosis in zygotes, mt-nucleoids at first coalesced with each other into a long string and then separated into spherical nucleoids in four spores . These changes paralleled those in mitochondria observed using DASPMI . The use of spheroplasts allowed us to examine the behaviour of mt-nucleoids at higher resolution and to identify several distinct meiotic prophase stages of the cell nucleus during early sporulation . In diploid spheroplasts at the stationary phase, 50-70 of the mt-nucleoids were observed to be separated from each other and each spherical mitochondrion contained only one mt-nucleoid . At the later stage of premeiotic DNA synthesis, a single branched giant mitochondrion was formed as a result of complete mitochondrial fusion . All of the mt-nucleoids were arranged in an array on a giant mitochondrion and coalesced into a string-like network . Through meiosis I and II, strings of mt-nucleoids were observed close to the dividing nuclei . At late meiosis II, a ring of mt-nucleoids enclosing each daughter nucleus was formed . In ascospores, discrete small nucleoids were visible close to each spore nucleus with a 'string-of-beads' appearance . Many mt-nucleoids were excluded from the ascospores and remained in the residual cytoplasm of the ascus.

J Gen Microbiol, 1984 Mar, 130 ( Pt 3), 615 - 24
Initiation of sporulation in Saccharomyces cerevisiae . Mutations preventing initiation; Calvert GR et al.; Mutants of Saccharomyces cerevisiae that are unable to initiate sporulation, but can continue vegetative growth under conditions in which the wild-type strain sporulates, have been isolated and characterized . The mutations arose spontaneously as suppressors of the spd1 mutations, restoring the ability of spd1 mutants to grow on glycerol, and also spontaneously in cultures of a wild-type diploid strain undergoing sporulation in continuous culture . The mutations all conferred asporogeny, and were recessive in this respect to the wild-type, but dominant in acting as suppressors of the spd1 mutation . They fell into three complementation groups which corresponded to three unlinked loci, designated spo50, spo51 and spo53 . None of these mutations was closely linked to the other initiation mutations defined by the spd1, spd3, spd4, cdc25, cdc28 loci, nor to the cell size control mutations whi1 and whi2 . Loose linkage was detected between spd1 and spo53, and spo50, spd3 and spo53 were linked to their respective centromeres . The spo50, spo51 and spo53 mutations are not nonsense suppressors . Mutations in all three genes conferred similar highly pleiotropic phenotypes including: asporogeny; dominant suppression of both spd1 and spd3 mutations; aberrant cell morphology and viability loss on starvation; constitutive ability to reduce tetrazolium (which is subject to carbon source repression in the wild-type); and complete repression of the synthesis of several polypeptides that are subject to carbon source repression in the wild-type strain and derepressed in spd1 mutants derepressed for sporulation . A diploid strain homozygous for the spo50 mutation did not undergo either premeiotic DNA replication or meiotic recombination when transferred to sporulation media.

Can J Microbiol, 1984 Mar, 30(3), 345 - 52
Protein synthesis in germinating Saccharomyces cerevisiae ascospores; Armstrong RL et al.; The uptake and incorporation of macromolecular precursors in germinating Saccharomyces cerevisiae ascospores were investigated . Addition of cycloheximide at various times during germination revealed that protein synthesis can occur within 20 min after the spores are shifted to glucose-containing media . The time of initiation of uptake and incorporation of several amino acids differed; this can be attributed to differing amino acid pool levels in the spores, as well as differing transport activities . Two-dimensional gel electrophoresis of proteins labeled with {35S}methionine for various 20-min periods after germination began showed at least one protein whose synthesis begins well after the bulk of the proteins.

Arch Microbiol, 1984 Mar, 137(3), 188 - 93
Selective inhibition of transition from sexual agglutination to zygote formation by ethyl N-phenylcarbamate in Saccharomyces cerevisiae; Hasegawa S et al.; Effects of ethyl N-phenylcarbamate (EPC) on the mating reaction of Saccharomyces cerevisiae were studied, with special attention on the effect on the alpha pheromone action . EPC inhibited zygote formation at a concentration which promoted induction of sexual agglutinability . EPC enhanced agglutinability induction by alpha pheromone, but inhibited alpha-pheromone-induced formation of large pearshaped cells in a mating type . The enhancement of agglutinability induction was accompanied with increased production of a agglutination substance and inhibition of alpha pheromone inactivation . EPC arrested the cell cycle of a cells probably in the step controlled by CDC19, CDC35, cAMP etc., just before the step controlled by CDC28, alpha pheromone etc.

J Cell Biol, 1984 Mar, 98(3), 934 - 45
Relationship of actin and tubulin distribution to bud growth in wild-type and morphogenetic-mutant Saccharomyces cerevisiae; Adams AE et al.; The distribution of actin in wild-type cells and in morphogenetic mutants of the budding yeast Saccharomyces cerevisiae was explored by staining cells with fluorochrome-labeled phallotoxins after fixing and permeabilizing the cells by several methods . The actin appeared to be localized in a set of cortical spots or patches, as well as in a network of cytoplasmic fibers . Bundles of filaments that may possibly correspond to the fibers visualized by fluorescence were observed with the electron microscope . The putative actin spots were concentrated in small and medium-sized buds and at what were apparently the sites of incipient bud formation on unbudded cells, whereas the putative actin fibers were generally oriented along the long axes of the mother-bud pairs . In several morphogenetic mutants that form multiple, abnormally elongated buds, the actin patches were conspicuously clustered at the tips of most buds, and actin fibers were clearly oriented along the long axes of the buds . There was a strong correlation between the occurrence of active growth at particular bud tips and clustering of actin spots at those same tips . Near the end of the cell cycle in wild-type cells, actin appeared to concentrate (as a cluster of spots or a band) in the neck region connecting the mother cell to its bud . Observations made using indirect immunofluorescence with a monoclonal anti-yeast-tubulin antibody on the morphogenetic mutant cdc4 (which forms multiple, abnormally elongated buds while the nuclear cycle is arrested) revealed the surprising occurrence of multiple bundles of cytoplasmic microtubules emanating from the one duplicated spindle-pole body per cell . It seems that most or all of the buds contain one or more of these bundles of microtubules, which often can be seen to extend to the very tips of the buds . These observations are consistent with the hypotheses that actin, tubulin, or both may be involved in the polarization of growth and localization of cell-wall deposition that occurs during the yeast cell cycle.

J Bacteriol, 1984 Mar, 157(3), 958 - 61
Asparaginase II of Saccharomyces cerevisiae: positive selection of two mutations that prevent enzyme synthesis; Kim KW et al.; A positive selection method, D-aspartic acid beta-hydroxamate resistance, was used to isolate Saccharomyces cerevisiae strains lacking the ability to synthesize asparaginase II . Of 100 such mutant strains, 93 exhibited mutations which were allelic with asp3, a previously characterized mutation . The other seven strains carried a new mutation, asp6 . The asp6 mutation segregated 2:2 in asp6 X wild-type crosses and assorted from the asp3 mutation in asp6 X asp3 crosses . All seven asp6 mutant isolates reverted at a relatively high frequency, whereas the asp3 mutant isolates did not revert under the same conditions . Various independent asp3 isolates were mated to give heteroallelic diploids, which when sporulated and spread on D-asparagine medium yielded no recombinant strains.

Mol Cell Biol, 1984 Mar, 4(3), 520 - 8
Temporal analysis of general control of amino acid biosynthesis in Saccharomyces cerevisiae: role of positive regulatory genes in initiation and maintenance of mRNA derepression; Penn MD et al.; In Saccharomyces cerevisiae, starvation for a single amino acid results in the derepression of enzyme activities in multiple amino acid biosynthetic pathways . Derepression is a consequence of increased transcription of the genes encoding these enzymes . Analysis of the kinetics of mRNA elevation established that derepression occurs within 5 min of a shift of the culture from rich medium to starvation medium . Any starvation condition was sufficient to trigger an initial high mRNA elevation; however, it was the severity of starvation which determined the steady-state mRNA levels that were subsequently established . The products of the positive regulatory genes AAS101, AAS103, and AAS2 were shown to be required in the initiation phase of this response, whereas the AAS102 gene product was required to maintain the new elevated steady-state mRNA levels . The AAS101 and AAS102 genes were cloned . Consistent with their respective roles in initiation and maintenance of derepression . AAS101 mRNA was found to be expressed at high levels in both rich and starvation media, whereas AAS102 mRNA was derepressed only under starvation conditions . The derepression of AAS102 mRNA is dependent on the AAS101 gene product.

Arch Microbiol, 1984 Mar, 137(3), 209 - 14
An examination of factors affecting the instability of Saccharomyces cerevisiae glucan synthetase in cell free extracts; Leal F et al.; Yeast beta(1--3) glucan synthetase is stimulated and stabilized by EDTA . Sucrose protects the enzyme from self-inactivation . Preincubation of cell free extracts at low sucrose concentrations indicates a slow transition of the enzyme towards dissociation . Transition kinetics at 30 degrees C and 0 degrees C in the presence and in the absence of sucrose are interpreted assuming that a subunit is thermolabile in the free state and that sucrose increases its stability . Magnesium is deleterous for glucan synthetase in cell-free extracts . Chaotropic agents inactivate glucan synthetase according to their capacity to solubilize and depolymerize biological compounds . Fluoride plays a special role in the activation of glucan synthetase . Its action appears to be dependent on the presence of GTP (or other nucleotides) . The role of all these agents on the activity and stability of the enzyme is interpreted in a unified scheme.

Biochim Biophys Acta, 1984 Feb 24, 781(1-2), 153 - 64
The formation of a defective small subunit of the mitochondrial ribosomes in petite mutants of Saccharomyces cerevisiae; Maheshwari KK et al.; The involvement of mitochondrial protein synthesis in the assembly of the mitochondrial ribosomes was investigated by studying the extent to which the assembly process can proceed in petite mutants of Saccharomyces cerevisiae which lack mitochondrial protein synthetic activity due to the deletion of some tRNA genes and/or one of the rRNA genes on the mtDNA . Petite strains which retain the 15-S rRNA gene can synthesize this rRNA species, but do not contain any detectable amounts of the small mitochondrial ribosomal subunit . Instead, a ribonucleoparticle with a sedimentation coefficient of 30 S (instead of 37 S) was observed . This ribonucleoparticle contained all the small ribosomal subunit proteins with the exception of the var1 and three to five other proteins, which indicates that the 30-S ribonucleoparticle is related to the small mitochondrial ribosomal subunit (37 S) . Reconstitution experiments using the 30-S particle and the large mitochondrial ribosomal subunit from a wild-type yeast strain indicate that the 30-S particle is not active in translating the artificial message poly(U) . The large mitochondrial ribosomal subunit was present in petite strains retaining the 21-S rRNA gene . The petite 54-S subunit is biologically active in the translation of poly(U) when reconstituted with the small subunit (37 S) from a wild-type strain . The above results indicate that mitochondrial protein synthetic activity is essential for the assembly of the mature small ribosomal subunit, but not for the large subunit . Since the var1 protein is the only mitochondrial translation product known to date to be associated with the mitochondrial ribosomes, the results suggest that this protein is essential for the assembly of the mature small subunit.

Nature, 1984 Feb 23-29, 307(5953), 740 - 2
Role of an upstream regulatory element in leucine repression of the Saccharomyces cerevisiae leu2 gene; Martinez-Arias A et al.; The expression of a number of eukaryotic genes has been shown to involve at least two sequences located upstream of the actual transcription unit: one of these sequences, centred on a widely conserved TATAAT sequence, is thought to be involved in determining the precise site of initiation of transcription; the other has a gene-specific sequence, can function at a variable distance upstream of the initiation site, and is involved in the regulation of transcription . By constructing beta-galactosidase gene fusions, to facilitate measuring gene expression in vivo, we have now defined a cis-acting regulatory element of the Saccharomyces cerevisiae leu2 gene . This element is located within a 280 base pair (bp) fragment which occurs 125 bp upstream of the leu2 translation initiation codon and which contains a short G + C-rich palindromic sequence . A fragment of the Escherichia coli transposable element Tn9 which contains a similar palindromic sequence can functionally replace the natural leu2 regulatory element . Our results are contrary to previous speculations that the leu2 gene is regulated by an attenuation mechanism.

Carbohydr Res, 1984 Feb 15, 125(2), 301 - 7
Analysis of linkage positions in Saccharomyces cerevisiae D-mannans by the reductive-cleavage method; Bowie JU et al.; The positions of linkage in the D-mannans derived from Saccharomyces cerevisiae X2180 and its mutants, mnn1, mnn2, and mnn4, were established by perethylation and subsequent reductive cleavage with triethylsilane in the presence of boron trifluoride etherate (BF3 . Et2O) or trimethylsilyl trifluoromethanesulfonate . With the latter as the catalyst, all glycosidic carbon-oxygen bonds were cleaved, to produce a mixture of ethylated 1,5-anhydro-D-mannitol derivatives . With BF3 . Et2O as the catalyst, 2-, 3-, and 6-linked residues were incompletely cleaved, and residues linked at both O-2 and O-6 were not cleaved at all . It was concluded that reductive cleavage is an attractive method for determination of the structure of polysaccharides.

Biochim Biophys Acta, 1984 Feb 15, 769(3), 601 - 10
Effects of phenothiazines on inhibition of plasma membrane ATPase and hyperpolarization of cell membranes in the yeast Saccharomyces cerevisiae; Eilam Y; The transmembranal potential, in Saccharomyces cerevisiae, has been calculated from the distribution ratio of the lipophilic cation tetraphenylphosphonium (TPP+) between the intracellular and extracellular water . Trifluoperazine at concentrations of 10 to 50 microM, caused a substantial increase in the membrane potential (negative inside) . This increase was observed only in the presence of a metabolic substrate and was eliminated by the addition of the protonophores 2,4-dinitrophenol and sodium azide, removal of glucose, replacement of glucose by the nonmetabolizable analog 3-O-methyl glucose, or by the addition of 100 mM KCl . An increase in 45CaCl2 accumulation from solutions of low concentrations (1 microM) was observed under all conditions where membrane potential was increased . Proton ejection activity was monitored by measurements of the rates of the decrease in the pH of unbuffered cell suspensions in the presence of glucose . Trifluoperazine inhibited the changes in medium pH; this inhibition was not the result of an increase in the permeability of cell membranes to protons since in the absence of glucose, trifluoperazine did not cause a change in the rate of pH change generated by proton influx . The activity of plasma membrane ATPase was measured in crude membrane preparations in the presence of sodium azide to inhibit mitochondrial ATPase . Trifluoperazine strongly inhibited the activity of the plasma membrane ATPase . The effect of phenothiazines on transport and on membrane potential reported in this study and in the previous one (Eilam, Y . (1983) Biochim . Biophys . Acta 733, 242-248) were observed only in the presence of a metabolic substrate . The possibility that energy is required for the uptake of phenothiazines into the cells was eliminated by results showing energy-independent uptake of {3H}chlorpromazine . The results strongly suggest that phenothiazines activate energy-dependent K+-extrusion pumps, which lead to increased membrane potential . Increased influx of calcium seems to be energized by membrane potential, and therefore stimulated under all conditions where membrane potential is increased . The analog which does not bind to calmodulin, trifluoperazine sulfoxide, had no effect on the cells, but the involvement of calmodulin in the processes altered by trifluoperazine cannot as yet, be determined.

Biochem J, 1984 Feb 15, 218(1), 147 - 55
gamma-Glutamyltransferase is not involved in the bulk uptake of amino acids, peptides or gamma-glutamyl-amino acids in yeast (Saccharomyces cerevisiae); Payne GM et al.; gamma-Glutamyltransferase activity has been measured in yeast (Saccharomyces cerevisiae) and shown to be associated mainly with the membrane fraction . A similar level of activity is found in a wild-type strain and in gap and gpp strains, the latter mutants being defective in the general amino acid and peptide permeases respectively . The activity is inhibited in whole cells by 6-diazo-5-oxo-L-norleucine (N2O-Nle), azaserine and serine-borate complex; this inactivation seemingly acts from without, for it is similar in (i) control and dicyclohexylcarbodi-imide-treated cells and in (ii) the wild-type and a gap mutant, a treatment and a mutation that it has been shown prevents uptake of the inhibitors . Thus a major portion of the gamma-glutamyltransferase activity appears to exist in a membrane-bound form that is orientated with its gamma-glutamyl-binding site facing the outside . Yeast cells in which gamma-glutamyltransferase has been inactivated by N2O-Nle show no significant change in their rates of uptake of a variety of amino acids, dipeptides and gamma-glutamyl-amino acids . The results preclude a major, direct role for gamma-glutamyltransferase in the transport of these substrates.

FEBS Lett, 1984 Feb 13, 167(1), 151 - 4
Coupling between phosphatidylinositol metabolism and cdc 28 gene product of Saccharomyces cerevisiae . On the possible mechanism of cdc 28 gene action; Dudani AK et al.; It was shown that the decrease in phosphatidylinositol (PI) content in cdc 28 G1-cells was due to a defect in inositol transport . This decrease in inositol transport was linked to microtubular function which was evident by the effect of a microtubular disrupting agent (colcemid) on inositol transport in stationary phase A364A cells . The involvement of PI in yeast G1 phase was further substantiated by the observation that o-phenanthroline, which blocks yeast cells in G1 phase, could inhibit inositol transport and PI levels as well . It is proposed that the regulation of PI metabolism is mediated by the gene cdc 28 and that microtubules may play a major role in the mechanism of action of this gene product.

Nucleic Acids Res, 1984 Feb 10, 12(3), 1627 - 40
Expression of Ty-lacZ fusions in Saccharomyces cerevisiae; Bowen BA et al.; We have determined the nucleotide sequence of about 520 bp spanning the 5' delta regions (Figure 1) of two Tyl elements . There is an open reading frame running out of the deltas for at least 180 nucleotides into the internal region of each element . The functional significance of these open reading frames has been tested by fusing them to a defective E.coli lacZ gene . Expression of B-galactosidase in yeast transformants containing these fusions shows that Tyl elements contain functional translation signals.

J Biol Chem, 1984 Feb 10, 259(3), 1375 - 7
Identification of Saccharomyces cerevisiae mutants deficient in DNA topoisomerase I activity; Thrash C et al.; Mutants of the yeast, Saccharomyces cerevisiae, deficient in DNA topoisomerase I activity have been identified . One mutant has normal topoisomerase I activity when assayed at 25 degrees C and about 20% of normal activity when assayed at 36 degrees C . Strains with this mutation grow normally at all temperatures tested . The mutation has been mapped to MAK1, a gene required for maintenance of killer RNA . Three previously isolated mak1 mutants exhibit less than 1% of normal topoisomerase I activity in our assay, but yet they grow normally . The implications of these results for the role of DNA topoisomerase I in the cell are discussed.

Gene, 1984 Feb, 27(2), 233 - 7
Molecular cloning of the ADE1 gene of Saccharomyces cerevisiae and stability of the transformants; Dimock K et al.; Plasmid YEp ( ADE1 )1a, containing a 2.7-kb Sau3A fragment of Saccharomyces cerevisiae DNA inserted at the BamHI site of the yeast shuttle vector pBTI -1 (Morris et al., 1981), results in high frequency, unstable transformation of ade 1 yeast strains . A second plasmid, YRp ( ADE1 )2, containing adjacent 0.5-kb and 3.0-kb BamHI fragments in pBR322 gave three types of yeast transformants: (1) transformants carrying extrachromosomal copies of the plasmid which indicate the presence of a functional ars sequence, (2) transformants indistinguishable from ade 1 strains by hybridization analysis, and (3) a transformant carrying a multimeric form of YRp ( ADE1 )2 . Cells transformed with either of the plasmids are free of the red pigment characteristic of ade 1 mutants and indicate potential for direct colour-based selection of yeast transformants using ADE1 plasmids.

Proc Natl Acad Sci U S A, 1984 Feb, 81(4), 1149 - 53
Specificity of polyamine requirements for the replication and maintenance of different double-stranded RNA plasmids in Saccharomyces cerevisiae; Tyagi AK et al.; We have shown previously that the M1 double-stranded (ds) RNA (i.e., the killer plasmid {KIL-k1}) that codes for a protein toxin requires spermidine or spermine for its replication . We now report that replication of two other ds RNA plasmids of yeast also requires polyamines: (i) M2 ds RNA {( KIL-k2}) and (ii) L-A-E, a ds RNA plasmid carrying the non-Mendelian genetic element {EXL} . Putrescine alone is sufficient to maintain L-A-E but is not sufficient to maintain either M1 ds RNA or M2 ds RNA, which require either spermidine or spermine . Once M1 or M2 or L-A-E is lost, it cannot be restored by the addition of polyamines . In contrast, L-A-HN, a ds RNA molecule that carries the cytoplasmic genes {HOK} and {NEX}, is not lost during polyamine deprivation . It is striking that polyamine deprivation differentially affects L-A-E and L-A-HN, even though these two ds RNA molecules have more than 99% homology . L-C, which is the same size as L-A but very different in sequence, is also not lost on polyamine starvation.

Proc Natl Acad Sci U S A, 1984 Feb, 81(4), 1144 - 8
Molecular cloning of hormone-responsive genes from the yeast Saccharomyces cerevisiae; Stetler GL et al.; A method for identifying yeast genes whose transcription is differentially regulated was developed . The technique is based on incorporation of the analog 4-thiouridine into nascent RNAs, which allows their purification . The purified RNAs are used to prepare cDNA copies for screening of genomic DNA libraries by hybridization . Using this procedure, several cloned yeast DNA segments were found whose transcription in MATa haploids in vivo is apparently modulated in dramatic fashion within 10-15 min after exposure to the mating pheromone, alpha factor . Subsequent analysis indicated that these sequences fall into three major classes: (i) genes expressed in vegetatively growing cells that are no longer transcribed after alpha-factor administration ("turn-off" genes); (ii) genes whose expression is increased 10- to 20-fold after exposure of the MATa cells to alpha factor ("turn-up" genes); and (iii) genes that are expressed only after alpha-factor treatment ("turn-on" genes) . The first class may encode products required for cell cycle progression; the third class may code for products uniquely involved in the mating process.

Mol Cell Biol, 1984 Feb, 4(2), 260 - 7
Primary structure of the Saccharomyces cerevisiae GAL4 gene; Laughon A et al.; The GAL4 gene encodes a positive regulator of the galactose-inducible genes in Saccharomyces cerevisiae . Recently, GAL4 has been cloned and its 2.8-kilobase mRNA has been identified . We report here the DNA sequence of GAL4 and the mapping of the 5' and 3' ends of its transcripts . The region sequenced contains a single open reading frame, 881 codons long, which could encode a 99,350-dalton protein . The 5' ends of the GAL4 transcripts fall into two clusters . Transcripts which begin at the upstream cluster would encode the 99,350-dalton protein, whereas those starting at the downstream cluster may result in the synthesis of a shorter, 91,600-dalton protein . The putative GAL4 proteins contain an amino acid sequence near their amino termini which resembles a DNA-binding motif found in bacterial and phage repressors and gene activator proteins.

Genetics, 1984 Feb, 106(2), 165 - 83
Enhanced gene conversion and postmeiotic segregation in pachytene-arrested Saccharomyces cerevisiae; Davidow LS et al.; Previous study has demonstrated that incubation of yeast cells of strain AP-1 in sporulation medium at 36 degrees permits them to begin meiosis but that they become arrested at pachytene and undergo enhanced intragenic recombination between ade2 heteroalleles . Tetrad analysis was undertaken to characterize the altered program of meiotic recombination more widely . In one set of experiments, pachytene-arrested cells were permitted to resume sporulation upon transfer to the permissive temperature . In the resulting asci, both postmeiotic segregation and gene conversion were increased several-fold at a number of loci relative to unarrested controls, whereas reciprocal recombination increased two- to threefold . Another set of experiments analyzed the genetic consequences of inducing the pachytene-arrested cells to revert directly to mitotic growth without completion of meiosis . The appearance of homozygous sectors from heterozygous markers revealed that these cells had become committed to appreciable recombination but that reciprocal exchange was less frequent than in normal asci . Taken together, the data indicate that pachytene arrest rendered the cells committed to enhanced recombination upon resumption of sporulation but that most of the crossing over did not occur until release from the arrest.--The genetic basis of pachytene arrest by AP-1 was investigated by mating each of its parents with progeny of strain Y55, which is able to sporulate at 36 degrees . Both of these diploids sporulated at 36 degrees, and asci from the one studied further exhibited 2:2 segregation of the sporulation defect, indicating that pachytene arrest is dependent on a recessive, temperature-sensitive allele at a chromosomal locus.

J Cell Biol, 1984 Feb, 98(2), 678 - 84
Bud formation by the yeast Saccharomyces cerevisiae is directly dependent on "start"; Singer RA et al.; Cells of the yeast Saccharomyces cerevisiae, which bear a cdc4 gene mutation, arrest early in the cell cycle but continue to produce buds in a periodic fashion . We show here that this periodic bud formation by cells already arrested at the CDC4 step is inhibited if the cell cycle regulatory step "start" is also specifically blocked by mutation or by the presence of the yeast mating pheromone alpha-factor . Thus, the characteristic periodic bud formation by cdc4 mutant cells requires the continued ability to perform start . This finding raises questions concerning the nature of start; these issues are discussed.

J Bacteriol, 1984 Feb, 157(2), 475 - 83
Sterol methylation in Saccharomyces cerevisiae; McCammon MT et al.; Various nystatin-resistant mutants defective in S-adenosylmethionine: delta 24-sterol-C-methyltransferase (EC 2.1.1.41) were shown to possess alleles of the same gene, erg6 . The genetic map location of erg6 was shown to be close to trp1 on chromosome 4 . Despite the single locus for erg6, S-adenosylmethionine: delta 24-sterol-C-methyltransferase enzyme activity was found in three separate fractions: mitochondria, microsomes, and the "floating lipid layer." The amount of activity in each fraction could be manipulated by assay conditions . The lipids and lipid synthesis of mutants of Saccharomyces cerevisiae defective in the delta 24-sterol-C-methyltransferase were compared with a C5(6) desaturase mutant and parental wild types . No ergosterol (C28 sterol) could be detected in whole-cell sterol extracts of the erg6 mutants, the limits of detection being less than 10(-11) mol of ergosterol per 10(8) cells . The distribution of accumulated sterols by these mutants varied with growth phase and between free and esterified fractions . The steryl ester concentrations of the mutants were eight times higher than those of the wild type from exponential growth samples . However, the concentration of the ester accumulated by the mutants was not as great in stationary-phase cells . Whereas the head group phospholipid composition was the same between parental and mutant strains, strain-dependent changes in fatty acids were observed, most notably a 40% increase in the oleic acid content of phosphatidylethanolamine of one erg6 mutant, JR5.

Genetics, 1984 Feb, 106(2), 185 - 205
Meiotic and mitotic behavior of dicentric chromosomes in Saccharomyces cerevisiae; Haber JE et al.; Meiotic recombination between a circular and a linear chromosome in Saccharomyces cerevisiae has been investigated . The circle was a haploid-viable derivative of chromosome III constructed by joining regions near the two chromosome ends via a recombinant DNA construction: (HMR/MAT-URA3-pBR322-MAT/HML) and was also deleted for MAL2 (which therefore uniquely marks a linear chromosome III) . Recombination along chromosome III was measured for eight intervals spanning the entire length of the circular derivative . Only 25% of all tetrads from a ring/rod diploid contained four viable spores . These proved to be cases in which there was either no recombination along chromosome III or in which there were two-strand double crossovers or higher order crossovers that would not produce a dicentric chromosome.--At least half of the tetrads with three viable spores included one Ura+ Mal+ spore that was genetically highly unstable . The Ura+ Mal+ spore colonies gave rise to as many as seven genetically distinct, stable ("healed") derivatives, some of which had lost either URA3 or MAL2 . Analysis of markers on chromosome III suggests that dicentric chromosomes frequently do not break during meiosis but are inherited intact into a haploid spore . In mitosis, however, the dicentric chromosome is frequently broken, giving rise to a variety of genetically distinct derivatives . We have also shown that dicentric ring chromosomes exhibit similar behavior: at least half the time they are not broken during meiosis but are broken and healed during mitosis.--The ring/rod diploid can also be used to determine the frequency of sister chromatid exchange (SCE) along an entire yeast ring chromosome . We estimate that an unequal number of SCE events occurs in approximately 15% of all cells undergoing meiosis . In contrast, the mitotic instability (and presumably SCE events) of a ring chromosome is low, occurring at a rate of about 1.2 X 10(-3) per cell division.

Biochim Biophys Acta, 1984 Jan 31, 784(2-3), 102 - 7
A cytoplasmic, cyclic nucleotide-independent casein kinase II from Saccharomyces cerevisiae; Kudlicki W et al.; Two molecular forms of casein kinase II (an ATP: protein phosphotransferase, EC 2.7.1.37) from yeast were isolated and characterized . The first form was composed of three polypeptide subunits with molecular weights of 41000, 37000 and 24000 . The second form contained two larger polypeptides and lacked an autophosphorylatable 24 kDa subunit . The properties of both enzyme forms were found to be practically the same in respect to the substrate and phosphate donor specificities, kinetics, their sensitivity to heparin, etc . The results obtained strongly indicate that isolated yeast casein kinase II does not necessarily require the smallest subunit for the enzyme activity.

J Biol Chem, 1984 Jan 25, 259(2), 878 - 83
Physicochemical and kinetic properties of acid phosphatase from Saccharomyces cerevisiae; Barbaric S et al.; Acid phosphatase from yeast Saccharomyces cerevisiae was purified, and its physicochemical and kinetic properties were investigated . The sedimentation coefficient has been determined to be s0(20,w) = 13.6 S . The diffusion constant has been found to be 3.9 X 10(-7) cm2s-1, and the calculated partial specific volume was v = 0.663 cm3/g . From these data, a molecular weight of 252,000 was calculated . Electrophoresis on gel slabs, with a linear concentration gradient of polyacrylamide (4-30%), showed size heterogeneity of the native enzyme preparation and indicated an apparent molecular weight in the range of 170,000 to 360,000 . In the presence of sodium dodecyl sulfate, the molecular weight was in the range of 82,000 to 165,000, indicating dimeric structure of the native enzyme, which was confirmed by cross-linking experiments . Isoelectric focusing demonstrated charge heterogeneity of enzyme preparation . From CD spectrum it was calculated that the enzyme contains about 29% of alpha-helical structure . Excitation at 278 nm gave an emission fluorescence spectrum with a maximum at 340 nm . Amino acid analysis revealed a high content of aspartic acid, serine, and threonine . Glycine is found as the NH2-terminal amino acid . Initial velocity dependence on substrate concentration, as well as on pH, and thermostability studies indicated the presence of at least two enzyme forms in the preparation.

Nucleic Acids Res, 1984 Jan 25, 12(2), 1049 - 68
Characterization of human chromosomal DNA sequences which replicate autonomously in Saccharomyces cerevisiae; Montiel JF et al.; We have characterised two restriction fragments, isolated from a "shotgun" collection of human DNA, which function as autonomously replicating sequences (ARSs) in Saccharomyces cerevisiae . Functional domains of these fragments have been defined by subcloning and exonuclease (BAL 31) deletion analysis . Both fragments contain two spatially distinct domains . One is essential for high frequency transformation and is termed the Replication Sequence (RS) domain, the other, termed the Replication Enhancer (RE) domain, has no inherent replication competence but is essential for ensuring maximum function of the RS domain . The nucleotide sequence of these domains reveals several conserved sequences one of which is strikingly similar to the yeast ARS consensus sequence.

J Biol Chem, 1984 Jan 25, 259(2), 1288 - 93
Regulation of NAD-dependent glutamate dehydrogenase by protein kinases in Saccharomyces cerevisiae; Uno I et al.; The active NAD-dependent glutamate dehydrogenase of wild type yeast cells fractionated by DEAE-Sephacel chromatography was inactivated in vitro by the addition of either the cAMP-dependent or cAMP-independent protein kinases obtained from wild type cells . cAMP-dependent inhibition of glutamate dehydrogenase activity was not observed in the crude extract of bcy1 mutant cells which were deficient in the regulatory subunit of cAMP-dependent protein kinase . The cAMP-dependent protein kinase of CYR3 mutant cells, which has a high K alpha value for cAMP in the phosphorylation reaction, required a high cAMP concentration for the inactivation of NAD-dependent glutamate dehydrogenase . An increased inactivation of partially purified active NAD-dependent glutamate dehydrogenase (Mr = 450,000) was observed to correlate with increased phosphorylation of a protein subunit (Mr = 100,000) of glutamate dehydrogenase . The phosphorylated protein was labeled by an NADH analog, 5'-p-fluorosulfonyl{14C}benzoyladenosine . Activation and dephosphorylation of inactive NAD-dependent glutamate dehydrogenase fractions were observed in vitro by treatment with bovine alkaline phosphatase or crude yeast cell extracts . These results suggested that the conversion of the active form of NAD-dependent glutamate dehydrogenase to an inactive form is regulated by phosphorylation through cAMP-dependent and cAMP-independent protein kinases.

Biochemistry, 1984 Jan 17, 23(2), 390 - 6
Acidic proteins of the large ribosomal subunit in Saccharomyces cerevisiae . Effect of phosphorylation; Vidales FJ et al.; Three strongly acidic proteins with pIs between 3.0 and 3.5 have been detected and purified from an ammonium-ethanol extract of Saccharomyces cerevisiae ribosomes . The three proteins, called L45, L44, and L44', have a similar amino acid composition, but differences were shown by tryptic peptide analysis . Nevertheless, the three polypeptides show total cross-reaction to antisera raised against one of them . Protein L44' is very unstable in the extract when treated at the basic pH 9.2, due to an enzymatic process not yet clarified . When purified, the protein is, however, stable . In solution, the proteins are present as dimers, as verified by ultracentrifugation, column filtration, and photochemical cross-linking . The tendency to dimerization is much lower in the case of protein L44' . On the average, 3.2 copies of these proteins are detected per ribosome . The proteins are monophosphorylated when present in the ribosome . Phosphorylation seems to regulate the affinity of the polypeptides for the particles because unphosphorylated proteins bind poorly to the ribosomes deprived of the acidic proteins . Since these proteins are unphosphorylated when present in the cytoplasm {Zinker, S . (1980) Biochim . Biophys . Acta 606, 76-82; Sanchez-Madrid, F., Vidales, F . J., & Ballesta, J . P . G . (1981) Eur . J . Biochem . 114, 609-613}, a regulatory mechanism of the ribosomal function based on a phosphorylation-dephosphorylation process of the acidic proteins is being studied.

Symp Soc Exp Biol, 1984, 38, 123 - 59
Molecular mechanisms of recombination in Saccharomyces cerevisiae: testing mitotic and meiotic models by analysis of hypo-rec and hyper-rec mutations; Esposito MS; Recombination in the yeast Saccharomyces cerevisiae has been the subject of extensive genetic studies documenting the general properties of intragenic and intergenic recombination and the differences between mitotic and meiotic gene conversion and reciprocal exchange . Spontaneous mitotic and meiotic events differ in the time of onset of recombination relative to chromosomal replication, symmetry versus asymmetry of putative heteroduplex DNA regions, polarity of conversion of intragenic markers, and the lengths of DNA segments that undergo coincident conversion . The differences observed and the properties of yeast rec mutations provide evidence for multiple modes or pathways of mitotic and meiotic recombination . Several molecular models of recombination have been proposed to account for the basic parameters of genetic recombination and the differences between mitotic and meiotic recombination . Since the models differ with respect to the partial reactions comprising recombination they predict the isolation of different classes of hypo-recombination and hyper-recombination rec mutants . We have isolated a broad spectrum of yeast REC gene mutations that includes both hyper-rec and hypo-rec mutants . Five phenotypic classes of rec variants have been identified based upon their effects on spontaneous mitotic gene conversion and intergenic recombination . Their characteristics demonstrate that mitotic gene conversion and intergenic recombination are under independent as well as coordinate genetic control . Four gene mutations affecting recombination rad50, rad52, rem1 and spo11 have been extensively examined in several laboratories and illustrate the information that can be obtained by characterization of double mutant strains, detailed genotypic analysis of recombinants, and studies of meiotic recombination in cells in which the reductional division of meiosis has been bypassed by the spo13 mutation.

Folia Microbiol (Praha), 1984, 29(6), 441 - 9
Papulacandin B: inhibitor of biogenesis of (1----3)-beta-D-glucan fibrillar component of the cell wall of Saccharomyces cerevisiae protoplasts; Kopecka M; The effect of papulacandin B on regenerating protoplasts of Saccharomyces cerevisiae was studied by light and electron microscopy . In liquid media it inhibited the biogenesis of (1----3)-beta-D-glucan fibrillar nets; as a result, the protoplasts did not grow polarly but only spherically . The effect was reversible . Instead of the nets the inhibited protoplasts synthesized only individual microfibrils soluble in hydroxide; these were not joined in the nets and were partially masked by amorphous material . The microfibrils disintegrated after lysis and did not maintain the shape of protoplasts . Protoplasts inhibited in solid media grew spherically up to 25 micron but they did not divide or revert, in spite of forming cell walls . These walls were amorphous and fragile and they disintegrated during preparation . Papulacandin B did not decrease the viability of protoplasts and did not interfere with their growth, biogenesis of alkali-soluble glucan microfibrils or amorphous wall matrix . It inhibited specifically the synthesis of alkali-insoluble branched (1----3)-beta-D-glucan, a necessary building unit required for the formation of the fibrillar component of the cell wall responsible for the cell wall shape, its rigidity and tensile strength.

Mol Gen Genet, 1984, 198(1), 62 - 8
Leakiness of termination codons in mitochondrial mutants of the yeast Saccharomyces cerevisiae; Weiss-Brummer B et al.; Seven mutants in exon 1 of the mitochondrial cob gene in yeast are described with respect to their translation products, RNA pattern, and deoxyribonucleotide sequence alteration(s) . Sequence analysis of the mutations, which previously were shown to cause premature termination of apocytochrome b, revealed that two of them directly transform sense codons to chain-termination codons, whereas the other four are frame-shift mutations (+1/-1, insertions/deletions) . Only the latter mutants are found to be leaky in that (a) RNA splicing occurs, and (b) in three of them, to a minor degree an apocytochrome b homologue is synthesized, which, however, does not lead to respiratory competence . Both require translation through exon 1 into downstream introns to produce 'RNA maturases' necessary for splicing the primary transcript (Lazowska et al . 1980; Weiss-Brummer et al . 1982) . These and other previously published data show that mitochondrial frame-shift mutants tend to be leaky to a variable degree . Several possible mechanisms of 'frame-shift suppression' are discussed.

Mol Gen Genet, 1984, 198(1), 50 - 4
Cloning of hexokinase isoenzyme PI from Saccharomyces cerevisiae: PI transformants confirm the unique role of hexokinase isoenzyme PII for glucose repression in yeasts; Entian KD et al.; Hexokinase isoenzyme PI was cloned using a gene pool obtained from a yeast strain having only one functional hexokinase, isoenzyme PI . The gene was characterized using 20 restriction enzymes and located within a region of 2.0 kbp . The PI plasmid strongly hybridized with the PII plasmids isolated previously (Frohlich et al . 1984) . Hence there was a close relationship between the two genes, one of which must have been derived from the other by gene duplication . In contrast, glucose repression was restored only in hexokinase PII transformants; PI transformants remained non-repressible . This observation provided additional evidence for the hypothesis of Entian (1980) that only hexokinase PII is necessary for glucose repression . Furthermore, glucose phosphorylating activity in PI transformants exceeded that of wild-type cells, giving clear evidence that the phosphorylating capacity is not important for glucose repression.

Mol Gen Genet, 1984, 195(1-2), 234 - 7
Genetic analysis of Saccharomyces cerevisiae SY 15 relaxed mutant; Stateva LI et al.; Evidence is presented showing that the relaxed phenotype of the SY15 mutant is determined by one nuclear recessive mutation . The most characteristic patterns of the relaxed phenotype in yeast - rRNA accumulation and rRNA processing in the absence of protein synthesis were found to segregate together in first and second generation crosses . Therefore, the interruption of rRNA processing that occurs after starvation for a required amino acid is a pleiotropic manifestation of the stringent control itself . It is suggested that the locus for the stringent response in Saccharomyces cerevisiae (designated STR) coordinates the synthesis of rRNA on transcriptional and post-transcriptional levels.

Mol Gen Genet, 1984, 195(1-2), 139 - 43
Gene conversion at different points in the mitotic cycle of Saccharomyces cerevisiae; Fabre F et al.; In the Saccharomyces cerevisiae mitotic cycle, the timing of radiation-induced gene conversion has been studied using thermosensitive cell division cycle mutants . The cells were found to perform conversion at different G1 or post-replication steps . A lower yield in induction is found during the G2 phase and is explained by the competition for recombinational repair between sister chromatids and homologous chromosomes . The results are discussed in relation to repair.

Mol Gen Genet, 1984, 196(1), 158 - 66
Cytochrome b of cob revertants in yeast . 1 . Isolation and characterization of revertants derived from cob exon mutants of Saccharomyces cerevisiae; Burger G; About 300 revertants were derived from 44 cob- mutants, mapping in the structure coding regions (exon 1, 3, 4, 5, or 6) of the mitochondrial apocytochrome b gene in Saccharomyces cerevisiae, strain 777-3A . Most of the revertants could not be distinguished from the wild-type by means of physiological properties . Twenty-two revertants different in phenotype are described here in more detail . The suppressor mutations (supa) that compensate the primary cob- mutations (i.e., restore growth on glycerol) are mitochondrially inherited . They were localized in the same cob exon regions as the respective primary mutations, except for one revertant with a primary mutation in exon 6 and a suppressor, 4.2 map units distant, which may be located either in intron 5 or downstream in exon 6 . Of 21 suppressors 17 are closely coupled to the primary mutation with recombination frequencies of less than or equal to 0.1%-0.3% . An estimate predicts that in more than 80% of these revertants only one amino acid is altered at that point of the polypeptide corresponding to the cob- site in the gene . The most interesting revertant phenotypes are: reduced growth rate on glycerol . The respective cob-/supa mutations are scattered over the whole cob region and cannot be correlated exclusively with special gene regions . decreased cytochrome b content . The most extreme reductions (28% and 30% of wild-type level) were observed to be due to mutations located in the 5' proximal part of exon 1 . The highest percentage of revertants with decreased cytochrome b content was predominantly found mapping in exon 3.(ABSTRACT TRUNCATED AT 250 WORDS)

Folia Microbiol (Praha), 1984, 29(4), 343 - 5
Denatured proteins are degraded more rapidly than abnormal proteins in cell-free extracts of Saccharomyces cerevisiae; Chopra AK et al.; Abnormal proteins synthesized in the presence of ethionine were degraded more rapidly than the normal ones in cell-free extracts of ethanol-grown yeast . The denatured proteins, however, were degraded in preference to their native counterparts which were either normal or abnormal.

Braz J Med Biol Res, 1984, 17(1), 17 - 20
Revision of the nucleotide sequence at the last intron of the mitochondrial apocytochrome b gene in Saccharomyces cerevisiae; Bonjardim CA et al.; The nucleotide sequence around a G + C-rich cluster in the last intron of the mitochondrial apocytochrome b gene (F.G . Nobrega and A . Tzagoloff, Journal of Biological Chemistry, 255: 9828-9837, 1980) was revised when restriction digests failed to show a predicted Rsa I site at position +2769 . The corrected sequence has four additional nucleotides, one Hae III and two Hpa II sites . Previously undetected typographical errors were also found in the A + T-rich sequence.

Int J Biochem, 1984, 16(6), 667 - 73
A rapid method for the purification of fatty acid synthetase from the yeast Saccharomyces cerevisiae; Karam GA et al.; A rapid method for the isolation and purification of small quantities of highly active fatty acid synthetase (FAS) from several strains of the yeast Saccharomyces cerevisiae, is presented . The purification procedure which is the shortest reported to this date (18 hr), involves the release of the enzyme by either cell wall digestion with Zymolyase 60000 or cell wall disruption by glass beads, followed by 35-50% ammonium sulfate fractionation, desalting by Sephadex G-25 chromatography, then calcium phosphate gel treatment, concentration by 50% ammonium sulfate precipitation, sedimentation of the enzyme in the ultracentrifuge and finally, column chromatography on DEAE Bio-Gel A . Fatty acid synthetase prepared by the cell breakage method, was found to be homogeneous according to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), SDS-Tris-glycine disc gel electrophoresis and immunoelectrophoresis criteria . However, enzyme prepared from Zymolyase treated cells showed several proteolytic bands in addition to FAS bands, on SDS-PAGE . Enzyme obtained by both methods of cell breakage, showed a similar behavior throughout the purification procedure and gave a similar yield of enzyme of high specific activity (4800-7200 nmol/min/mg) that remained stable for several months at -85 degrees C.

Mol Cell Biochem, 1984, 61(2), 173 - 82
Regulation of galactokinase (GAL1) enzyme accumulation in Saccharomyces cerevisiae; Yarger JG et al.; The regulation of GAL1 RNA and enzyme synthesis has been investigated in Saccharomyces cerevisiae . We have shown that the induction of GAL10 and GAL1 RNAs is coordinate . GAL1 RNA transcripts appear within 4.5 to 6 min and galactokinase synthesis within 6 to 9 min . Steady-state RNA levels were reached within 50 min and the steady-state rate of galactokinase enzyme synthesis within 40-50 min . From these kinetic studies, the initial induction of GAL1 enzyme activity is apparently under transcriptional control . In addition, during early induction, two galactokinase enzyme activities were detected; a major stable form and a minor unstable form.

Mol Gen Genet, 1984, 194(1-2), 299 - 302
Effect of aspirin on mitochondrial mutagens in Saccharomyces cerevisiae; Bruce IJ et al.; The mitochondrial mutation petite was induced in yeast cells by ethidium bromide (EB), Adriamycin (ADR) and 4-nitroquinoline-N-oxide (NQO) . In the presence of aspirin in concentrations ranging from 0.1 to 1.0 mg/ml, the mutagenicity of EB and ADR was reversed but petite induction by NQO was unaffected . At these concentrations, aspirin also reversed mitochondrial inhibition by oligomycin, a non-mutagenic inhibitor of the organellar ATPase complex . Cells grown in the presence of aspirin alone showed a significantly higher rate of oxygen uptake than untreated control cultures when the drug concentration ranged from 0.05 to 1.0 mg/ml . At concentrations of 2 mg/ml and above, aspirin inhibited mitochondrial respiration.

Folia Microbiol (Praha), 1984, 29(2), 115 - 9
Lysis of growing cells of Saccharomyces cerevisiae induced by papulacandin B; Kopecka M; Light and electron microscopy was used to study the effect of papulacandin B on Saccharomyces cerevisiae in the exponential growth phase . At 1-2 micrograms/mL cell division in the culture continued almost in parallel with the control, at 4 micrograms/mL cell proliferation was reduced and the culture contained some cells with 2-9 buds which were not separated from the mother cell by a septum, and at higher concentrations (8, 16 and 32 micrograms/mL) the proliferation stopped within 2 h . Cessation of proliferation was due to lysis of budding cells in the bud region including perforation of thinned cell wall (most often at the bud basis and sometimes at its apex), extrusion of cytoplasm and death of cell . Lysis was also observed in cells without visible buds . Dividing cells died without visible lysis.

DNA, 1984, 3(2), 167 - 71
Homologous in vitro transcription of linear DNA fragments containing the tRNAArg-tRNAAsp gene pair from Saccharomyces cerevisiae; Kjellin-Straby K et al.; Transcription of a tRNAArg-tRNAAsp gene pair from Saccharomyces cerevisiae by an homologous yeast extract results in a dimeric percursor molecule which is processed to mature-sized tRNAArg and tRNAAsp molecules . We have transcribed linear DNA fragments cleaved within the gene sequences to show that precursor synthesis is not dependent on the internal promoter of the second gene (tRNAAsp) . Furthermore, the second gene does not support independent transcription when the normal upstream initiation site is removed.

Folia Biol (Praha), 1984, 30(1), 1 - 14
Evidence for interaction of 5.8S rRNA with the 5'- and 3'- terminal segments of Saccharomyces cerevisiae 25S rRNA; Georgiev OI et al.; The possible sites of 5.8S:25S rRNA interaction in Saccharomyces cerevisiae are investigated by blot-hybridization of in vivo 32P-labelled 5.8S rRNA with restriction fragments from the 25S rRNA gene . Strong hybridization signals are obtained with fragments from the 5'-end (nucleotides 1 to 494) and the 3'-end (3066 to 3391) of the gene . The fragments from the remaining part of the gene are negative . A computer analysis of the known Saccharomyces cerevisiae 5.8S (Rubin 1973) and 25S (Georgiev et al . 1981) rRNA sequences show a markedly higher complementarity between 5.8S rRNA and the 5'- and 3'-terminal segments of 25S rRNA corresponding to the hybridization positive rDNA fragments . In accordance with the experimental and sequence analysis data, two alternative end-to-end base pairing models of possible 5.8S:25S rRNA binding are proposed . Both models imply that 5.8S rRNA connects the 5'- and 3'-terminal segments of 25S rRNA, but differ in the extent of preservation of the secondary structure typical of free 5.8S rRNA . It is suggested that the requirement for 5'- and 3'-end binding of 23S rRNA in Escherichia coli is conserved in evolution and that in eukaryotes 5.8S rRNA plays the role of joining together the two ends of L-rRNA molecules.

Mol Gen Genet, 1984, 193(3), 389 - 94
Genetic study of the role of calcium ions in the cell division cycle of Saccharomyces cerevisiae: a calcium-dependent mutant and its trifluoperazine-dependent pseudorevertants; Ohya Y et al.; A cal1-1 mutant of the yeast Saccharomyces cerevisiae showing Ca2+-dependent growth was isolated . Its growth continued exponentially in Ca2+-rich medium, but stopped in Ca2+-poor medium at 37 degrees C . Mg2+ ions could not replace Ca2+ ions . In Ca2+-poor medium, the mutant cells stopped growing homogeneously at the stage of cell division cycle with a tiny bud . The nucleus in these arrested cells was in the G2 stage, judging from observation after nuclear staining and determination of the DNA content . Trifluoperazine-dependent pseudorevertants, which could grow in the presence of 20 microM to 80 microM trifluoperazine in Ca2+-poor medium at 37 degrees C, were obtained from this cal1-1 mutant . The suppressor mutation, tfr1, itself conferred trifluoperazine resistance . Other calmodulin inhibitors structurally unrelated to trifluoperazine had similar effects to trifluoperazine on these pseudorevertants . These results suggest that Ca2+ ions and a calmodulin play important roles in the yeast cell division cycle at the stage of bud growth and nuclear division.

J Cell Biol, 1984 Jan, 98(1), 341 - 6
Identification of coated vesicles in Saccharomyces cerevisiae; Mueller SC et al.; Clathrin-coated vesicles were found in yeast, Saccharomyces cerevisiae, and enriched from spheroplasts by a rapid procedure utilizing gel filtration on Sephacryl S-1000 . The coated vesicles (62-nm diam) were visualized by negative stain electron microscopy and clathrin triskelions were observed by rotary shadowing . The contour length of a triskelion leg was 490 nm . Coated vesicle fractions contain a prominent band with molecular weight of approximately 185,000 when analyzed by SDS PAGE . The presence of coated vesicles in yeast cells suggests that this organism will be useful for studying the function of clathrin-coated vesicles.

Mol Cell Biol, 1984 Jan, 4(1), 92 - 100
Multiple L double-stranded RNA species of Saccharomyces cerevisiae: evidence for separate encapsidation; Thiele DJ et al.; The L double-stranded (ds) RNA component of Saccharomyces cerevisiae may contain up to three dsRNA species, each with a distinct sequence but with identical molecular weights . These dsRNAs have been separated from each other by denaturation and polyacrylamide gel electrophoresis . The 3' terminal sequences of the major species, LA dsRNA, were determined . Secondary structural analysis supported the presence of two stem and loop structures at the 3' terminus of the LA positive strand . In strain T132B NK-3, both the LA and LC species are virion encapsidated . Two distinct classes of virions were purified from this strain, each with a different RNA polymerase activity and with distinct protein components . The heavy virions harbored LA dsRNA, whereas the LC dsRNA species co purified with the light virion peak . Thus, LA and LC dsRNAs, when present in the same cell, may be separately encapsidated.

Mol Cell Biol, 1984 Jan, 4(1), 181 - 7
Two new double-stranded RNA molecules showing non-mendelian inheritance and heat inducibility in Saccharomyces cerevisiae; Wesolowski M et al.; Certain strains of Saccharomyces cerevisiae were found to have a complex nuclear defect (designated clo-) that makes cells unable to maintain some L-B and some L-C double-stranded RNAs at 25 degrees C . The clo- strains were not defective in maintenance of L-A, M1, or M2 double-stranded RNAs . Most clo-strains lacking L and M carry small amounts of two double-stranded RNA species intermediate in size between L and M and denoted T (2.7 kilobase pairs) and W (2.25 kilobase pairs) . Some strains carry both T and W, some carry neither, and some carry only W; no strains carrying only T have been found . Both T and W show 4+:0 segregation in meiosis and efficient transmission by cytoplasmic mixing (cytoduction), indicating that they are non-Mendelian genetic elements . T and W do not cross-hybridize with each other or with L-A, L-B, L-C, M1, M2, or chromosomal DNA . T and W are apparently distinct from other known non-Mendelian genetic elements (2mu DNA, {rho}, {psi}, 20S RNA, {URE3}) . In most strains the copy number of both T and W is increased about 10-fold by the growth of cells at 37 degrees C . This heat inducibility of T and W is under control of a cytoplasmic gene . T and W double-stranded RNAs are not found in a purified L-containing virus-like particle preparation from a strain containing L-B, T, and W double-stranded RNAs . The role, if any, of T or W in the killer systems is not known.

Mol Cell Biol, 1984 Jan, 4(1), 142 - 50
Isolation of DNA sequences preferentially expressed during sporulation in Saccharomyces cerevisiae; Percival-Smith A et al.; A differential hybridization screen has been used to identify genes cloned from the yeast Saccharomyces cerevisiae that are expressed preferentially during sporulation . Duplicate copies of a partial Sau3A yeast DNA library prepared in the vector pBR322 were hybridized with radioactive cDNA probes representing the mRNA populations of sporulating a alpha cells and asporogenous alpha alpha cells at various times after transfer to sporulation medium . Thirty-eight clones showed an enhanced hybridization signal with the a alpha sporulation probe relative to the alpha alpha control cDNA probe . A comparison of the array of fragments produced by restriction endonuclease digestion of these plasmids suggested that 15 different sequences had been cloned . An RNA blot analysis using these cloned DNAs to probe RNAs purified from aa, a alpha, and alpha alpha cells harvested either during vegetative growth or at 10 h after transfer to sporulation medium indicated that 14 different sporulation-specific genes had been identified . Transcripts complementary to these genes are present only in a alpha cells after transfer to sporulation medium . Three of these clones contain two sporulation-specific genes . Three genes have been identified that are expressed in all cell types during vegetative growth and only in a alpha cells in sporulation medium.

Nauchnye Doki Vyss Shkoly Biol Nauki, 1984, (1), 35 - 7
{Role of DNA damage in the inactivation of Saccharomyces cerevisiae yeasts by 313-nm ultraviolet light}; Pospelov ME et al.; The relative contribution of respiration photoinhibition and DNA damage in the lethal effect induced by 313 nm ultraviolet light (UV) has been investigated in some strains of the yeast Saccharomyces cerevisiae . It has been shown that cells inactivation is essentially due to photo-induced damage to DNA . By photoreactivation experiments it has been found that dimers of the pyrimidine bases are the main lethal photoproducts induced in the DNA by 313 nm ultraviolet light.

Arch Biochem Biophys, 1984 Jan, 228(1), 1 - 12
Reversible inactivation of Saccharomyces cerevisiae glutathione reductase under reducing conditions; Pinto MC et al.; Glutathione reductase from Saccharomyces cerevisiae was rapidly inactivated following aerobic incubation with NADPH, NADH, and several other reductants, in a time- and temperature-dependent process . The inactivation had already reached 50% when the NADPH concentration reached that of the glutathione reductase subunit . The inactivation was very marked at pH values below 5.5 and over 7, while only a slight activity decrease was noticed at pH values between these two values . After elimination of excess NADPH the enzyme remained inactive for at least 4 h . The enzyme was protected against redox inactivation by low concentrations of GSSG, ferricyanide, GSH, or dithiothreitol, and high concentrations of NAD(P)+; oxidized glutathione effectively protected the enzyme at concentrations even lower than GSH . The inactive enzyme was efficiently reactivated after incubation with GSSG, ferricyanide, GSH, or dithiothreitol, whether NADPH was present or not . The reactivation with GSH was rapid even at 0 degree C, whereas the optimum temperature for reactivation with GSSG was 30 degrees C . A tentative model for the redox interconversion, involving an erroneous intramolecular disulfide bridge, is put forward.

Proc Natl Acad Sci U S A, 1984 Jan, 81(1), 120 - 4
Identification of a labile protein involved in the G1-to-S transition in Saccharomyces cerevisiae; Popolo L et al.; The biochemical nature of the start process that commits budding yeast to DNA synthesis is not known . Kinetic evidence has suggested recently that short-lived protein(s) may have to accumulate to a critical level before the cell cycle may progress towards DNA synthesis and cell division . We investigated by high-resolution two-dimensional electrophoresis whether, in a cdc25-1 mutant strain of Saccharomyces cerevisiae that had been blocked at the regulatory step called "start" by growth at a restrictive temperature, short-lived proteins are synthesized during the recovery of growth at a permissive temperature . Of the approximately equal to 500 proteins resolved by the two-dimensional electrophoresis, 6 were short-lived . Only one of them (Mr = 100,000, pI approximately equal to 4.8-5) appears to be specifically made during the G1-to-S transition at start . A regulatory role for cell cycle progression in yeast is suggested for this protein, p100.

Mol Gen Genet, 1984, 193(1), 192 - 4
Cell cycle parameters in radiation sensitive strains of Saccharomyces cerevisiae; Fingerhut R et al.; Cell cycle parameters in different radiation-sensitive strains of diploid yeast were determined by flow cytofluorometry . The cell generation time was increased in homozygous rad2 and rad51 mutants but was not significantly different from the wild type in rad9 and rad6 mutants . All mutants had a longer G1-phase than the wild type . A lengthened S-phase was found in rad2 cells . Rad51 mutants displayed a considerably longer duration of G2.

Mol Gen Genet, 1984, 193(1), 188 - 9
Benomyl prevents nuclear fusion in Saccharomyces cerevisiae; Delgado MA et al.; Benomyl prevents nuclear fusion in mating mixtures of Saccharomyces cerevisiae . Cytoductants, heterokaryons and diploids may be recovered from these mixtures.

J Bacteriol, 1984 Jan, 157(1), 283 - 90
Copy number and the stability of 2-micron circle-based artificial plasmids of Saccharomyces cerevisiae; Futcher AB et al.; The copy number and stability of artificial 2-micron circle-based plasmids have been accurately measured in {Cir+} and {Cir0} strains of Saccharomyces cerevisiae . We conclude that (i) instability and copy number vary greatly from plasmid to plasmid; (ii) instability and copy number are negatively correlated--that is, high copy number is associated with low instability; (iii) it is difficult to reconcile this variability with a strict and direct system of copy number control; (iv) instabilities are much higher than expected from random partition and the observed copy numbers: this may imply partition which is less efficient than random . Even so, (v) the partitioning of 2-micron circle-like plasmids is more efficient than that of ARS-based plasmids, which hints at the existence of a system for the (inefficient) distribution of 2-micron circles.

J Bacteriol, 1984 Jan, 157(1), 277 - 82
Identification of the structural gene and nonsense alleles for adenylate cyclase in Saccharomyces cerevisiae; Matsumoto K et al.; Tetraploid strains of Saccharomyces cerevisiae carrying different dosages of the CYR1+ gene have been constructed . Adenylate cyclase activity observed in these tetraploid strains was proportional to the dosage of the active CYR1+ gene . Of the 57 mutants requiring adenosine 3',5'-monophosphate for growth at 35 degrees C, two allelic temperature-sensitive cyr1 mutants produced thermolabile adenylate cyclase . Crude extract and plasma membrane fraction of cyr1 mutant cells had no adenylate cyclase activity when assayed with GTP or 5'-guanylyl imidodiphosphate in the presence of Mn2+ or Mg2+ . Plasma membrane and Lubrol-soluble plasma membrane fractions obtained from the temperature-sensitive cyr1 mutant were thermolabile compared with those from the wild-type strain . Three cyr1 mutants carried nonsense mutations susceptible to ochre (UAA) suppressors, SUP3 and SUP-o, and had no detectable level of adenylate cyclase activity . It is concluded that the cyr1 mutants carry lesions in the structural gene for adenylate cyclase.

Mol Gen Genet, 1984, 194(3), 395 - 401
Ty1-promoted expression of aspartate transcarbamylase in the yeast Saccharomyces cerevisiae; Bach ML; An entire copy of a Ty1 yeast transposon has been found inserted between two regions comprising the single transcriptional and translational URA2 units in yeast that code respectively for carbamylphosphate synthetase (CPSase) and aspartate transcarbamylase (ATCase) . The mutant Rev 16 was obtained from an ATCase- strain blocked by multiple nonsense mutations in the proximal CPSase region and submitted to selective pressure for recovery of the enzyme activity coded by the distal part of the gene . The inserted Ty1 has one XhoI site in both delta elements, delimiting a 5.6 kb piece of DNA that shows a classical Ty1 restriction pattern . The orientation of this sequence in URA2 is the same as in the previously described examples in which Ty1 has positive effects on the expression of adjacent genes . In this case the Ty1 is situated more than 1 kb from the URA2 region in which ATCase structural mutants have been mapped . Nevertheless, transcription of the entire sequence distal to the Ty1 is restored and has become subject to mating-type control, leading to a weak enzyme activity . Our observations are in agreement with generally accepted ideas regarding the way in which Ty1 elements affect gene expression, and additionally, represent the first example of a Ty1 -promoted reinitiation occurring in the middle of a single transcription unit.

Mol Gen Genet, 1984, 194(1-2), 144 - 8
Cloning and restriction analysis of the hexokinase PII gene of the yeast Saccharomyces cerevisiae; Frohlich KU et al.; Carbon catabolite repression in yeast depends on catalytic active hexokinase isoenzyme PII ( Entian 1980a ) . A yeast strain lacking hexokinase isoenzymes PI and PII was transformed, using a recombinant pool with inserts of yeast nuclear DNA up to 10 kbp in length . One hundred transformants for hexokinase were obtained . All selected plasmids coded for hexokinase isoenzyme PII, none for hexokinase isoenzyme PI, and carbon catabolite repression was restored in the transformants . Thirty-five independently isolated stable plasmids were investigated further . Analysis with the restriction enzyme EcoRI showed that these plasmids fell into two classes with different restriction behaviour . One representative of each class was amplified in Escherichia coli and transferred back into the yeast hexokinase-deficient strain with concomitant complementation of the nuclear mutation . The two types of insert were analysed in detail with 16 restriction enzymes, having 0-3 cleavage sites on transformant vector YRp7 . The plasmids differed from each other by the orientation of the yeast insert in the vector . After yeast transformation with fragments of one plasmid the hexokinase PII gene was localised within a region of 1.65 kbp.

Xenobiotica, 1984 Jan-Feb, 14(1-2), 187 - 206
Studies on the properties of highly purified cytochrome P-448 and its dependent activity benzo{a}pyrene hydroxylase, from Saccharomyces cerevisiae; King DJ et al.; The yeast Saccharomyces cerevisiae, produces a cytochrome P-450 enzyme with a Soret peak in the reduced-CO difference spectrum at 448 nm . The enzyme purified to homogeneity (88-97% pure on a specific content basis) has a molecular wt . of 55 500 as determined by SDS-PAGE . Amino acid analysis of yeast cytochrome P-448 revealed 407 amino acid residues per molecule with a 43% complement of hydrophobic residues . Although the number of residues is smaller than cytochrome P-448 enzymes from mammalian sources, the percentage of hydrophobic residues is almost identical . Estimation of the haem content of yeast cytochrome P-448 showed that one haem group was present per molecule . Phospholipid was present at very low levels . The molecular wt . of the polypeptide chain plus an estimated 5-6 units of hexose and of hexosamine is in good agreement with the molecular wt . value obtained from SDS-PAGE . A reconstituted system of purified cytochrome P-448, purified NADPH-cytochrome P-450 (c) reductase and phospholipid showed aryl hydrocarbon hydroxylase activity towards benzo{a}pyrene . Both protein components, NADPH and dilauroyl phosphatidylcholine (or emulgen 911) were necessary for full activity . The NADPH requirement could be replaced by cumene hydroperoxide or H2O2 generated in situ from a glucose oxidase system; in each case Vmax is increased, but the apparent affinity for benzo{a}pyrene, as measured by an increased Km, is lowered . The spin state of purified yeast cytochrome P-448 was 94% low spin (22 degrees C) as determined from the temperature-dependent spin-state equilibrium . The addition of benzo{a}pyrene to this enzyme resulted in a change to higher spin state (18% high spin at 22 degrees C) . Equilibrium gel filtration analysis of the number of benzo{a}pyrene binding sites per mole of enzyme monomer showed a value of 1 for purified yeast cytochrome P-448 and 6 for this enzyme in microsomal form . The corresponding values for purified and microsomal cytochrome P-450 from phenobarbital-pretreated rats are 1 and 6, respectively . However, purified cytochrome P-448 from beta-naphthoflavone-induced rats gave a value of 6 benzo{a}pyrene binding sites . Type I binding spectra with purified yeast cytochrome P-448 were observed with benzo{a}pyrene, lanosterol, ethylmorphine, dimethylnitrosamine, sodium phenobarbitone and perhydrofluorene . Type II spectral changes were observed with imidazole, aniline and benzphetamine . Cytochrome P-448 from Saccharomyces cerevisiae is identified as a distinct enzyme of the P-450 family . This enzyme however has many properties in common with cytochrome P-448 from mammalian sources.(ABSTRACT TRUNCATED AT 400 WORDS)

Gene, 1984 Jan, 27(1), 35 - 46
Expression of calf prochymosin in Saccharomyces cerevisiae; Goff CG et al.; A yeast strain which synthesizes activatable calf prochymosin (also known as prorennin) has been constructed by transformation with a vector carrying the methionyl-prochymosin coding sequence attached to efficient yeast transcriptional promoter and terminator sequences . Cloned preprochymosin cDNA was altered by restriction endonuclease cleavage and addition of a synthetic oligonucleotide to yield a DNA sequence encoding methionyl-prochymosin . This methionyl-prochymosin gene was ligated to a yeast chromosomal fragment containing the GAL1 promoter, and the construction was placed in an Escherichia coli-Saccharomyces cerevisiae shuttle vector with or without a transcriptional terminator DNA fragment from the yeast SUC2 gene . In yeast the two constructions result in equal amounts of prochymosin protein and mRNA . The prochymosin from yeast is activatable to chymosin by incubation at low pH and exhibits milk-clotting activity indistinguishable from calf chymosin.

Mol Gen Genet, 1984, 193(3), 525 - 31
Cloning and mapping of the RAD50 gene of Saccharomyces cerevisiae; Kupiec M et al.; The RAD50 gene was cloned as a 4.8 kb fragment in the 2 mu derived plasmid pFL1 . The gene resides in a 3.9 kb segment that was subcloned into the plasmid YRp7 . The cloned gene complements the deficiency caused by the rad50-1 mutation with respect to gamma-rays, MMS resistance and UV-induced mitotic recombination . Restoration of the Rad+ phenotype occurs when the cloned gene is on a freely replicating multiple-copy plasmid or in the integrated form . Mapping of the cloned gene following integration of the 2 mu plasmid, and of the subclone in plasmid YRp7, showed it to be located on the left arm of chromosome XIV . Tetrad analysis of various crosses involving two different strains carrying rad50-1 showed the mutation to map next to pet2 on chromosome XIV, and not on the right arm of chromosome IV, as previously published.

Mol Gen Genet, 1984, 193(3), 507 - 12
Isolation and characterization of a pleiotropic glucose repression resistant mutant of Saccharomyces cerevisiae; Bailey RB et al.; A new mutation has been described which confers resistance to catabolite repression in Saccharomyces cerevisiae . The mutant allele, termed grr-1 for glucose repression-resistant, is characterized by insensitivity to glucose repression for the cytoplasmic enzymes invertase, maltase, and galactokinase, as well as the mitochondrial enzyme cytochrome c oxidase . Hexokinase levels in grr-1 mutants are approximately 3-fold higher than the corresponding activity of the parental strain . Although the grr-1 allele is expressed phenotypically similarly to the hex-1 (hxk-2) and hex-2 mutations described by Entian et al . (1977) and Zimmermann and Scheel (1977) respectively, we have shown genetically and physiologically that grr-1 represents a new class of mutation.

Mol Gen Genet, 1984, 193(3), 406 - 13
A DNA sequence from Dictyostelium discoideum complements ura3 and ura5 mutations of Saccharomyces cerevisiae; Boy-Marcotte E et al.; A 3.7 kilobase fragment of Dictyostelium discoideum genomic DNA has been cloned by its ability to complement a yeast ura3 mutation affecting the activity of orotidine-5'-phosphate carboxy-lyase (EC 4.1.1.23) . This fragment also complements a yeast ura5 mutation that leads to a defect in orotate phosphoribosyl transferase (EC 2.4.2.10) . The orotidine-5'-phosphate carboxy-lyase and the orotate phosphoribosyl transferase activities that result from Dictyostelium gene expression in yeast have been detected . The size of the DNA required for both complementations has been localised to a segment of less than 2 kb . A unique Dictyostelium RNA species of 1,600 base pairs hybridizes to this fragment . In vitro deletions in this fragment lead to the simultaneous loss of the two activities . The two enzymatic activities coelute as a protein of 120,000 daltons during gel filtration of a Dictyostelium extract . These results favour the existence, on the cloned Dictyostelium DNA fragment, of a unique structural gene which codes for a bifunctional enzyme carrying the two activities, orotidine-5'-phosphate carboxy-lyase and orotate phosphoribosyl transferase.

EMBO J, 1984 Jan, 3(1), 207 - 12
Amino acid sequence of a new mitochondrially synthesized proteolipid of the ATP synthase of Saccharomyces cerevisiae; Velours J et al.; The purification and the amino acid sequence of a proteolipid translated on ribosomes in yeast mitochondria is reported . This protein, which is a subunit of the ATP synthase, was purified by extraction with chloroform/methanol (2/1) and subsequent chromatography on phosphocellulose and reverse phase h.p.l.c . A mol . wt . of 5500 was estimated by chromatography on Bio-Gel P-30 in 80% formic acid . The complete amino acid sequence of this protein was determined by automated solid phase Edman degradation of the whole protein and of fragments obtained after cleavage with cyanogen bromide . The sequence analysis indicates a length of 48 amino acid residues . The calculated mol . wt . of 5870 corresponds to the value found by gel chromatography . This polypeptide contains three basic residues and no negatively charged side chain . The three basic residues are clustered at the C terminus . The primary structure of this protein is in full agreement with the predicted amino acid sequence of the putative polypeptide encoded by the mitochondrial aap1 gene recently discovered in Saccharomyces cerevisiae . Moreover, this protein shows 50% homology with the amino acid sequence of a putative polypeptide encoded by an unidentified reading frame also discovered near the mitochondrial ATPase subunit 6 gene in Aspergillus nidulans.

Mol Cell Biol, 1984 Jan, 4(1), 61 - 8
Carbon source dependence of transposable element-associated gene activation in Saccharomyces cerevisiae; Taguchi AK et al.; Seven cis-dominant mutations leading to the overproduction of the glucose-repressible alcohol dehydrogenase isozyme ADHII (structural gene, ADH2) in Saccharomyces cerevisiae have previously been shown to be due to insertion of a transposable element, Ty, in the 5' regulatory region of the ADH2 gene . We showed that although mating-competent cells (a, alpha, a/a, or alpha/alpha cells) overproduced both ADHII enzyme and ADH2 mRNA, mating-incompetent cells (a/alpha or ste-cells) produced much less ADHII enzyme and ADH2 mRNA . This mating type effect on ADH2 expression was greatest in the presence of a normally derepressing carbon source, glycerol, and much less apparent in the presence of a repressing carbon source, glucose . In addition, Ty insertion led to an aberrant carbon source response in mating-incompetent cells--the normally glucose-repressible ADHII becomes glycerol repressible . The mating type effect and aberrant carbon source response in mating-incompetent cells was specific for Ty-associated mutations in the 5' flanking region of the ADH2 gene in that a non-Ty mutation in the same region did not show these effects . Finally, Ty1 RNA levels also showed a/alpha, suppression, which was apparent only during growth on a nonfermentable carbon source such as glycerol . This suggests that Ty-mediated gene expression is subject to regulation by both mating competence and carbon catabolites.

Mol Cell Biol, 1984 Jan, 4(1), 203 - 11
Mating type control in Saccharomyces cerevisiae: a frameshift mutation at the common DNA sequence, X, of the HML alpha locus; Tanaka K et al.; A mutation defective in the homothallic switching of mating type alleles, designated hml alpha-2, has previously been characterized . The mutation occurred in a cell having the HO MATa HML alpha HMRa genotype, and the mutant culture consisted of ca . 10% a mating type cells, 90% nonmater cells of haploid cell size, and 0.1% sporogenous diploid cells . Genetic analyses revealed that nonmater haploid cells have a defect in the alpha 2 cistron at the MAT locus . This defect was probably caused by transposition of a cassette originating from the hml alpha-2 allele by the process of the homothallic mating type switch . That the MAT locus of the nonmater cells is occupied by a DNA fragment indistinguishable from the Y alpha sequence in electrophoretic mobility was demonstrated by Southern hybridization of the EcoRI-HindIII fragment encoding the MAT locus with a cloned HML alpha gene as the probe . The hml alpha-2 mutation was revealed to be a one-base-pair deletion at the ninth base pair in the X region from the X and Y boundary of the HML locus . This mutation gave rise to a shift in the open reading frame of the alpha 2 cistron . A molecular mechanism for the mating type switch associated with the occurrence of sporogenous diploid cells in the mutant culture is discussed.

Mol Gen Genet, 1984, 193(1), 149 - 52
Genetics of oxidative phosphorylation: petite deletion mapping of the Oli 2 region of the mitochondrial genome of Saccharomyces cerevisiae; Connerton IF et al.; Petite deletion mapping has been carried out for the Oli 2 region of the mitochondrial genome of Saccharomyces cerevisiae to produce a fine structure genetic map . Previously unlocated mit- mutants together with the drug resistant loci Oli 2 and Oss 1 have been ordered between the cytochrome oxidase and apocytochrome b genes . As a result of this study a series of isogenic p- clones have been isolated spanning the Oli 2 region.

Arch Biochem Biophys, 1984 Jan, 228(1), 22 - 30
Involvement of oxygen and mitochondrial function in the metabolism of D-xylulose by Saccharomyces cerevisiae; Maleszka R et al.; Mitochondrial function associated with oxygen was required for growth of Saccharomyces cerevisiae on D-xylulose . The requirement was shown by (i) the inhibition of growth of a wild-type strain under anaerobic conditions, (ii) the inhibition of aerobic growth after treatment with inhibitors of mitochondrial function, and (iii) the lack of aerobic and anaerobic growth of nuclear and cytoplasmic petites . The mitochondrial function was associated with the channeling of catabolites of D-xylulose to growth processes, since ethanol was formed even when growth was inhibited . Mitochondrial function was implicated as well in determining the extent of growth and the concentration of ethanol in aerobic cultures of the wild-type . In such cultures, the concentration of ethanol decreased and growth increased concomitantly as aeration rate increased . A factor in this relation was considered to be the relatively poor ability of D-xylulose to inhibit the oxidative utilization of ethanol.

Acta Microbiol Pol, 1984, 33(1), 25 - 35
Protoplast fusion in Saccharomyces cerevisiae; Skala J et al.; The ability of different Saccharomyces cerevisiae yeast strains to form protoplasts and protoplast fusion were studied . The protoplast formation depended mainly on strains used and the time of snail gut enzyme action . The percentages of the regenerating protoplasts varied, depending on strain, from 3 to 33 per cent . From the fusion experiments one can establish that kariogamy is prerequisite for stable for stable diploid formation . The yields of protoplast fusion were higher when both strains were rho+ as compared with rho+ and rho 0 combinations.

Mol Cell Biol, 1984 Jan, 4(1), 101 - 9
Saccharomyces cerevisiae killer virus transcripts contain template-coded polyadenylate tracts; Hannig EM et al.; The M double-stranded RNA component of type 1 killer strains of the yeast Saccharomyces cerevisiae contains an internal 200-base pair adenine- and uracil-rich region . The plus strands of this viral genomic RNA contain an internal adenine-rich region which allows these strands to bind to polyuridylate-Sepharose as tightly as do polyadenylated RNAs with 3'-terminal polyadenylated tracts of 70 to 100 residues . Internal template coding of an adenine-rich tract in positive polarity in vivo and in vitro transcripts of M double-stranded RNA may serve as an alternate method of transcript polyadenylation . The 3'-terminal residue of the in vitro m transcript is a non-template-encoded purine residue . The 5' terminus of this transcript is involved in a stem-and-loop structure which includes an AUG initiation codon, along with potential 18S and 5.8S rRNA binding sites . Except for the 3'-terminal residue, transcription in in vitro shows complete fidelity.

Mutat Res, 1984 Jan, 139(1), 21 - 4
Genetic effects of 5-azacytidine in Saccharomyces cerevisiae; Zimmermann FK et al.; The base analog 5-azacytidine induced a variety of genetic and epigenetic effects in different organisms . It was tested in two diploid strains of the yeast Saccharomyces cerevisiae to study the induction of point mutation, mitotic reciprocal crossing-over, mitotic gene conversion (strain D7) and mitotic aneuploidy (strain D61.M) . It was used on cells growing in its presence for 4-5 generations . There was a strong induction of both types of mitotic recombination and point mutation . However, there was no induction of mitotic chromosomal malsegregation under the same conditions.

Teratog Carcinog Mutagen, 1984, 4(2), 201 - 10
Genetic effects of procarbazine in the yeast Saccharomyces cerevisiae, strain D4; Frezza D et al.; Procarbazine ( PCZ ) was tested for its ability to induce mitotic gene conversions at the ade and trp loci of Saccharomyces cerevisiae, strain D4 . The influence of the following factors was examined: growth phase of the yeast cells (log vs stationary phase), pH of the treatment solution, and addition of mouse S9 fractions prepared from different organs . The drug was found more toxic and mutagenic at low doses (up to 25 mg/ml) for log phase cells, and scarcely toxic but highly mutagenic, even at high doses, for stationary phase cells . PCZ activity was reduced by acidic pH, and suppressed by S9 mix . Gene conversions were also analyzed in the intrasanguineous host-mediated assay performed in mice orally administered with PCZ . In such conditions PCZ was ineffective in stimulating mitotic gene conversions, probably owing to its inactivation in the acidic environment of the gastroenteric tract.

Acta Biochim Pol, 1984, 31(4), 375 - 82
Phosphorylation of ribosomal proteins during differentiation of Saccharomyces cerevisiae; Szyszka R et al.; Four major phosphoproteins of yeast ribosomes: S2, S6, L44 and L45, were identified in germinating spores . It was found that protein S6 became phosphorylated at an early stage of germination and that only the phosphorylation of this protein responded well to changes in growth conditions of yeast culture . The results obtained give support for the suggested functional role of modification of S6 protein.

Microbios, 1984, 41(165-166), 177 - 89
Effects of inhibitors of plasma-membrane ATPase on potassium and calcium fluxes, membrane potential and proton motive force in the yeast Saccharomyces cerevisiae; Eilam Y et al.; The plasma membrane ATPase inhibitors N,N-dicyclohexylcarbodiimide (DCCD), diethylstilboestrol and sodium orthovanadate caused inhibition of proton ejection in Saccharomyces cerevisiae at a concentration range of 0.1-0.4 mM . At this concentration K+ efflux was not observed; DCCD caused K+ efflux only at a much higher concentration (2mM) . It was shown that induction of K+ efflux by DCCD was not mediated via inhibition of ATPase and that it may be the result of a direct effect of the drug on the cell membrane or on the K+ carrier . All three inhibitors also caused mild hyperpolarization of the membrane and an increase in the carrier mediated Ca2+ uptake into the cells . The increase in delta psi balanced the decrease in delta pH so that the value of delta mu H+ changed very little following incubation of the cells with the inhibitors . The respiratory deficient mutant ('petite') displayed similar phenomena to the wild type, but membrane potential values were lower than in the wild type.

Mol Gen Genet, 1984, 195(1-2), 275 - 80
Molecular cloning and genetic mapping of the PET494 gene of Saccharomyces cerevisiae; Muller PP et al.; The activity of the nuclear gene PET494 is required to allow expression of the yeast mitochondrial gene oxi2 . To aid the study of the mechanism of action of PET494 we have isolated this gene from yeast DNA . A clone bank of yeast DNA fragments in a yeast-E . coli shuttle vector was screened by transformation for a plasmid able to complement the pet494-1 amber mutation . A complementing plasmid was obtained that contained a unique 4.4 kb yeast sequence . This 4.4 kb sequence contains the PET494 gene . Integration of a plasmid containing it into chromosomal DNA by homologous recombination, and subsequent genetic analysis, demonstrated that the 4.4 kb fragment was tightly linked to the pet494-1 mutation . In addition, the corresponding 4.4 kb sequence isolated from a pet494-1 mutant failed to complement the mutation . A 2 kb fragment, subcloned from the original plasmid retained the ability to complement the mutation . The pet494-1 mutation maps to chromosome XIV between rna2 and lys9, approximately 2.4 cm from lys9.

Mol Gen Genet, 1984, 195(3), 500 - 6
Molecular cloning and biosynthetic regulation of cry1 gene of Saccharomyces cerevisiae; Himmelfarb HJ et al.; The cryptopleurine resistance gene, cry1, of Saccharomyces cerevisiae has been molecularly cloned using genetic complementation of cryptopleurine sensitivity by the cryptopleurine resistance gene contained in a clone library prepared from DNA of a cryptopleurine resistant strain . Analysis of RNA transcripts indicated that the cry1 gene is the template for a transcript of approximately 900 bases and that the primary transcript contains an intron of approximately 300 bases . In vitro hybrid selection translation experiments indicated that this transcript encodes a protein of molecular weight 17 kilodaltons which on two-dimensional SDS polyacrylamide gels exactly coincides with ribosomal protein rp59 . Further analysis showed that when the gene was present on a plasmid of about five copies per cell the amount of messenger RNA was elevated approximately five-fold compared to a cell that had only a single chromosomal copy . The rate of synthesis of ribosomal protein rp59 was not detectably elevated . These data suggest that the cry1 gene is regulated, at least in part, post-transcriptionally.

Eur J Biochem, 1983 Dec 15, 137(3), 501 - 7
Purification and properties of a double-stranded ribonuclease from the yeast Saccharomyces cerevisiae; Mead DJ et al.; A double-stranded ribonuclease has been purified more than 90-fold to near homogeneity from the yeast, Saccharomyces cerevisiae . The enzyme shows a high specificity for double-stranded RNA as its substrate . It has a molecular weight of 27000 as determined by sodium dodecyl sulphate/polyacrylamide gel electrophoresis . The enzyme degrades dsRNA optimally at 30 degrees C; it is stimulated by KCl and by the -SH reagent, dithiothreitol . In contrast to RNase III from Escherichia coli, the yeast enzyme is inhibited by divalent cations . Physiological studies have demonstrated that in vivo levels of the enzyme activity fell during the latter part of the exponential growth phase but rose during stationary phase . The specific activity of the enzyme in nitrogen-starved yeast cells was 2-3-fold higher than in non-starved cells . The enzyme could be detected in yeast strains containing both, one or none of the species of cytoplasmic dsRNA (L and MdsRNAs) and may, therefore, have some wider role.

Nature, 1983 Dec 15-21, 306(5944), 707 - 9
ras-Related gene sequences identified and isolated from Saccharomyces cerevisiae; DeFeo-Jones D et al.; The oncogenes of Harvey and Kirsten murine sarcoma viruses (v-rasH and v-rasK) and their cellular homologues (c-rasH and c-rasK) constitute two members of the ras gene family . Each functional member of the ras gene family encodes a 21,000 molecular weight protein (p21ras) . ras genes have been detected in a wide variety of vertebrate species, including Xenopus laevis (R . E . Steele, personal communication), and in Drosophila melanogaster . We report here the detection of ras-related genes in the yeast Saccharomyces cerevisiae, and the isolation of two ras-related molecular clones, c-rassc-1 and c-rassc-2, from the DNA of Saccharomyces . Both c-rassc-1 and c-rassc-2 hybridize specifically to probes prepared from mammalian ras DNA . Sequencing of c-rassc-1 reveals extensive amino acid homology between the protein encoded by c-rassc-1 and the p21 encoded by c-rasH . Our studies suggest that these clones can be used to elucidate the normal cellular functions of ras-related genes in this relatively simple eukaryotic organism.

Biochem J, 1983 Dec 1, 215(3), 471 - 4
Post-secretional modification of exo-1,3-beta-D-glucanases from Saccharomyces cerevisiae; Sanchez A et al.; Exo-1,3-beta-D-glucanase secreted by Saccharomyces cerevisiae undergoes extracellular modifications in its carbohydrate moiety that change the affinity towards the lectin concanavalin A . The transition of negatively reacting enzyme form into positively reacting one depends on temperature . Results from experiments with glucono-delta-lactone and from treatments in vitro with hydrolases suggest a glycosidase-mediated mechanism.

Proc Natl Acad Sci U S A, 1983 Dec, 80(24), 7566 - 70
Enhancement of spontaneous mitotic recombination by the meiotic mutant spo11-1 in Saccharomyces cerevisiae; Bruschi CV et al.; Both nonreciprocal and reciprocal mitotic recombination are enhanced by the recessive mutant spo11-1, which was previously shown to affect meiosis by decreasing recombination and increasing nondisjunction . The mitotic effects are not distributed equally in all chromosomal regions . The genotypes of mitotic recombinants in spo11-1/spo11-1 diploid cells provide further evidence that widely spaced chromosomal markers undergo coincident conversion in mitosis.

Gene, 1983 Dec, 26(2-3), 119 - 26
Molecular cloning and characterization of the RAD1 gene of Saccharomyces cerevisiae; Higgins DR et al.; We have cloned the RAD1 gene of Saccharomyces cerevisiae and physically mapped it to a 4.0-kb DNA fragment from chromosome XVI . The RAD1 gene determines a transcript of 3.1 kb, and the direction of transcription was found to be leftwards, from EcoRI towards Bg/II (Fig . 1) . Deletions of the RAD1 gene were made and were found to have no effect on viability of vegetative cells or spores, or on sporulation.

Anal Biochem, 1983 Dec, 135(2), 447 - 52
Detection of phospholipid biosynthetic enzyme activities in Saccharomyces cerevisiae by colony autoradiography; Homann MJ et al.; The colony autoradiography method (C . R . H . Raetz (1975) Proc . Nat . Acad . Sci . USA 72, 2274-2278) was modified for the detection of CDP-diacylglycerol synthase, phosphatidylinositol synthase, phosphatidylserine synthase, and phosphatidylglycerophosphate synthase activities of Saccharomyces cerevisiae colonies on filter paper replica prints . Colonies were replica printed onto filter paper, permeabilized by air drying, and assayed for enzyme activities with labeled substrates . Autoradiograms of replica prints, following enzyme assays, showed dark halos indicating the enzymatic synthesis of labeled phospholipid products . The method was also used to detect a cho 1 mutant defective in phosphatidylserine synthase and a strain that overproduces phosphatidylserine synthase . The method should become a valuable tool in isolating yeast strains defective in phospholipid biosynthetic enzyme activities and strains with overproduced enzyme activities.

Radiat Res, 1983 Dec, 96(3), 532 - 48
Wavelength dependence of inactivation and membrane damage to Saccharomyces cerevisiae cells by monochromatic synchrotron vacuum-uv radiation (145-190 nm); Ito T et al.; Using an electron storage ring as a source of radiation, the wavelength dependence of inactivation and membrane damage in yeast cells (Saccharomyces cerevisiae) was investigated in the range from 145 to 254 nm, with special reference to the effects of vacuum-uv radiation . The cells were irradiated on a Millipore filter in a moist chamber filled with water vapor (deoxygenated) at saturation pressure . Fluence-survival curves taken at 5-nm intervals were generally sigmoidal . Action spectra of the two types of effects were nearly identical in shape . The maximum occurred in both spectra at 160 nm, decreasing sharply toward 180 nm . The spectra remarkably resembled the calculated absorption spectrum of (liquid) water in the range from 145 to 170 nm; the spectra had no similarity at all to the absorption spectra of DNA, proteins, or lipids . These data support the theory that inactivation of wet cells by vacuum-uv radiation may be attributable to damage in the cell membrane initiated by the absorption of water molecules . Above 210 nm the spectrum for inactivation paralleled the absorption of DNA . Genetic changes (induction of gene conversion) were also observed above 210 nm . Photoreversion for the induced convertants was detectable only above 220 nm . These characteristics are consistent with the expectation that above 210 nm the site of major lethal damage shifts to DNA.

Mutat Res, 1983 Dec, 122(3-4), 305 - 8
Effect of post-irradiation inhibition of protein synthesis on UV-induced mitotic gene conversion in Saccharomyces cerevisiae; Vashishat RK et al.; The effect of post-irradiation inhibition of protein synthesis with cycloheximide was studied on UV-induced mitotic gene conversion in yeast . The frequency of UV-induced mitotic gene convertants as well as survival were reduced when post-irradiation protein synthesis was inhibited beyond 8 h . It is concluded that proteins required for mitotic recombination are not induced by UV irradiation and are already present in mitotic cells.

Cell, 1983 Dec, 35(3 Pt 2), 805 - 13
Genetic recombination of homologous plasmids catalyzed by cell-free extracts of Saccharomyces cerevisiae; Symington LS et al.; We have developed an in vitro system utilizing yeast cell-free extracts to catalyze recombination events between homologous plasmids containing different mutant alleles of the Tet or ARG4 genes . The reaction increased the frequency of Tcr or Arg+ transformants (recombinants) from 2 X 10(-6) to 1-3 X 10(-3) . Linearizing one substrate between the two tet mutations stimulated the reaction 2 to 4 fold . The reaction required rATP, Mg++, NAD, and DTT . The rad52-1 mutation decreased the reaction between linear and circular substrates 5 to 6 fold but had little effect with circular substrates . The structures of Tcr plasmids was analyzed by restriction endonuclease mapping and was consistent with a recombination reaction involving crossing-over and gene conversion . Recombination products were also observed directly by subjecting reaction mixtures to electrophoretic analysis . These results indicate that recombination events were catalyzed by the yeast extract.

J Bacteriol, 1983 Dec, 156(3), 1282 - 91
Structural heterogeneity in populations of the budding yeast Saccharomyces cerevisiae; Vanoni M et al.; Bud scar analysis integrated with mathematical analysis of DNA and protein distributions obtained by flow microfluorometry have been used to analyze the cell cycle of the budding yeast Saccharomyces cerevisiae . In populations of this yeast growing exponentially in batch at 30 degrees C on different carbon and nitrogen sources with duplication times between 75 and 314 min, the budded period is always shorter (approximately 5 to 10 min) than the sum of the S + G2 + M + G1* phases (determined by the Fried analysis of DNA distributions), and parent cells always show a prereplicative unbudded period . The analysis of protein distributions obtained by flow microfluorometry indicates that the protein level per cell required for bud emergence increases at each new generation of parent cells, as observed previously for cell volume . A wide heterogeneity of cell populations derives from this pattern of budding, since older (and less frequent) parent cells have shorter generation times and produce larger (and with shorter cycle times) daughter cells . A possible molecular mechanism for the observed increase with genealogical age of the critical protein level required for bud emergence is discussed.

Gene, 1983 Dec, 26(2-3), 223 - 30
The mitochondrial genome of Saccharomyces cerevisiae contains numerous, densely spaced autonomously replicating sequences; Hyman BC et al.; Restriction fragments produced by a complete Sau3A cleavage of Saccharomyces cerevisiae grande mitochondrial DNA were ligated into the yeast-Escherichia coli shuttle vector YIp5 to establish a clone library representing the mitochondrial genome . 30 hybrid plasmids with an average insert size of 1200 bp were chosen at random and tested for the presence of an autonomously replicating sequence (ars) . Over two-thirds of these plasmids transformed yeast at high frequency, indicating the mitochondrial genome contains a large number of ars elements . Our calculations suggest there may be over 40 ars elements contained within the mitochondrial DNA with an average spacing of less than 1700 bp . Mapping experiments indicate that ars elements can be found at many locations on the mitochondrial genome, and in the initial example we have tested, the locations of ars elements derived from grande and petite mtDNAs appear to coincide . If we assume that these ars elements represent mitochondrial DNA replication origins used in vivo, these observations would explain in part the fact that petite mtDNAs can be derived from any location on the grande mitochondrial genome.

Eur J Biochem, 1983 Nov 2, 136(2), 275 - 81
Purification and characterization of two ribosomal proteins of Saccharomyces cerevisiae . Homologies with proteins from eukaryotic species and with bacterial protein EC L11; Juan-Vidales F et al.; Two non-acidic proteins, extracted from the ribosomes of Saccharomyces cerevisiae using 1 M ammonium chloride in the presence of 50% ethanol, have been purified and characterized . Similar proteins are present in other eukaryotic ribosomes tested, as determined by two-dimensional gel electrophoresis and cross-reaction with antisera . One of the two yeast proteins, protein YL23, seems to be very well preserved during evolution, since antisera specific for YL23 cross-react with protein EC L11 from Escherichia coli . The structural similarity between these two proteins parallels a functional equivalence shown by the ability of the bacterial protein to reconstitute the activity of protein-deficient core particles from yeast . However, in contrast to protein EC L11, protein YL23 interacts with the yeast acidic proteins, forming a complex probably similar to the one made by bacterial protein EC L10 with proteins EC L7 and EC L12 in the E . coli ribosome . Protein YL23 might play similar roles to those of proteins EC L10 and EC L11 in bacteria.

Ann Microbiol (Paris), 1983 Nov-Dec, 134B(3), 379 - 85
Saccharomyces cerevisiae: heat and gluculase sensitivities of starved cells; Paris S et al.; Exponentially growing populations were abruptly shifted to media lacking a nitrogen source, a sulphur source or a phosphorus source . When proliferation ceased, cells were homogeneously arrested at the beginning of the cell cycle and were resistant to killing by exposure to 52 degrees C and to cell wall degrading enzymes . The results suggest that these two types of resistance represent a general response to nutrient limitation and are characteristic of resting cells.

J Cell Sci, 1983 Nov, 64, 307 - 22
Residual cell division measurements are unreliable as indicators of the timing of events in the Saccharomyces cerevisiae cell cycle; Richmond KM et al.; We report here an analysis of the execution point of the temperature-sensitive Saccharomyces cerevisiae cell cycle mutant, cdc27-47 . When a logarithmically growing culture was shifted from standard growth conditions (strain 27.8B growing in YEPD at 25 degrees C) to the restrictive temperature cell division ceased abruptly and reproducibly within one population doubling time, the extent of cell division indicating an execution point early in the cell cycle . Approximately 50% of stationary-phase cells were able to divide when refed with fresh medium at 37 degrees C, showing that the execution point could be passed before 'start' . This makes the sharp cut-off in cell division difficult to explain . This difficulty was compounded by observations of the cell cycle stage at which individual cells acquired the capacity to divide at 37 degrees C . Half the cells that were budded at the time of a temperature shift-up formed three division-blocked cells, and in 11 of these 13 cases, two were descended from the original mother cell and one from the original bud . Thus, mother and daughter cells pass the execution point independently; daughters usually during G1, and mothers usually in the budded phase of the previous cycle . The sharp cut-off in cell division is therefore spurious, and a mechanism is proposed to account for it, which has implications for the interpretation of the execution points of other cdc mutants . In addition, the expression of the cdc27-47 execution point was modified by both genetic and environmental factors, being affected by carbon source, by the petite condition, and by genetic background . This illustrates the difficulties of interpreting execution point data and the dangers of extrapolation of cell cycle parameters between strains and growth conditions.

Radiat Res, 1983 Nov, 96(2), 374 - 9
A comparison between rates of cell death and DNA damage during irradiation of Saccharomyces cerevisiae in N2 and N2O; Mitchel RE et al.; Yeast and several other organisms are more sensitive to the lethal effects of ionizing irradiation if exposed in the presence of N2O as compared to N2 . It has been suggested that this increased sensitivity is due to the cooperative effects of OH and H2O2 generated external to the cell wall . Using diploid yeast, wild type for radiation resistance, we have compared the rates of cell death due to gamma irradiation in N2 and N2O with the rates of DNA damage measured by gene conversion of trp- to trp+ (a recombinational repair event) . We find that DNA damage as measured by gene conversion increases at a faster rate, per unit dose, during irradiation in N2O as compared to N2, just as lethality was higher in N2O . When DNA damage was compared in N2 and N2O at equal levels of survival, however, there was no significant difference between the two irradiation conditions . Therefore, increased lethality during irradiation in N2O seems to be directly due to increased DNA damage . If the observed increased lethality results from external OH and H2O2, the effect of these highly reactive species is expressed by increased internal damage at the level of DNA.

Proc Natl Acad Sci U S A, 1983 Nov, 80(22), 6912 - 6
Gene conversion and associated reciprocal recombination are separable events in vegetative cells of Saccharomyces cerevisiae; Roman H et al.; Mitotic gene conversion occurs in both the G1 and G2 phases of cell growth . We postulate that the DNA strand or strands that connect the duplexes in G1 as a consequence of strand transfer are cleaved by an endonuclease . When this takes place in G1, crossing-over occurs in G2 at a frequency that is higher than would be expected from independent events . Conversion and associated reciprocal recombination are therefore separable with respect to the stage in the cell cycle when each occurs and possibly with respect to mechanism.

Exp Cell Res, 1983 Nov, 149(1), 15 - 26
Growth and the cell cycle of the yeast Saccharomyces cerevisiae . II . Relief of cell-cycle constraints allows accelerated cell divisions; Singer RA et al.; For cells of the yeast Saccharomyces cerevisiae, conditions which limit S phase or nuclear division allow steady-state division kinetics without significant effects on growth . Such cells become unusually large . When large proliferating cells were released from any one of several conditions which slowed progress through the DNA-division sequence, they underwent a period of accelerated division with a cell cycle devoid of a G1 interval, as evidenced by low proportions of unbudded cells and shifted execution points for the 'start' cell cycle step . We interpret these results to mean that when released from conditions slowing the DNA-division sequence these large cells continue for several cell doublings to accumulate mass fast enough to eliminate the need for a G1 interval . The results support the conclusion that the yeast G1 interval is the for most part only an interval of growth.

Exp Cell Res, 1983 Nov, 149(1), 1 - 13
Growth and the cell cycle of the yeast Saccharomyces cerevisiae . I . Slowing S phase or nuclear division decreases the G1 cell cycle period; Johnston GC et al.; Cells of the yeast Saccharomyces cerevisiae were subjected to a number of treatments which protracted S phase without proportional effects on growth processes . These treatments allowed steady-state exponential growth but lengthened the overall generation time . Such cells exhibited larger cell sizes and earlier performance of the cell cycle regulatory event 'start' and the two prereplicative steps defined by cdc4 and cdc7 mutations . Similarly, inhibiting progress through nuclear division with sub-arresting concentrations of methyl-benzimidazole-2-yl-carbamate also caused longer steady-state cell cycle times and earlier performance of 'start' . These findings underscore and extend earlier conclusions that most of the G1 interval of the yeast cell cycle is simply a period of ongoing growth . Conditions which protract any one of the periodic events in the division process without affecting growth will lead to the virtual elimination of the G1 interval.

J Bacteriol, 1983 Nov, 156(2), 907 - 8
Procedure for mutagenizing spores of Saccharomyces cerevisiae; Romano P et al.; A procedure for inducing mutants of a homothallic strain of Saccharomyces cerevisiae is described . The essential parts of the procedure are long incubation in Glusulase, which preferentially kills vegetative cells instead of spores, and treatment in 9% ethyl methanesulfonate, which also preferentially kills vegetative cells instead of spores . Consequently, the viable population is virtually 100% spores.

Gene, 1983 Nov, 25(2-3), 179 - 88
Plasmid-encoded hygromycin B resistance: the sequence of hygromycin B phosphotransferase gene and its expression in Escherichia coli and Saccharomyces cerevisiae; Gritz L et al.; The plasmid-borne gene hph coding for hygromycin B phosphotransferase (HPH) in Escherichia coli has been identified and its nucleotide sequence determined . The hph gene is 1026 nucleotides long, coding for a protein with a predicted Mr of 39 000 . The hph gene was placed in a shuttle plasmid vector, downstream from the promoter region of the cyc 1 gene of Saccharomyces cerevisiae, and an hph construction containing a single AUG in the 5' noncoding region allowed direct selection following transformation in yeast and in E . coli . Thus the hph gene can be used in cloning vectors for both pro- and eukaryotes.

J Bacteriol, 1983 Nov, 156(2), 898 - 900
Cyclic AMP may not be involved in catabolite repression in Saccharomyces cerevisiae: evidence from mutants unable to synthesize it; Matsumoto K et al.; Yeast cells with a nonsense adenylate cyclase mutation, cyr1-3, required cyclic AMP for growth . This phenotype was suppressed by the byc1 mutation; however, cyr1-3 bcy1 cells produced no detectable level of adenylate cyclase or cyclic AMP . On induction, the bcy1 and cyr1-3 bcy1 mutant cells produced the same levels of galactokinase and alpha-D-glucosidase as did the wild-type cells and fourfold-higher levels of invertase . Since galactokinase synthesis was severely repressed by glucose in the constitutive GAL81 mutants, irrespective of the cyr1-3 bcy1 genotype, cyclic AMP may not be involved in catabolite repression.

J Bacteriol, 1983 Nov, 156(2), 625 - 35
The presence of a defective LEU2 gene on 2 mu DNA recombinant plasmids of Saccharomyces cerevisiae is responsible for curing and high copy number; Erhart E et al.; The copy number of 2 mu DNA-derived plasmids in CIR+ Saccharomyces cerevisiae transformants is determined by its selective marker and is usually much lower than that of the endogenous plasmid . Only plasmids containing the leu2 allele of pJDB219, designated as leu2-d, under selective conditions displayed a higher copy number than did endogenous 2 mu DNA and by displacement generated cured cells . Spontaneous loss of 2 mu DNA occurred with a frequency of about 0.02% per generation . Curing plasmids, like pMP78, have copy numbers of 35; noncuring plasmids, like pDB248 or YEp6, have copy numbers of 4 to 8 . The 2 mu DNA copy number in strains AH22 and YNN27 were determined to be 40 and 100, respectively . The high copy number of leu2-d-containing plasmids can be explained by its weak expression of less than 5% that of the wild-type LEU2 gene . The leu2-d allele has a deletion of the 5'-end sequence starting from 29 base pairs before the ATG initiation codon, but surprisingly, its expression is still regulated . On YRp7, which contains the chromosomal autonomic replication sequence ARS1, the defective leu2-d allele could not complement a leu2 host strain . This suggests a more stringent control of replication of ARS1-containing plasmids than of 2 mu-containing plasmids.

Mol Cell Biol, 1983 Nov, 3(11), 1889 - 97
Oxalurate induction of multiple URA3 transcripts in Saccharomyces cerevisiae; Buckholz RG et al.; The URA3 gene from Saccharomyces cerevisiae is localized on a 1.1-kilobase (kb) DNA fragment . By using this fragment as a hybridization probe, we found that oxalurate, a gratuitous inducer of the allantoin degradative system, also serves to induce URA3 specific RNA . This response is restricted to oxalurate; other conditions which bring about high-level synthesis of the allantoin degradative enzymes did not produce the effect . Two classes of RNA (1.0 and 1.5 kb) were found to be oxalurate induced . Both classes are encoded by the URA3 gene, overlap, and probably do not significantly differ at their 5' termini . Northern blot mapping of the transcripts indicated that the 1.5-kb transcript was likely encoded by sequences extending up to 0.5 kb downstream from the 3' terminus of the 1.0-kb transcript . Analysis of the endpoints of the major 1.0-kb URA3 transcript by S1 nuclease mapping revealed the existence of two 5' termini, separated by 5 to 10 nucleotides, and seven 3' termini, separated by 5 to 20 nucleotides each, over a range of about 70 bases.

J Biol Chem, 1983 Oct 10, 258(19), 11648 - 53
Molecular cloning of fatty acid synthetase genes from Saccharomyces cerevisiae; Kuziora MA et al.; Fatty acid synthetase from Saccharomyces cerevisiae is a multifunctional enzyme which catalyzes the synthesis of long chain fatty acids from acetyl- and malonyl-CoA . The enzyme is composed of two nonidentical subunits, alpha (Mr = 212,000) and beta (Mr = 203,000), which are coded for by two unlinked genes FAS2 and FAS1, respectively . Individual yeast strains containing mutations in either of the FAS genes were transformed with a bank of yeast DNA sequences in the vector YEp13 . Plasmids YEpFAS1 and YEpFAS2 were selected by their ability to complement the fas1 or fas2 mutations, respectively . Additionally, we utilized an immunologic screening of a second yeast DNA bank and selected two clones 33F1 and 102B5 which produce antigenically reactive material to anti-yeast fatty acid synthetase antibodies . Through Southern hybridization experiments and restriction endonuclease mapping, a region of 5.3 kilobase pairs of 33F1 was shown to be homologous with YEpFAS1, and a span of 3.4 kilobase pairs of 102B5 was homologous with YEpFAS2 . These experiments identify the yeast DNA sequences cloned into 33F1 as originating from the FAS1 gene and those DNA sequences in 102B5, from the FAS2 gene.

Mol Cell Biol, 1983 Oct, 3(10), 1730 - 7
Cloning of Saccharomyces cerevisiae DNA replication genes: isolation of the CDC8 gene and two genes that compensate for the cdc8-1 mutation; Kuo CL et al.; The CDC8 gene, whose product is required for DNA replication in Saccharomyces cerevisiae, has been isolated on recombinant plasmids . The yeast vector YCp50 bearing the yeast ARS1, CEN4, and URA3 sequences, to provide for replication, stability, and selection, respectively, was used to prepare a recombinant plasmid pool containing the entire yeast genome . Plasmids capable of complementing the temperature-sensitive cdc8-1 mutation were isolated by transformation of a cdc8-1 mutant and selection for clones able to grow at the nonpermissive temperature . The entire complementing activity is carried on a 0.75-kilobase fragment, as revealed by deletion mapping . This fragment lies 1 kilobase downstream from the well-characterized sup4 gene, a gene known to be genetically linked to CDC8, thus confirming that the cloned gene corresponds to the chromosomal CDC8 gene . Two additional recombinant plasmids that complement the cdc8-1 mutation but that do not contain the 0.75-kilobase fragment or any flanking DNA were also identified in this study . These plasmids may contain genes that compensate for the lack of CDC8 gene product.

Radiat Res, 1983 Oct, 96(1), 95 - 9
Heat-shock induction of ultraviolet light resistance in Saccharomyces cerevisiae; Mitchel RE et al.; When exponentially growing diploid wild type Saccharomyces cerevisiae cells were subjected to a sudden rise in temperature (heat shock) they responded by increasing their resistance to the lethal effects of ultraviolet light . We have previously reported heat shock-induced increases in heat and ionizing radiation resistance . The shock-induced rise in resistance to uv light reported here was examined in terms of DNA repair capacity, and we find that the increase is due to induction of the recombinational repair system with no significant response from the uv-excision repair process.

Radiat Res, 1983 Oct, 96(1), 113 - 7
Assessment of the role of oxygen and mitochondria in heat shock induction of radiation and thermal resistance in Saccharomyces cerevisiae; Mitchel RE et al.; In response to a heat shock, the yeast Saccharomyces cerevisiae undergoes a large increase in its resistance to heat and, by the induction of its recombinational DNA repair capacity, a corresponding increase in resistance to radiation . Yeast which lack mitochondrial DNA, mitochondria-controlled protein synthetic apparatus, aerobic respiration, and electron transport rho 0 strain) were used to assess the role of O2, mitochondria, and oxidative processes controlled by mitochondria in the induction of these resistances . We have found that rho 0 yeast grown and heat shocked in either the presence or absence of O2 are capable of developing both radiation and heat resistance . We conclude that neither the stress signal nor its cellular consequences of induced heat and radiation resistance are directly dependent on O2, mitochondrial DNA, or mitochondria-controlled protein synthetic or oxidative processes.

J Bacteriol, 1983 Oct, 156(1), 36 - 48
Isolation and characterization of aminopeptidase mutants of Saccharomyces cerevisiae; Trumbly RJ et al.; Mutants of Saccharomyces cerevisiae were isolated which have decreased ability to hydrolyze leucine beta-naphthylamide, a chromogenic substrate for amino-peptidases . The mutations were shown by starch gel electrophoresis to affect one of four different aminopeptidases . Mutations affecting a given enzyme belong to a single complementation group . The four genes were symbolized lap1, lap2, lap3, and lap4, and the corresponding enzymes LAPI, LAPII, LAPIII, and LAPIV . Both lap1 and lap4 were mapped to the left arm of chromosome XI, and lap3 was mapped to the left arm of chromosome XIV . Strains which possessed only one of the four leucine aminopeptidases (LAPs) were constructed . Crude extracts from these strains were used to study the properties of the individual enzymes . Dialysis against EDTA greatly reduced the activity of all the LAPs except for LAPIII . Of the cations tested, Co2+ was the most effective in restoring activity . LAPIV was the only LAP reactivated by Zn2+ . LAPI was purified 331-fold and LAPII was purified 126-fold from cell homogenates . Both of the purified enzymes had strong activity on dipeptides and tripeptides . The activity levels of the LAPs are strongly dependent on growth stage in batch culture, with the highest levels in early-stationary phase . Strains lacking all four LAPs have slightly lower growth rates than wild-type strains . The ability of leucine auxotrophs to grow on dipeptides and tripeptides containing leucine is not impaired in strains lacking all four LAPs.

J Bacteriol, 1983 Oct, 156(1), 212 - 21
Temperature-sensitive Saccharomyces cerevisiae mutant defective in lipid biosynthesis; Letts VA et al.; A temperature-sensitive mutant of Saccharomyces cerevisiae (DAM303) is described that exhibits an early defect in lipid biosynthesis at the restrictive growth temperature, 37 degrees C . This strain rapidly lost viability after 1 h of incubation at 37 degrees C, and this was accompanied by a significantly reduced incorporation of 32Pi into cellular lipid and an accumulation of {1-14C}acetate into the free fatty acid fraction . The temperature-sensitive DAM303 mutation failed to complement the sec13 mutation described by Novick et al . (Cell 21:205-215, 1980), and from analysis of invertase secretion in the temperature-sensitive DAM303 strain, it is clear that the loss of invertase secretion in the mutant occurs after the loss of phospholipid synthesis . Although the precise nature of the temperature-sensitive lesion in the DAM303 strain has still to be identified, the results from the study of this mutant indicate that a defect in lipid biosynthesis can be correlated with subsequent alterations in extracellular protein secretion and loss of other macromolecular functions including DNA, RNA, and protein syntheses . From studies of this mutant, two procedures of enriching for other temperature-sensitive mutants with defects in lipid biosynthesis have emerged: inositol overproduction and screening for increased buoyant densities.

Can J Microbiol, 1983 Oct, 29(10), 1452 - 7
Phosphatidylglycerophosphate synthase activity in Saccharomyces cerevisiae; Carman GM et al.; Cytidine 5'-diphospho-1,2-diacyl-sn-glycerol (CDP-diacylglycerol): sn-glycerol-3-phosphate phosphatidyltransferase (phosphatidylglycerophosphate synthase, EC 2.7.8.5) activity was characterized from the mitochondrial fraction of Saccharomyces cerevisiae . The pH optimum for the reaction was 7.0 . Maximum activity was dependent on manganese (0.1 mM), magnesium (0.3 mM), or cobalt (1 mM) ions and the nonionic detergent Triton X-100 (1 mM) . The apparent Km values for CDP-diacylglycerol and glycerol-3-phosphate were 33 and 27 microM, respectively . Optimal activity was at 30 degrees C with an energy of activation of 5.4 kcal/mol (1 cal = 4.1868 J) . Phosphatidylglycerophosphate synthase activity was thermally labile above 40 degrees C . p-Chloromecuriphenylsulfonic acid, N-ethylmaleimide, and mercurous ions inhibited activity . Phosphatidylglycerophosphate synthase activity was partially solubilized from the mitochondrial fraction with 1% Triton X-100.

Braz J Med Biol Res, 1983 Oct, 16(3), 203 - 13
Regulation of porphyrin biosynthesis in yeast . Level of delta-aminolevulinic acid in porphyrin mutants of Saccharomyces cerevisiae; Malamud DR et al.; Saccharomyces cerevisiae mutants bearing mutations at the cyc4 locus are partially deficient in cytochrome synthesis . Although the mutation is not in the structural gene for delta-aminolevulinic acid (Alv) synthase, the mutants are deficient in Alv synthesis in vivo as indicated by abnormally low intracellular Alv concentrations . The cyc4 mutation causes cells to grow very slowly in minimal glucose medium, but not in yeast extract-peptone-glucose medium . A simple nutritional defect caused by the cyc4 mutation is not involved because cytochrome deficiency is enhanced by growing cyc4 cells in yeast extract-peptone medium . A regulatory role for CYC4 is indicated . Evidence for negative feed-back control of Alv synthase by heme is provided by the observation of enhanced intracellular Alv accumulation in yeast mutants partially deficient in decarboxylation of uroporphyrinogen and coproporphyrinogen, respectively.

J Bacteriol, 1983 Oct, 156(1), 421 - 3
Membrane-associated phosphatidylinositol kinase from Saccharomyces cerevisiae; McKenzie MA et al.; Membrane-associated phosphatidylinositol kinase (ATP:phosphatidylinositol 4-phosphotransferase, EC 2.7.1.67) was partially purified 93-fold from Saccharomyces cerevisiae . Activity was dependent on magnesium ions (10 mM) and the optimum pH was 8.5 . The apparent Km values for ATP and phosphatidylinositol were 0.21 mM and 71 microM, respectively . Activity was stimulated by sodium cholate and inhibited by sodium, potassium, lithium, and fluoride ions.

J Bacteriol, 1983 Oct, 156(1), 222 - 9
Two distinct subfractions in isolated Saccharomyces cerevisiae plasma membranes; Tschopp J et al.; The plasma membrane from Saccharomyces cerevisiae X2180-1A and a secretion-blocked mutant, secl (P . Novick and R . Schekman, Proc . Natl . Acad . Sci . U.S.A . 76:1858-1862, 1979) has been purified . Cell walls were digested by treatment with lyticase followed by concanavalin A coating of spheroplasts . alpha-Methylmannoside treatment after lysis, sonication at high salt concentration, and fractionation on a Renografin gradient resulted in two highly purified membrane fractions sedimenting at densities of 1.15 and 1.17 g/cm3 . Yields determined by recovery of vanadate-sensitive ATPase activity were 11 to 18%, and those determined by recovery of the spheroplast surface label 125I were 17 to 29% . Iodinated cells have most of their label in sedimentable, nonspheroplast material . However, both membrane populations contain some 125I surface label and show ATPase activity with pH optima only at 5.5 . The apparent Vmax of the plasma membrane ATPase equals 360 to 560 nmol of ATP hydrolyzed per min per mg of protein, with a Km for ATP of 0.7 mM . ATPase specific activity is not decreased in mutant plasma membrane . Analysis of 125I-labeled plasma membrane proteins by two-dimensional gel electrophoresis revealed seven major proteins on the plasma membrane surface.

Proc Natl Acad Sci U S A, 1983 Oct, 80(19), 5847 - 51
13C NMR studies of acetate metabolism during sporulation of Saccharomyces cerevisiae; Dickinson JR et al.; The sporulation of Saccharomyces cerevisiae in the presence of {2-13C}acetate was studied by 13C NMR spectroscopy . The fate of 13C label was analyzed in vivo and in cell extracts . During the first 4 hr of sporulation the major metabolite produced from {2-13C}acetate utilization was glutamate . From the labeling pattern observed it is concluded that both the tricarboxylic acid cycle and the glyoxylate cycle are operating . After about 4 hr trehalose is made . Comparison of the doublet/singlet ratios for C-1,1(1) and C-6,6(1) of trehalose shows a steady drop in the ratio of C-1, C-2-coupled species over trehalose labeled only at C-1 in the C-1, 2 segment of the molecule . The negative correlation of this ratio with that for the C-5, 6 segment indicates a cycling of glucose through the hexose monophosphate shunt . Subsequently fatty acid biosynthesis commences . Large amounts of saturated fatty acid were made . There were conspicuous differences observed in the metabolism of {2-13C}acetate between sporulating and vegetatively growing cells.

FEBS Lett, 1983 Sep 19, 161(2), 190 - 4
Detection of inactive precursors of beta-glucanases in Saccharomyces cerevisiae; Hernandez LM et al.; Accumulation and secretion of beta-glucanases have been studied in vivo by using a thermosensitive secretory mutant of Saccharomyces cerevisiae blocked at the endoplasmic reticulum level (sec 18-1) . When incubated at the restrictive temperature no accumulation of active glucanases was observed . Following a shift to permissive conditions in the presence of cycloheximide a rise in the internal activity took place . The increase in total glucanase activity was partially due to the activation of an exo-glucanase that hydrolyzes PNPG . It is concluded that glucanases are synthesized in inactive precursor forms and are converted to the active forms in their secretory pathway.

FEBS Lett, 1983 Sep 19, 161(2), 201 - 6
Non-random distribution of the Ty1 elements within nuclear DNA of Saccharomyces cerevisiae; Oyen TB et al.; Ty1 homologous sequences appear to be non-randomly distributed among different density classes of nuclear yeast DNA . Characteristic patterns of Ty1 containing EcoRI fragments can be generated from the various DNA fractions . The sequences are particularly enriched in the A + T rich part of the main nuclear DNA fraction, while the frequency in the rDNA containing heavy satellite DNA is low . The transposon however, seems to be present in this dense fraction, at least for some strains.

Biochem J, 1983 Sep 15, 214(3), 785 - 94
Correlation between the rate of proteolysis of mitochondrial translation products and fluidity of the mitochondrial inner membrane in Saccharomyces cerevisiae yeast . Alteration of the rate of proteolysis under glucose repression; Luzikov VN et al.; Our previous results {Kalnov, Novikova, Zubatov & Luzikov (1979) FEBS Lett . 101, 355-358; Biochem . J . 182, 195-202} suggested that in yeast the mitochondrial translation products localized in the mitochondrial inner membrane are rapidly broken down by a proteolytic system inherent in the membrane . In the present work, it is demonstrated that, on glucose repression in undividing cells of Saccharomyces cerevisiae, there is no proteolysis of the mitochondrial translation products . This effect is not likely to be associated with lower activity of the proteolytic system of the mitochondrial inner membrane . Nor is the cessation of proteolysis due to qualitative changes in the composition of mitochondrial translation products . What repression does cause is a considerable alteration in the physical state (i.e . structure of the lipid bilayer) of the mitochondrial inner membrane; this was established by experiments involving lipid-soluble spin probes . The conclusion is reached that the rate of proteolysis of mitochondrial translation products in the mitochondrial inner membrane depends on the physical state of the membrane, which in its turn is controlled by the relative content of unsaturated fatty acid chains in the mitochondrial phospholipids.

J Biol Chem, 1983 Sep 10, 258(17), 10720 - 6
Polypeptide structure of DNA polymerase I from Saccharomyces cerevisiae; Badaracco G et al.; DNA polymerase I of the yeast Saccharomyces cerevisiae has been purified to near homogeneity . The enzyme sediments under high salt conditions as a band at 7.4 S and two polypeptides of Mr = 140,000 and 110,000 are resolved by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate . Both polypeptides react with rabbit anti-yeast DNA polymerase I serum and can be shown to be enzymatically active by renaturation in situ after electrophoresis on polyacrylamide gels in the presence of sodium dodecyl sulfate . This high molecular weight form of yeast DNA polymerase I is very sensitive to inhibition by aphidicolin . The biochemical properties of the enzyme and inhibitors that may aid in distinguishing yeast DNA polymerases I and II are also described.

J Biol Chem, 1983 Sep 10, 258(17), 10200 - 3
The isolation and characterization of a mutant strain of Saccharomyces cerevisiae that requires a long chain base for growth and for synthesis of phosphosphingolipids; Wells GB et al.; A mutant of Saccharomyces cerevisiae has been obtained that shows an absolute growth requirement for long chain bases found in sphingolipids . In the absence of a long chain base, the cells are unable to synthesize the phosphoinositol-containing sphingolipids characteristic of yeast . These results suggest that one or more of the yeast sphingolipids plays a vital biological role.

Mikrobiologiia, 1983 Sep-Oct, 52(5), 744 - 9
{Component activity of the isoenzyme spectra of Saccharomyces cerevisiae, Saccharomyces carlsbergensis and their hybrids}; Orlova VS et al.; The object of this work was to study the activity and the isozyme spectra of hexokinase (the triggering enzyme of glycolysis), glucose-6-phosphate dehydrogenase (the key enzyme of the pentose-phosphate shunt), malate dehydrogenase and isocitrate dehydrogenase (the enzymes of the citric acid cycle) and alcohol dehydrogenase (the enzyme involved in the first steps of ethanol oxidation) in Saccharomyces cerevisiae, race Ya, S . carlsbergensis, race 4228, and their hybrid 67 . The parent organisms and their hybrid were shown to differ from one another in the qualitative composition and the activity of the isozyme spectra of the above enzymes.

Mikrobiologiia, 1983 Sep-Oct, 52(5), 719 - 22
{Effect of saccharose and lactose on the resistance of Saccharomyces cerevisiae yeast to desiccation}; Rapoport AI et al.; Incubation of Saccharomyces cerevisiae cells in solutions containing different sucrose or lactose concentrations (0.25 to 1.0 M) makes the organism more resistant to dehydration . The effect is increased when the cells are incubated for longer periods of time . Apparently, certain intracellular reactions making the yeast survive upon dehydration are initiated under these conditions.

Z Ernahrungswiss, 1983 Sep, 22(3), 205 - 12
{Effect of cadmium, zinc, lead and mercury on the enzyme activity in Saccharomyces cerevisiae in vitro}; Grafl HJ et al.; The difference between cadmium, zinc, lead, and mercury in regard of their effects on the activity of the enzymes tested is very slight . Concentrations higher than 10(-5) M reduce significantly the activity of the enzymes, and concentrations of approximately 10(-3) M inhibit it completely . An increase of the activity cannot be detected . The addition of combinations of cadmium, zinc, and lead results in a summing up of the toxic effects, whereas the interaction between mercury and the other three heavy metals shows a cumulative effect, which is appointed nearly completely by the heavy metal more toxic . The findings suggest that under in-vitro conditions there exists a direct interaction between the heavy metals and the enzymes.

Mol Cell Biol, 1983 Sep, 3(9), 1665 - 9
Stably denatured regions in chromosomal DNA from the cdc2 Saccharomyces cerevisiae cell cycle mutant; Conrad MN et al.; DNA isolated from Saccharomyces cerevisiae strains carrying temperature-sensitive mutations in the CDC2 gene after incubation at the restrictive temperature contains multiple stably denatured regions 200 to 700 base pairs long . These regions are probably stabilized by a DNA-binding protein . They are found in both replicated and unreplicated portions of DNA molecules, suggesting that they are not an early stage in the initiation of DNA replication.

Mol Cell Biol, 1983 Sep, 3(9), 1615 - 24
Transcriptional analysis of the Saccharomyces cerevisiae mitochondrial var1 gene: anomalous hybridization of RNA from AT-rich regions; Zassenhaus HP et al.; A family of mitochondrial RNAs hybridizes specifically to the var1 region on Saccharomyces cerevisiae mitochondrial DNA (Farrelly et al., J . Biol . Chem . 257:6581-6587, 1982) . We constructed a fine-structure transcription map of this region by hybridizing DNA probes containing different portions of the var1 region and some flanking sequences to mitochondrial RNAs isolated from var1-containing petites . We also report the nucleotide sequence of more than 1.2 kilobases of DNA flanking the var1 gene . Our primary findings are: (i) The family of RNAs we detect with homology to var1 DNA is colinear with the var1 gene . Their direction of transcription is olil to cap, as it is for most other mitochondrial genes . (ii) Extensive hybridization anomalies are present, most likely due to the high A-T (A-U) content of the hybridizing species and to the asymmetric distribution of their G-C residues . An important conclusion is that failure to detect transcripts from A-T-rich regions of the yeast mitochondrial genome by standard blot transfer hybridizations cannot be interpreted to mean that such sequences, which are commonly supposed to be spacer DNA, are noncoding or lack direct function in the expression of mitochondrial genes.

Arch Biochem Biophys, 1983 Sep, 225(2), 562 - 75
In situ behavior of the pyrimidine pathway enzymes in Saccharomyces cerevisiae . I . Catalytic and regulatory properties of aspartate transcarbamylase; Penverne B et al.; A permeabilization procedure was adapted to allow the in situ determination of aspartate transcarbamylase activity in Saccharomyces cerevisiae . Permeabilization is obtained by treating cell suspensions with small amounts of 10% toluene in absolute ethanol . After washing, the cells can be used directly in the enzyme assays . Kinetic studies of aspartate transcarbamylase (EC 2.1.3.2) in such permeabilized cells showed that apparent Km for substrates and Ki for the feedback inhibitor UTP were only slightly different from those reported using partially purified enzyme . The aspartate saturation curve is hyperbolic both in the presence and absence of UTP . The inhibition by this nucleotide is noncompetitive with respect to aspartate, decreasing both the affinity for this substrate and the maximal velocity of the reaction . The saturation curves for both substrates give parallel double reciprocal plots . The inhibition by the products is linear noncompetitive . Succinate, an aspartate analog, provokes competitive and uncompetitive inhibitions toward aspartate and carbamyl phosphate, respectively . The inhibition by phosphonacetate, a carbamyl phosphate analog, is uncompetitive and noncompetitive toward carbamyl phosphate and aspartate, respectively, but pyrophosphate inhibition is competitive toward carbamyl phosphate and noncompetitive toward aspartate . These results, as well as the effect of the transition state analog N-phosphonacetyl-L-aspartate, all exclude a random mechanism for aspartate transcarbamylase . Most of the data suggest an ordered mechanism except the substrates saturation curves, which are indicative of a ping-pong mechanism . Such a discrepancy might be related to some channeling of carbamyl phosphate between carbamyl phosphate synthetase and aspartate transcarbamylase catalytic sites.

Proc Natl Acad Sci U S A, 1983 Sep, 80(17), 5374 - 8
Positive regulation in the general amino acid control of Saccharomyces cerevisiae; Hinnebusch AG et al.; Starvation of yeast for a single amino acid leads to derepression of enzymes in many different amino acid biosynthetic pathways . This general control is regulated by several transacting genes . Mutations in the TRA3 gene result in constitutive derepression, whereas mutations in AAS genes lead to the inability to derepress . We have isolated aas mutations as suppressors of the tra3-1 mutation . Some of these suppressors are alleles of AAS2 and others define a heretofore unidentified gene, AAS3 . We have studied the regulatory behavior of strains containing both aas and tra3 mutations and strains containing the cloned AAS genes in high copy number . Either aas1- or aas2- in combination with tra3- has the Tra- phenotype, whereas aas3- in combination with tra3- has the Aas- phenotype . These interactions suggest that the AAS1 and AAS2 products act indirectly to bring about derepression by disabling the repressive effect of TRA3, whereas the AAS3 product functions more directly and is required even in the absence of the TRA3 function . When present in high copy number, the AAS3 gene complements mutations in AAS1 and AAS2, whereas AAS1 and AAS2 only complement their cognate mutations . Taken together these data suggest that AAS1 and AAS2 are negative regulators of TRA3, which in turn is a negative regulator of AAS3 . AAS3 is a positive regulator, which is required for the general control response . This model of negative and positive interactions is formally identical to those proposed for the regulation of the galactose and phosphatase systems in yeast.

Proc Natl Acad Sci U S A, 1983 Sep, 80(17), 5185 - 9
39K, 23Na, and 31P NMR studies of ion transport in Saccharomyces cerevisiae; Ogino T et al.; The relationship between efflux and influx of K+, Na+, and intracellular pH (pHin) in yeast cells upon energizing by oxygenation was studied by using the noninvasive technique of 39K, 23Na, and 31P NMR spectroscopy . By introducing an anionic paramagnetic shift reagent, Dy3+(P3O5(-10))2, into the medium, NMR signals of intra- and extracellular K+ and Na+ could be resolved, enabling us to study ion transport processes by NMR . Measurements showed that 40% of the intracellular K+ and Na+ in yeast cells contributed to the NMR intensities . By applying this correction factor, the intracellular ion concentrations were determined to be 130-170 mM K+ and 2.5 mM Na+ for fresh yeast cells . With the aid of a home-built solenoidal coil probe for 39K and a double-tuned probe for 23Na and 31P, we could follow time courses of K+ and Na+ transport and of pHin with a time resolution of 1 min . It was shown that H+ extrusion is correlated with K+ uptake and not with Na+ uptake upon energizing yeast cells by oxygenation . When the cells were deenergized after the aerobic period, K+ efflux, H+ influx, and Na+ influx were calculated to be 1.6, 1.5, and 0.15 mumol/min per ml of cell water, respectively . Therefore, under the present conditions, K+ efflux is balanced by exchange for H+ with an approximate stoichiometry of 1:1.

J Bacteriol, 1983 Sep, 155(3), 995 - 1000
Transport of 6-deoxyglucose in Saccharomyces cerevisiae; Bisson LF et al.; The uptake of 6-deoxyglucose was measured in wild-type Saccharomyces cerevisiae, in a double mutant strain lacking activity for hexokinases A and B (hxkl hxk2), in a triple mutant strain lacking activity for both hexokinases and glucokinase (hxkl hxk2 glk), and in the triple mutant with high levels of activity of single kinases restored by introduction of the cloned genes . In the wild-type strain, uptake of the glucose analog showed two components, with Km values of ca . 20 mM ("high affinity") and 250 mM ("low affinity"), respectively . The double mutant also had high- and low-affinity components, but the triple mutant showed only low-affinity uptake . Reintroduction of the single kinases to the triple mutant restored high-affinity uptake . (Other experiments on 6-deoxyglucose uptake are also presented, including the apparent use of the galactose transport system when induced.) These results show that the recent implication of the kinases in transport of glucose (L.F . Bisson and D.G . Fraenkel, Proc . Natl . Acad . Sci . U.S.A . 80:1730-1734, 1983) applies equally to the nonmetabolized analog 6-deoxyglucose and suggests that the role of the kinases in transport is not merely a consequence of metabolism of the transported compound.

J Bacteriol, 1983 Sep, 155(3), 1178 - 84
Events associated with restoration by zinc of meiosis in apomictic Saccharomyces cerevisiae; Bilinski CA et al.; The effects of nutritional alterations (carbon source and zinc) on nuclear division and protein synthesis during apomictic and meiotic development in Saccharomyces cerevisiae 19e1 were investigated . Unlike cells cultivated under meiosis-promoting conditions, cells cultured under apomixis-promoting conditions exhibited extensive protein synthesis during the first 3 h of incubation in sporulation medium, and nuclear divisions were evident during this time . Cycloheximide treatment of the latter cells induced meiosis, and maximum yields of meiotic asci resulted when this treatment was given for the first 3 h in sporulation medium . The results indicate that the decision concerning which developmental route cells will follow is made shortly after transfer to sporulation medium . Electrophoretic analysis of labeled proteins synthesized during sporulation revealed bands unique to both developmental routes.

Can J Microbiol, 1983 Sep, 29(9), 1200 - 4
Control of catalase and peroxidase biosynthesis by carbon source and oxygen in the yeast Saccharomyces cerevisiae; Valdivia E et al.; The effect of carbon source and oxygen tension on catalase and peroxidase levels and on the intermediates of the biosynthesis of the prosthetic group of both enzymes has been studied . Oxygen produces an increase of both enzymatic activities, even in presence of glucose . On the other hand it seems probable that glucose does not have a direct inhibitory effect on the biosynthesis of 5-aminolevulinic acid (ALA) and porphyrins.

Gene, 1983 Sep, 24(1), 1 - 14
Efficient synthesis of enzymatically active calf chymosin in Saccharomyces cerevisiae; Mellor J et al.; We have constructed a high-efficiency expression vector to direct the synthesis of heterologous polypeptides in yeast . The vector is termed a sandwich expression vector as the heterologous gene is inserted between the 5' and 3' control regions of the efficiently expressed yeast PGK gene . We have used this vector to direct the expression of three derivatives of the calf chymosin cDNA gene; preprochymosin, prochymosin and chymosin . Prochymosin is synthesised to at least 5% of total yeast-cell protein and furthermore, it can be readily activated to produce an enzyme which has milk-clotting activity.

Proc Natl Acad Sci U S A, 1983 Sep, 80(17), 5345 - 9
pif mutation blocks recombination between mitochondrial rho+ and rho- genomes having tandemly arrayed repeat units in Saccharomyces cerevisiae; Foury F et al.; Three allelic nuclear mutants affected in the recombination of mtDNA have been characterized in Saccharomyces cerevisiae and assigned to the PIF locus . In the mutants, the general recombination measured by the recombination frequency between linked or unlinked alleles is normal . However, the pif mutations prevent the integration into the rho+ genome of the markers (oli1, oli2, diu1, ery, oxi1, oxi2) of those rho- genomes that have tandemly arrayed repeat units . Therefore, these rho- genomes characterize a PIF-dependent recombination system . The pif mutations have also revealed the existence of a PIF-independent recombination system used by those rho- genomes that have an inverted organization of their repeat units . The markers of such palindromic rho- genomes exhibit high integration frequency into the rho+ genome even in the presence of the pif mutation . In addition, the pif mutations greatly increase suppressiveness in crosses between pif rho+ strains and PIF-dependent as well as PIF-independent rho- clones . We conclude that the recombination between rho+ and rho- genomes involves at least two distinct systems that depend on the organization of the rho- genome.

Zentralbl Bakteriol Mikrobiol Hyg {B}, 1983 Sep, 177(6), 514 - 26
{Effect of heavy metal interactions on the growth of Saccharomyces cerevisiae}; Grafl HJ et al.; The influence of interactions of cadmium, zinc, lead, and mercury on the growth of Saccharomyces cerevisiae has been studied . Generally the experiments resulted in the following findings: Low zinc concentrations reduce the toxicity of cadmium while high zinc concentrations intensify the effects of cadmium . Zinc does not decrease the growth inhibition by mercury . The actions of cadmium and mercury are not changed by lead, but 5 X 10(-4) M lead prevent completely the increase of the growth intensity caused by zinc . Combinations of toxic concentrations of cadmium and mercury show a synergistic lengthening of the lag period, but their cumulative influence on the growth rate is lower than the sum of the corresponding inhibition effects.

FEBS Lett, 1983 Aug 22, 160(1-2), 195 - 7
Calcium uptake during the cell cycle of Saccharomyces cerevisiae; Saavedra-Molina A et al.; Synchronous culture of the budding yeast Saccharomyces cerevisiae was obtained by sucrose density gradient selection with 90-100% of yeast synchronized by using the cells in the bottom . In these adult cells bud emergence is coincident with an increase in calcium uptake at 100 min of the culture, followed by a return to basal values which are maintained until the end of the first cell cycle of study . The phenothiazine derivatives, trifluoperazine and chlorpromazine inhibit bud emergence and trifluoperazine also increases calcium uptake.

Biochim Biophys Acta, 1983 Aug 16, 746(3), 133 - 7
Studies on aconitase species from Saccharomyces cerevisiae, porcine and bovine heart, obtained by a modified isolation method; Scholze H; Aconitase (citrate (isocitrate) hydro-lyase, EC 4.2.1.3) was isolated from Saccharomyces cerevisiae, porcine and bovine heart by a simplified method including affinity chromatography on Blue Dextran-Sepharose . Partial characterisation reveals that the aconitase species are all similar due to molecule size, amino acid composition, isoelectric point and enzymatic activity . Aconitase appears as a single polypeptide chain with a small carbohydrate content . A molecular weight of 79000 +/- 2000 and a Svedberg constant of s20,w = 4.75 +/- 0.2 S indicate a compact structure of aconitase . Due to different properties among the yeast aconitase species concerning isoelectric point and enzymatic activity a coherence between net charge of the protein and redox state of the Fe-S cluster can be expected.

J Mol Biol, 1983 Aug 15, 168(3), 505 - 23
A family of Saccharomyces cerevisiae repetitive autonomously replicating sequences that have very similar genomic environments; Chan CS et al.; We have characterized a family of moderately repetitive autonomously replicating sequences (ARSs) in Saccharomyces cerevisiae . Restriction mapping, deletion studies and hybridization studies suggest that these ARSs, which are probably less than 350 base-pairs in size, share one common feature: each is located close to, but not within, a repetitive sequence (131) of approximately 10(3) to approximately 1.5 X 10(3) base-pairs in length . These ARSs can be divided into two classes (X and Y) by their sequence homology and genomic environments . Each of the class X ARSs is embedded within a repetitive sequence (X) of variable length (approximately 0.3 X 10(3) to approximately 3.75 X 10(3) base-pairs); each of the class Y ARSs is embedded within a highly conserved repetitive sequence (Y) of approximately 5.2 X 10(3) base-pairs in length . Both of these sequences are located directly adjacent to the 131 sequence.

J Biol Chem, 1983 Aug 10, 258(15), 9245 - 9
Fructose 2,6-bisphosphate and fructose-6-P 2-kinase in Saccharomyces cerevisiae in relation to metabolic state in wild type and fructose-6-P 1-kinase mutant strains; Clifton D et al.; In wild type Saccharomyces cerevisiae, fructose-6-P is known to be in much lower amounts than needed to saturate fructose-6-P 1-kinase in vitro, and the same is true for a mutant with reduced affinity for fructose-6-P, even though its in vivo fructose-6-P concentration is much higher than normal . Both the wild type and mutant fructose-6-P 1-kinases were activated in vitro by fructose-2,6-P2 in the 0.1 microM concentration range, and the effector was present in more than adequate amounts . Hence, it is likely to be necessary for sufficient flux through the fructose-6-P 1-kinase reaction in vivo, and the data also fit with fructose-2,6-P2 acting at different sites on the enzyme from fructose-6-P . In growth on glucose, a variety of wild type strains contained 5-10 microM fructose-2,6-P2, and various fructose-6-P 1-kinase mutant strains had levels of up to 150 microM in the presence of glucose . Fructose-2,6-P2 was also found (0.5-10 microM) in derepressed cultures after glucose exhaustion and in growth on pyruvate . Activities of fructose-6-P 2-kinase in the various strains and situations are also presented . The data generally indicate a correlation between levels of fructose-2,6-P2 and fructose-6-P and suggest that fructose-2,6-P2 is not rapidly degraded after glucose exhaustion.

J Biochem (Tokyo), 1983 Aug, 94(2), 501 - 10
Characterization of a Saccharomyces cerevisiae mutant, N22, defective in ergosterol synthesis and preparation of {28-14C}ergosta-5,7-dien-3 beta-ol with the mutant; Hata S et al.; Analysis of sterols was made with a Saccharomyces cerevisiae mutant, N22, which was resistant to nystatin and defective in ergosterol synthesis, and with its parent strain, M10 (haploid type, methionineless, petite) . The main sterol of M10 was ergosterol, whereas that of N22 was ergosta-5,7-dien-3 beta-ol . A small amount of ergosterol was found also in N22 . This indicates that the delta 22-desaturation reaction in N22 is blocked, though not completely . Ergosta-5,8-dien-3 beta-ol, an unusual sterol, was detected in N22 but not in M10 . Variations in the amounts and compositions of free and esterified sterols of both strains were examined during cultivation and the subsequent aerobic adaptation . The contents of free sterols, which were mostly composed of the respective main sterols of both strains, did not change markedly . In contrast, the contents and compositions of esterified sterols of both strains varied depending on the growth conditions . When the cells of both strains were aerobically adapted, the total contents of esterified sterols increase, as did, as the intermediate sterols . Upon aerobic adaptation in the presence of {methyl-14C}methionine, {28-14C}ergosta-5,7-dien-3 beta-ol accumulated markedly in N22 cells . Using this mutant, we devised a convenient method for preparation of the radioactive sterol in high yield.

Genetika, 1983 Aug, 19(8), 1384 - 6
{Detection of the loss of 1 chromosome from pair III in Saccharomyces cerevisiae yeasts}; Bandas EL; A diploid strain of Saccharomyces cerevisiae, T6 is described which monitors both mitotic crossing over and induction of aneuploidy . The chromosome III carries recessive markers: rgh12 of "rough colony" phenotype closely linked to centromere, the left arm is marked with his4, the right arm is marked both with thr4 and the locus of mating type alpha . Expression of all the markers on chromosome III leads to formation of colonies which are rough, require histidine and threonine, and are of alpha mating type . These colonies arise as a result of the loss of a chromosome during mitosis, which makes the strain allow detection of monosomic cells formation . Chromosome XV carries two phenotypically distinguishable and recessive alleles of the gene ade2: ade2-192 (causes red coloration of colonies) and ade2-G45 (causes pink coloration of colonies) . Mitotic crossing over generates two reciprocal products which can be revealed together in colonies as pink and red sectors in double-spotted colonies . Both double-spotted and monosomic colonies have been obtained after treatment with gamma-rays . The frequency of mitotic crossing over after irradiation by 1000-3000 Gray increased up to 2-3.2% (the spontaneous level was 0.006%), the frequency of aneuploidy was 0.12 to 0.57% at plating immediately after irradiation (the spontaneous monosomics were not observed among 1.5 X 10(5) cells scored) . Induction of mitotic crossing over and aneuploidy by benomyl was rather slight (up to 0.05 and 0.006%, respectively).

Arch Microbiol, 1983 Aug, 135(1), 63 - 7
Nucleotide pools of growing, synchronized and stressed cultures of Saccharomyces cerevisiae; Ditzelmuller G et al.; High pressure liquid chromatography has been used to study the acid soluble nucleotide pool of Saccharomyces cerevisiae under different conditions of growth . ATP, ADP, AMP, NAD, GTP, UTP, UDP, CTP, CDP, and UDP-sugars plus UMP could be separated and were found in concentrations higher than 0.1 mumol per g yeast cell dry weight (= detection limit) . During glucose-limited continuous culture the levels of individual nucleotides depended on the growth rate, which was most pronounced with pyrimidine (uridine, cytidine) nucleotides . The energy charge (E.C.) remained high (0.9) at all growth rates (0.07-0.3 h-1) . During synchronized growth at a constant growth rate (0.11 h-1) almost all nucleotide levels and the E.C . remained at constant values with the only exception of UDP-sugars and UMP of which increased levels were found during the phase of budding . Under conditions of metabolic stress (addition of antimycin A, deoxyglucose plus iodoacetate) pronounced changes in the levels of purine (adenine and guanine) nucleotides and the E.C . were observed . All other nucleotides were less influenced by these conditions . Only under these conditions IMP accumulation was observed . The results strongly argue against the significance of purine nucleotide or E.C . measurements under viable conditions . In contrast, changes in the levels of pyrimidine nucleotides seem to be indicative of changes in the flux through the metabolic pathways where they act as coenzymes.

Mutat Res, 1983 Aug, 121(2), 117 - 23
Cytochrome P-450 factors determining synthesis in strain D7 Saccharomyces cerevisiae . An alternative system to microsomal assay; Del Carratore R et al.; Certain aspects of cytochrome P-450 induction were studied in a diploid strain (D7) of the yeast Saccharomyces cerevisiae in order to obtain cells containing a high level of metabolizing enzymes . The highest level of cytochrome P-450 was reached during the logarithmic growth phase in a 20%-glucose liquid medium . Yeast cells harvested in these conditions were used in the mutagenesis test with dimethyl nitrosamine (DMNA) as a positive control and with styrene (Sty) . Both substances gave positive results, whereas Sty never showed any mutagenic activity in the conventional test with stationary growth phase cells and external metabolic activation . The test with cells from the logarithmic growth phase is proposed as a possible alternative to the liver-microsome assay, and its reliability is discussed.

J Bacteriol, 1983 Aug, 155(2), 903 - 6
Extracellular suppression allows mating by pheromone-deficient sterile mutants of Saccharomyces cerevisiae; Chan RK et al.; MAT alpha haploids with mutations in the STE13 or KEX2 gene, and MATa haploids with mutations in the STE6 or STE14 gene, do not mate with wild-type cells of the opposite mating type . We found that such mutants were able to mate with partners that carry mutations (sst1 and sst2) that cause cells to be supersensitive to yeast mating pheromone action . Mating ability of MAT alpha ste13 and MAT alpha kex2 mutants could also be restored by adding normal MAT alpha cells to mating mixtures or by adding just the appropriate purified pheromone (alpha-factor) . Therefore, the mating deficiencies caused by the ste13 and kex2 lesions, and by inference, the ste6 and ste14 mutations, appear to result only from secretion of an insufficient amount of pheromone or a nonfunctional pheromone.

J Bacteriol, 1983 Aug, 155(2), 488 - 92
Oxygen requirements for formation and activity of the squalene epoxidase in Saccharomyces cerevisiae; Jahnke L et al.; The effect of oxygen on squalene epoxidase activity in Saccharomyces cerevisiae was investigated . In cells grown in standing cultures, the epoxidase was localized mainly in the "mitochondrial" fraction . Upon aeration, enzyme activity increased and the newly formed enzyme was associated with the "microsomal" fraction . At 0.03% (vol/vol) oxygen, epoxidase levels doubled, whereas the ergosterol level was only slightly increased . Cycloheximide inhibited the increase in epoxidase under these conditions . An apparent Km for oxygen of 0.38% (vol/vol) was determined from a crude particulate preparation for the epoxidase.

Biochimie, 1983 Aug-Sep, 65(8-9), 501 - 12
Potentiation of oxygen toxicity by menadione in Saccharomyces cerevisiae; Chaput M et al.; The cytotoxicity of molecular oxygen can be sharply increased in the yeast Saccharomyces cerevisiae by the use of redox compounds capable of shunting electrons in vivo and of spontaneous reoxidation under aerobic conditions . Among these redox compounds, menadione (Vitamin K3) is particularly able to stimulate the cyanide-resistant respiration of the yeast cells . Under steady-state conditions, the efficiency of menadione is modulated by the physiological state of the yeast cells and also depends on the availability of reducing agents within the cell . Menadione shows lethal effects towards yeast cells in the presence of O2 only, as a result of the production of toxic metabolites like O2- . and H2O2 which are actually detected in the extracellular fluid . Inhibitors of the enzymes scavenging O2- . and H2O2 generally potentiate the lethal effects of this redox compound . On the other hand, superoxide dismutase and/or catalase supplemented into the incubation buffer have been found to protect the cells to various extents from the cytotoxic effects of menadione . Our data support the following conclusions: When the cellular enzymatic defences are functional, the moderate lethality induced by menadione is principally mediated by O2- . ions acting on the outer side of the cell (peripheral region) . In the presence of cyanide, but not of azide, the loss of viability also results from additional damage occurring within the inner cell region . In this case, intracellular injury can be caused by H2O2 alone but our data also suggest that during redox cycling more reactive species--O2- . and probably OH.--are generally intracellularly and are involved in the cytotoxic process.

Proc Natl Acad Sci U S A, 1983 Aug, 80(15), 4818 - 21
A DNA repair gene required for the incision of damaged DNA is essential for viability in Saccharomyces cerevisiae; Naumovski L et al.; A diploid strain (RAD3/RAD3) of Saccharomyces cerevisiae was transformed with an integration plasmid containing an internal fragment of the cloned yeast RAD3 gene . Integration by homologous recombination inactivated one of the diploid RAD3 genes, creating a recessive mutation . This mutation is inferred to be lethal in haploid cells since sporulation of diploid transformants segregated two viable and two inviable spores per tetrad, while integration of plasmids containing one or the other end of the RAD3 gene resulted in diploid transformants that segregated normally--i.e., four viable spores in each tetrad . Evidence that integration of the internal fragment occurred specifically at one of the RAD3 genes in the diploid is provided by DNA . DNA hybridizations . In addition, transformation of a diploid strain heterozygous for the RAD3 gene (RAD3/rad3-2) (carrying a rad3 mutation that does not affect the viability of haploid cells) results in the rad- phenotype in half of the transformants, indicating that the RAD3 gene was inactivated in these cells.

J Bacteriol, 1983 Aug, 155(2), 747 - 54
In vivo ligation of linear DNA molecules to circular forms in the yeast Saccharomyces cerevisiae; Suzuki K et al.; The yeast Saccharomyces cerevisiae was transformed with restriction endonuclease-digested (linear) DNAs containing the replication origin of the yeast 2 microns plasmid and selectable markers with efficiencies of 10(3) to 10(4), 10(3), and 10(2) to 10(3) transformants per microgram of DNA in the cases of transformations with linear DNAs containing the same cohesive ends, flush ends, and non-complementary cohesive ends, respectively . The results of a restriction analysis of the circular plasmids recovered from transformed cells suggested that the linear DNA molecules were ligated to produce circular forms in the recipient protoplasts.

J Bacteriol, 1983 Aug, 155(2), 623 - 7
What is the function of nitrogen catabolite repression in Saccharomyces cerevisiae?
Cooper TG, Sumrada RA.
In contrast to the previously held notion that nitrogen catabolite repression is primarily responsible for the ability of yeast cells to use good nitrogen sources in preference to poor ones, we demonstrate that this ability is probably the result of other control mechanisms, such as metabolite compartmentation . We suggest that nitrogen repression is functionally a long-term adaptation to changes in the nutritional environment of yeast cells.

J Immunol Methods, 1983 Jul 29, 61(3), 351 - 7
A new method of preparing hapten-carrier immunogens by coupling with Saccharomyces cerevisiae by periodate oxidation; Bernard D et al.; A new method of preparing hapten-carrier immunogens is described . The carrier used was Saccharomyces cerevisiae yeast . Coupling was carried out with bovine serum albumin (BSA) . BSA was conjugated with mannose residues of Saccharomyces cerevisiae cell walls, following periodate oxidation to produce aldehyde groups reactive with side chain amino groups of BSA . The coupling was stabilized and non-reactant aldehydes were blocked with sodium borohydride reduction . A hapten, 3,5,3',5'-tetraiodo-L-tyrosyl-L-tyrosine, was also coupled with the yeast . Anti-hapten antibodies were obtained in rabbit with this conjugate.

FEBS Lett, 1983 Jul 25, 158(2), 247 - 51
Saccharomyces cerevisiae alpha-factor prevents formation of glycoproteins in a cells; Orlean P et al.; alpha-Factor inhibits incorporation of {14C}glucosamine into water-soluble and into SDS-extractable glycoproteins to about 90% . The incorporation into chitin is not affected and the same is true for {14C}phenylalanine incorporation into protein . The inhibition of glycoprotein synthesis is specific for a cells.

Eur J Biochem, 1983 Jul 15, 134(1), 117 - 21
Partial purification and characterization of mRNA (guanine-7-) methyltransferase from the yeast Saccharomyces cerevisiae; Locht C et al.; As a tool for the study of the capping-methylation process of yeast mRNA, we developed a procedure for the purification of the mRNA (guanine-7-)methyltransferase using the commercial cap analog guanosine(5')triphospho(5')guanosine as a substrate and radioactive S-adenosylmethionine (AdoMet) as the methyl group donor . The osmotic-sensitive yeast strain VY 1160 was used as the enzyme source . Little methyltransferase activity was detectable in a crude lysate obtained after osmotic shock . We showed that this was due to the presence of a low-molecular-weight inhibitor which could easily be eliminated by Sephadex G-25 gel filtration . The 10000 X g supernatant from the crude lysate was submitted to DEAE-cellulose and DNA-agarose chromatography . The resulting preparation was enriched about 450-fold in specific activity . Under standard assay conditions, the incorporation rate remained constant for at least 6 h at 30 degrees C . Transmethylation was not stimulated by KCl nor NaCl . Divalent cations were strong inhibitors . The partially purified enzyme was able to methylate undermethylated poly(A)-rich mRNA isolated from an AdoMet auxotrophic yeast strain briefly exposed to AdoMet-free medium.

Biochim Biophys Acta, 1983 Jul 13, 732(1), 186 - 92
All-or-none interactions of inhibitors of the plasma membrane ATPase with Saccharomyces cerevisiae; Borst-Pauwels GW et al.; The inhibitors of the yeast plasma membrane ATPase, Dio-9, miconazole, suloctidil, N,N'-dicyclohexycarbodiimide, triphenyltin and diethylstilbestrol provoke an all-or-none K+ loss from the cells . Some of the K+-depleted cells also show an increased permeability for the dye, Bromophenol blue, indicating that the barrier properties of these cells are drastically changed . Apart from this all-or-none process, a graded loss of K+ is also observed . These observations warn against the use of the inhibitors in studies aimed at evaluating the role of the ATPase in the energy transduction of membrane transport processes of yeasts.

Nucleic Acids Res, 1983 Jul 11, 11(13), 4435 - 51
Biogenesis of mitochondria: the mitochondrial gene (aap1) coding for mitochondrial ATPase subunit 8 in Saccharomyces cerevisiae; Macreadie IG et al.; A mitochondrial gene (denoted aap1) in Saccharomyces cerevisiae has been characterized by nucleotide sequence analysis of a region of mtDNA between the oxi3 and oli2 genes . The reading frame of the aap1 gene specifies a hydrophobic polypeptide containing 48 amino acids . The functional nature of this reading frame was established by sequence analysis of a series of mit- mutants and revertants . Evidence is presented that the aap1 gene codes for a mitochondrially synthesized polypeptide associated with the mitochondrial ATPase complex . This polypeptide (denoted subunit 8) is a proteolipid whose size has been previously assumed to be 10 kilodaltons based on its mobility on SDS-polyacrylamide gels, but the sequence of the aap1 gene predicts a molecular weight of 5,815 for this protein.

Biochem Int, 1983 Jul, 7(1), 15 - 22
Amino acid transport: its role in cell division and growth of Saccharomyces cerevisiae cells; Dudani AK et al.; The transport of six amino acids was studied in exponentially growing and G1 arrested temperature sensitive mutants of the yeast Saccharomyces cerevisiae . The uptake of three amino acids viz . Gly, Ala and Val was selectively reduced in G1 arrested cells . The reduction in transport was only noticeable when cdc 28 mutant cells were arrested at their non-permissive temperature . Cdc 28 is known to arrest at the point of commitment to the cell cycle 'start' . The uptake, however, was unaffected in G1 arrested cdc 4 and cdc 7 cells . The wild type cells (A364A) when arrested at stationary phase, also demonstrated the reduction in the uptake of Ala and Val . It appears that amino acid transport may be involved in growth and cell division of Saccharomyces cerevisiae.

Genetika, 1983 Jul, 19(7), 1063 - 9
{Molecular nature of direct gene mutations induced by gamma and ultraviolet irradiation in Saccharomyces cerevisiae yeasts}; Ivanov EL et al.; The spectrum of gamma- and UV-induced mutations for ADE2 locus of the yeast Saccharomyces cerevisiae was determined as follows (respectively): 27 and 41% GC leads to AT transitions, 8 and 11% AT leads to GC transitions, 59 and 40% transversions, 6 and 8% frameshifts . Our results indicate a specificity of UV-light for GC leads to AT transitions . Experimental data are discussed in a view of molecular mechanisms of radiation mutagenesis.

Arch Biochem Biophys, 1983 Jul 1, 224(1), 69 - 76
Reactivity of proteins in ribosomes from Saccharomyces cerevisiae with trypsin; Lee JC et al.; The susceptibility of 40S and 60S ribosomal subunits from Saccharomyces cerevisiae to digestion with varying concentrations of trypsin was studied by two-dimensional electrophoresis and quantitative measurements of the protein remaining with the ribosomal particles after trypsin treatment . Proteins from both subunits can be classified into three groups according to their rate of digestion by trypsin . These results are in good agreement with those obtained on the order of ribosomal assembly in vivo, i.e., proteins which are most susceptible to trypsin digestion have been shown to associate with the ribosomal particles at a relatively late stage of ribosome assembly.

J Bacteriol, 1983 Jul, 155(1), 64 - 8
Requirement for a second sterol biosynthetic mutation for viability of a sterol C-14 demethylation defect in Saccharomyces cerevisiae; Taylor FR et al.; Genetic analysis of a nystatin-resistant sterol mutant (strain JR4) of Saccharomyces cerevisiae defective in C-14 demethylation revealed the presence of a second mutation in 5,6-desaturation . It appeared from complementation tests that a defect in delta 5-desaturase enzyme activity was required for the viability of the C-14 demethylation mutant . Growth studies with a sterol auxotrophic strain indicated that the major sterol of strain JR4, 14 alpha-methyl-ergosta-8,24(28)-dien-3 beta-ol, could satisfy "bulk" membrane requirements but not the second, structurally specific, sterol function that we defined previously (Rodriguez et al., Biochem . Biophys . Res . Commun . 106:435-441, 1982) . Leakiness in the sterol mutations in strain JR4 provided a small amount of ergosterol which could satisfy this second function.

J Bacteriol, 1983 Jul, 155(1), 291 - 301
Expression of the BAR1 gene in Saccharomyces cerevisiae: induction by the alpha mating pheromone of an activity associated with a secreted protein; Manney TR; We have demonstrated and partially characterized the genetic control and pheromonal regulation of a soluble activity, produced only by mating-type a cells, that inhibits the action of the alpha mating pheromone, alpha-factor, on mating-type a cells . This activity was found to be associated with a heat-stable protein and to be secreted by MATa BAR1, mat alpha 2 BAR1, and mat alpha 1 mat alpha 2 BAR1 strains, but not by MAT alpha BAR1, MATa/MAT alpha BAR1, mat alpha 1 BAR1, or MATa barl strains, demonstrating that it is under the control of both the MAT alpha 2 and the BAR1 genes . Secretion of this activity was also found to be stimulated to as much as five times the basal level by exposure of the cells to alpha-factor . This stimulation was maximal after 6 h at a pheromone concentration of approximately 2 U/ml . An assay for this activity was developed by using a refined, quantitative assay for alpha-factor . The pheromone activity of samples added to wells in an agar plate was related to the size of the halo of growth inhibition produced in a lawn of mutant cells that are abnormally sensitive . The alpha-factor-inhibiting activity was related to a reduction of the halo size when active samples were added to the lawn . Although the assay for alpha-factor was found to be relatively insensitive to pH over a range of several units, the alpha-factor-inhibiting activity displayed a sharp pH optimum at approximately 6.5 . The properties of this activity have important implications concerning the role of the BAR1 gene product in recovery of mating-type a cells from cell division arrest by alpha-factor.

J Bacteriol, 1983 Jul, 155(1), 8 - 14
Nucleotide sequence of the tcml gene (ribosomal protein L3) of Saccharomyces cerevisiae; Schultz LD et al.; The yeast tcml gene, which codes for ribosomal protein L3, has been isolated by using recombinant DNA and genetic complementation . The DNA fragment carrying this gene has been subcloned and we have determined its DNA sequence . The 20 amino acid residues at the amino terminus as inferred from the nucleotide sequence agreed exactly with the amino acid sequence data . The amino acid composition of the encoded protein agreed with that determined for purified ribosomal protein L3 . Codon usage in the tcml gene was strongly biased in the direction found for several other abundant Saccharomyces cerevisiae proteins . The tcml gene has no introns, which appears to be atypical of ribosomal protein structural genes.

Nucleic Acids Res, 1983 Jun 25, 11(12), 4049 - 63
Saccharomyces cerevisiae contains two discrete genes coding for the alpha-factor pheromone; Singh A et al.; Two genes, MF alpha 1 and MF alpha 2, coding for the alpha-factor in yeast Saccharomyces cerevisiae were identified by in situ colony hybridization of synthetic probes to a yeast genomic library . The probes were designed on the basis of the known amino acid sequence of the tridecapeptide alpha-pheromone . The nucleotide sequence revealed that the two genes, though similar in their overall structure, differ from each other in several striking ways . MF alpha 1 gene contains 4 copies of the coding sequence for the alpha-factor, which are separated by 24 nucleotides encoding the octapeptide Lys-Arg-Glu-Ala-Glu(or Asp)-Ala-Glu-Ala . The first alpha-factor coding block is preceded by a sequence for the hexapeptide Lys-Arg-Glu-Ala and 83 additional amino acids . MF alpha 2 gene contains coding sequences for two copies of the alpha-factor that differ from each other and from alpha-factor encoded by MF alpha 1 gene by a Gln leads to Asn and a Lys leads to Arg substitution . The first copy of the alpha-factor is preceded by a sequence coding for 87 amino acids which ends with Lys-Arg-Glu-Ala-Val-Ala-Asp-Ala . The coding blocks of the two copies of the pheromone are separated by the sequence for Lys-Arg-Glu-Ala-Asn-Ala-Asp-Ala . Thus, the alpha-factor can be derived from 2 different precursor proteins of 165 and 120 amino acids containing, respectively, 4 and 2 copies of the pheromone.

Biochim Biophys Acta, 1983 Jun 10, 731(2), 361 - 72
Amino acid uptake by yeasts . IV . Effect of thiol reagents on L-leucine transport in Saccharomyces cerevisiae; Ramos EH et al.; (1) N-Ethylmaleimide (a penetrating SH- reagent) inactivated L-{14C}leucine entrance (binding and translocation) into Saccharomyces cerevisiae, the extent of inhibition depending on the time of preincubation with N-ethylmaleimide, N-ethylmaleimide concentration, the amino acid external and internal concentration, and the energization state of the yeast cells . With D-glucose-energized yeast, N-ethylmaleimide inhibited L-{14C}leucine entrance in all the assayed experimental conditions, but with starved yeast and low (0.1 mM) amino acid concentration, it did not inhibit L-{14C}leucine binding, except when the cells were preincubated with L-leucine . With the rho- respiratory-deficient mutant (energized cells), N-ethylmaleimide inhibited L-{14C}leucine entrance as with the energized wild-type, though to a lesser extent . (2) Analysis of the N-ethylmaleimide effect as a function of L-{14C}leucine concentration showed a significant decrease of Jmax values of the high- (S1) and low- (S2) affinity amino acid transport systems, but KT values were not significantly modified . (3) When assayed in the presence of D-glucose, N-ethylmaleimide inhibition of D-glucose uptake and respiration contributed significantly to inactivation of L-{14C}leucine entrance . Pretreatment of yeast cells with 2,4-dinitrophenol enhanced the effect of L-{14C}leucine binding and translocation . (4) Bromoacetylsulfanilic acid and bromoacetylaminoisophthalic acid, two non-penetrating SH- reagents, did not inactivate L-{14C}leucine entrance, while p-chloromercuribenzoate, a slowly penetrating SH-reagent, inactivated it to a limited extent . When compared with the effect of N-ethylmaleimide, these negative results indicate that thiol groups of the L-{14C}leucine carrier were not exposed on the outer surface of the yeast cell permeability barrier.

Biochim Biophys Acta, 1983 Jun 9, 757(3), 342 - 51
Studies on compartmentation of S-adenosyl-L-methionine in Saccharomyces cerevisiae and isolated rat hepatocytes; Farooqui JZ et al.; The existence of metabolically distinct pools of S-adenosyl-L-methionine in Saccharomyces cerevisiae and isolated rat hepatocytes was investigated . Utilizing a relatively long labeling period with {methyl-14C}methionine, a metabolically 'stable' pool was labeled . A subsequent short labeling with {methyl-3H}methionine selectively labeled a putative metabolically 'labile' pool . The existence of these distinguishable pools was ascertained by following the 3H and 14C label disappearance in S-adenosyl-L-methionine during the chase-period in label-free media containing cycloleucine to prevent further synthesis of S-adenosyl-L-methionine . In both yeast and hepatocytes, the 3H/14C ratio in S-adenosyl-L-methionine decreased sharply . The individual 3H and 14C decrease in S-adenosyl-L-methionine showed t1/2 values of 3 and 8 min for yeast and 4 and 18 min for hepatocytes . The results strongly indicate that at least two metabolically distinct S-adenosyl-L-methionine pools actually do exist in both systems . Subcellular fractionation revealed that the 'labile' pool exist in the cytosol for both yeast and hepatocytes while the 'stable' pool exists in the vacuolar and the mitochondrial fraction for the yeast and hepatocytes respectively . The S-adenosyl-L-methionine pools were also studied in normal yeast under anaerobic chase condition and petite mutant yeast . Sharply contrasting with aerobically chased normal yeast, both showed closely parallel 3H and 14C decreases in S-adenosyl-L-methionine.

Biochim Biophys Acta, 1983 Jun 2, 762(3), 466 - 70
NMR analysis of a cell division cycle mutant of Saccharomyces cerevisiae; Gillies RJ et al.; cdc 19.1 is a temperature-sensitive lesion in the genome of Saccharomyces cerevisiae . The phenotype of this mutant is a cell cycle specific arrest in G1, which is expressed at 37 degrees C . In the present study, 31P- and 13C-NMR spectroscopy were used to analyze the metabolism of the mutant at the permissive and restrictive temperatures . Our results confirm previous findings which have indicated that cdc 19.1 contains temperature-sensitive pyruvate kinase activity . In contrast to previous findings, however, the present investigation demonstrates that restriction of pyruvate kinase activity in vivo takes as long as 24 h to be fully expressed . In addition, analysis by NMR has allowed us to assess the metabolic consequences of pyruvate kinase restriction which may contribute to the arrest of cell growth in the early G1 phase of the cell division cycle.

Genetika, 1983 Jun, 19(6), 921 - 6
{Molecular nature of the mutations in gene ADE2 in Saccharomyces cerevisiae yeasts . III . Determination of the correlation of the GC AT pairs in genes ADE1 and ADE2}; Korolev VG; Lethal and mutagenic effects and the nature of mutations induced by decay of 32P incorporated into yeast cell DNA as 32P-deoxyguanosine monophosphate (32PdGMP) and 32P-thymidine monophosphate (32P-TMP), were studied . The lethal efficiency per 32P decay is independent of a labelled nucleotide incorporated into DNA . However, the mutagenic efficiency in ADE1, ADE2 genes per 32P decay is approximately 3 times greater for 32PdGMP than for 32P-TMP . This suggests that ADE1, ADE2 genes contain about 3 times more GC base pairs than AT pairs . Variations in a relative frequencies of GC leads to AT and AT leads to GC transitions were obtained depending upon a nucleotide labelled.

Can J Microbiol, 1983 Jun, 29(6), 681 - 8
Studies on mutants affecting amidophosphoribosyltransferase activity in Saccharomyces cerevisiae; Nieto DJ et al.; Mutants at the ade4 locus of yeast were isolated following mutagenesis of ade+ and ade2 with ultraviolet light (UV), ethylmethane sulphonate, and the acridine half mustard ICR-170 . Tests for interallelic complementation, osmotic remediality, temperature sensitivity, and mutagen-specific reversion were carried out on 19 mutants . Six mutants showed interallelic complementation and fell into four groups, defining three complons . Three mutants were osmotic remedial and the same three were temperature sensitive . Three mutants induced by ICR-170 gave purine-excreting revertants, designated Pur6 or ade4.RCF, after exposure to UV . Activity of amidophosphoribosyltransferase (PRPPAT) was assayed in the ade4 mutants and other alleles at this locus . The ade4 mutants lacked activity of the enzyme; the alleles su-pur+, su-pur, PUR6, and Pur6, showed different levels of activity . The enzyme was subject to feedback inhibition by AMP and IMP in su-pur+ and PUR6; su-pur was hypersensitive to inhibition by AMP, whereas Pur6 was slightly resistant . Purine synthesis de novo was shown to be repressible in su-pur+ and constitutive in PUR6 and Pur6 by following the accumulation of aminoimidazole ribotide in the presence and absence of cycloheximide . These observations were confirmed by direct assay of enzyme activity.

Mol Cell Biol, 1983 Jun, 3(6), 1000 - 12
Saccharomyces cerevisiae cdc2 mutants fail to replicate approximately one-third of their nuclear genome; Conrad MN et al.; Chromosomal DNA replication was examined in temperature-sensitive mutants of Saccharomyces cerevisiae defective in a gene required for the completion of S phase at the nonpermissive temperature, 37 degrees C . Based on incorporation of radioactive precursors and density transfer experiments, strains carrying three different alleles of cdc2 failed to replicate approximately one-third of their nuclear genome at 37 degrees C . Whole-cell autoradiography experiments demonstrated that 93 to 96% of the cells synthesized DNA at 37 degrees C . Therefore, all cells failed to replicate part of their genome . DNA isolated from terminally arrested cells was of normal size as measured on neutral and alkaline sucrose gradients, suggesting that partially replicated DNA molecules do not accumulate and that DNA strands are ligated properly in cdc2 mutants . In addition, electron microscopic examination of the equivalent of more than one genome's DNA from arrested cells failed to reveal any partially replicated molecules . The sequences which failed to replicate at 37 degrees C were not highly specific; eight different cloned sequences replicated to the same extent as total DNA . The 2-microns plasmid DNA and rDNA replicated significantly less well than total DNA, but approximately one-half of these sequences replicated at 37 degrees C . These observations suggest that cdc2 mutants are defective in an aspect of initiation of DNA replication common to all chromosomes such that a random fraction of the chromosomes fail to initiate replication at 37 degrees C, but that once initiated, replication proceeds normally.

Genetics, 1983 Jun, 104(2), 219 - 34
A mutation allowing expression of normally silent a mating-type information in Saccharomyces cerevisiae; Gruenspan H et al.; Mating type in haploid cells of the yeast Saccharomyces cerevisiae is determined by a pair of alleles MATa and MAT alpha . Under various conditions haploid mating types can be interconverted . It has been proposed that transpositions of silent cassettes of mating-type information from HML OR HMR to MAT are the source of mating type conversions . A mutation described in this work, designated AON1, has the following properties . (1) MAT alpha cells carring AON1 are defective in mating . (2) AON1 allows MAT alpha/MAT alpha but not MATa/MATa diploids to sporulate; thus, AON1 mimics the MATa requirement for sporulation . (3) mata-1 cells that carry AON1 are MATa phenocopies, i.e., MAT alpha/mata-1 AON1 diploids behave as standard MAT alpha/MATa cells; therefore, AON1 suppresses the defect of mata-1 . (4) AON1 maps at or near HMRa . (5) Same-site revertants from AON1 lose the ability to convert mating type to MATa, indicating that reversion is associated with the loss of a functional HMRa locus . In addition, AON1 is a dominant mutation . We conclude that AON1 is a regulatory mutation, probably cis-acting, that leads to the constitutive expression of silent a mating-type information located at HMRa.

Radiat Res, 1983 Jun, 94(3), 464 - 79
Differential inactivation analysis of diploid yeast exposed to radiation of various LET . I . Computerized single-cell observation and preliminary application to X-ray-treated Saccharomyces cerevisiae; Grundler W et al.; This series of investigations was designed to observe growth and division of single, diploid yeast cells within the first four generations after irradiation with ionizing radiation . Evidence exists that cell reactions important for the final cell fate occur during this period, and therefore the analysis of cell kinetics and of stationary forms of inactivated cells can be performed . A large number of experiments is necessary to obtain statistically confirmed results of single-cell observation . An automatically steered microphotographic registration device has been developed to facilitate the collection of large numbers of observations . Optical data scanned by a TV camera and digitally stored in a computer are processed by pattern recognizing programs to achieve the correct correlation of newly built cells to existing ones and to deliver a pedigree over four generations of at least eight cells for every irradiated single cell . The pooled data of many pedigrees of this kind allow the analysis of the differential behavior of a total population . From the analysis of X-irradiated cells one can conclude that a single cell that produces at least a microcolony of five cells is eventually able to form a microcolony and thus can be considered a survivor . That means the division probability of cells to go from generation zero to three corresponds to the survival curve of the colony-forming ability test . Therefore this method is suitable for the differential description of the important phenomenological cell reactions after irradiation.

J Cell Biol, 1983 Jun, 96(6), 1592 - 600
Two temperature-sensitive mutants of Saccharomyces cerevisiae with altered expression of mating-type functions; Manney TR et al.; Two mutants of Saccharomyces cerevisiae have been isolated from normal haploid MAT alpha strains and characterized as having temperature-sensitive, pleiotropic phenotypes for functions associated with mating . At the permissive temperature, 23 degrees C, they were found to behave as normal MAT alpha haploids with respect to mating efficiency, sporulation in diploids formed with MAT a strains, secretion of alpha-factor, and failure to secrete the MATa-specific products, a-factor and Barrier . At higher temperatures they were found to decline in mating and sporulation efficiency and to express the a-specific functions . Genetic analysis established that one of these mutants, PE34, carries a temperature-sensitive allele of the MAT alpha 2 gene and that the other, PD7, carries a temperature-sensitive allele of the TUP1 gene.

J Bacteriol, 1983 Jun, 154(3), 1476 - 9
Nonsense mutations in the can1 locus of Saccharomyces cerevisiae; Ono BI et al.; Yeast mutants resistant to L-canavanine were selected . All were recessive and fell into the can1 complementation group . Nonsense mutations were identified among them by using a set of different suppressors . Frequencies of UAA, UAG, and presumed UGA mutations were 14.8, 0.8, and 0.4%, respectively . A high incidence of nonsense mutations having discriminatory suppression patterns was characteristic of the locus.

J Bacteriol, 1983 Jun, 154(3), 1276 - 83
Plasma membrane expansion terminates in Saccharomyces cerevisiae secretion-defective mutants while phospholipid synthesis continues; Ramirez RM et al.; Phospholipid synthesis activity and plasma membrane growth have been studied in the Saccharomyces cerevisiae temperature-sensitive, secretion-defective mutants isolated by Novick and Schekman (Proc . Natl . Acad . Sci . U.S.A . 76:1858-1862, 1979; Novick et al., Cell 21:205-215, 1980) . The mutants, sec1 through sec23, do not grow at 37 degrees C and exhibit lower rates of phospholipid synthesis than does the wild-type strain X2180 . None of the mutants exhibits a decline in lipid synthesis rapid enough to explain secretion failure . Plasma membrane growth was assessed indirectly by examining the osmotic sensitivity of spheroplasts derived from cultures transferred from 24 to 37 degrees C . Spheroplasts from the normal-growing strain X2180 exhibited a small rapid increase in osmotic sensitivity and stabilized at a more sensitive state . Spheroplasts from the sec mutants exposed to the same temperature shift exhibited progressively increasing osmotic sensitivity . Cycloheximide treatment prevented progressive increases in osmotic fragility . These data are compatible with the hypothesis that plasma membrane expansion is restricted in the sec mutants . During incubation at 37 degrees C, the accumulation of intracellular materials within the no-longer expanding plasma membrane exerts osmotic stress on the membrane, increasing with time . The gene products defective in Novick and Schekman's sec mutants appear to be required for both extracellular protein secretion and plasma membrane growth in yeast cells.

Eur J Biochem, 1983 Jun 1, 133(1), 141 - 4
Study of the positive control of the general amino-acid permease and other ammonia-sensitive uptake systems by the product of the NPR1 gene in the yeast Saccharomyces cerevisiae; Grenson M; Mutations at the NPR1 genetic locus are known to inactivate (totally or partially) at least five distinct ammonia-sensitive permeases . Mutants with thermosensitive NPR1 gene product (nprts) have been used to discriminate between three possible roles of this protein, namely (a) a common constituent of a set of ammonia-sensitive permeases; (b) a common activator of these permeases; (c) a common positive factor necessary for their synthesis . Inactivation of the general amino-acid permease was observed upon transfer of nprts mutant cells to a non-permissive temperature . Under the same conditions, the general amino-acid permease of the wild-type cells remained active for several hours even when protein synthesis was inhibited by nitrogen starvation or by cycloheximide . Mutations at three unlinked loci, namely the PGR site (located in the GAP1 structural gene of the permease), and the unlinked MUT2 and MUT4 loci restore the general amino-acid permease activity in npr1 mutants . The results are interpreted as indicating that the NPR1 product is necessary for the reactivation of the general amino-acid permease which seems to be continuously inactivated by a regulatory process mediated by the MUT2 and the MUT4 gene products acting at the level of the PGR site of the general amino-acid permease molecule . The proline permease and the ureidosuccinic-acid permease seem to be subject to the same double regulation by inactivation-reactivation of the permeases and by repression of their synthesis . A tentative scheme of the regulation of the general amino-acid permease is presented.

Eur J Biochem, 1983 Jun 1, 133(1), 135 - 9
Inactivation-reactivation process and repression of permease formation regulate several ammonia-sensitive permeases in the yeast Saccharomyces cerevisiae; Grenson M; Two distinct regulatory mechanisms are responsible for the absence of general amino-acid permease activity in cells of the wild-type strain sigma 1278b of Saccharomyces cerevisiae grown in the presence of ammonium ions . One is a reversible inactivation process which progressively develops upon addition of ammonium ions to a proline-grown culture, and completely suppresses the permease activity within one hour . This inactivation process is absent in mutants altered at the MUT2, MUT4, or PGR genetic loci . In these mutants, a repression of the formation of active permease may clearly be observed in the presence of ammonium ions . This second regulatory mechanism is absent in mutants affected at the GDHCR locus, which might code for a repressor molecule . It is also relieved in the presence of a glnts mutation (which makes the glutamine synthetase thermosensitive) suggesting glutamine as an effector . Two other ammonia-sensitive permeases, namely the proline permease and the ureidosuccinic-acid permease, seem to be subject to the same double regulation . Mutations affecting the structural gene of the anabolic NADP-linked glutamate dehydrogenase (gdhA) seem to completely prevent repression of the general amino-acid permease, while they partially suppress its inactivation in the presence of ammonium ions.

Exp Cell Res, 1983 Jun, 146(1), 151 - 61
Control of cell division in Saccharomyces cerevisiae mutants defective in adenylate cyclase and cAMP-dependent protein kinase; Matsumoto K et al.; Examination of the proportion of unbudded cells, terminal nuclear phenotype and DNA content of nuclei indicated that cyr1 mutants of yeast defective in adenylate cyclase activity were arrested at the G1 phase of the cell cycle . The step of G1 arrest due to the cyr1 mutation preceded the step sensitive to the mating pheromone . The temperature-sensitive cyr1 cells did not continue growth, nor retain the capacity to conjugate at a restrictive temperature . The phenotypes of the cyr1 mutant mimicked those of nutritionally limited cells . The G1 arrest caused by the cyr1 mutation was overcome by the presence of a suppressor mutation, bcy1, that resulted in deficiency of a regulatory subunit of cAMP-dependent protein kinase and production of high level of cAMP-independent protein kinase . The bcy1 mutation suppressed G1 arrest caused by nutritional limitation, and continued bud emergence for multiple cycles without further nuclear division . The data suggest that cAMP works as a positive effector at the start of a yeast cell cycle via activation of cAMP-dependent protein kinase.

J Biol Chem, 1983 May 25, 258(10), 6293 - 9
Structure and expression of two aminoacyl-tRNA synthetase genes from Saccharomyces cerevisiae; Meussdoerffer F et al.; We have isolated the structural genes for methionyl-tRNA synthetase (L-methionine:tRNAMet ligase, EC 6.1.1.10) and isoleucyl-tRNA synthetase (L-isoleucine:tRNAIle ligase, EC 6.1.1.5) from the yeast Saccharomyces cerevisiae by transformation . Both genes exist in a single copy in the yeast genome and show no homology to either yeast mitochondrial or Escherichia coli DNA as judged by Southern analysis . The region corresponding to the DNA segment altered in mes1 or ils1-1 mutants was defined by transformation of synthetase mutants with subclones of the respective gene . Although the isolated fragment containing the methionyl-tRNA synthetase gene complements the mes1 mutation in yeast, it does not complement the analogous metG mutation in E . coli . A single transcript of about 3100 nucleotides codes for the isoleucyl-tRNA synthetase . This transcript is about 2-fold elevated in strains starved for isoleucine, histidine, or arginine as compared to wild type . It is regulated by the general control of amino acid biosynthesis, which affects several biosynthetic pathways . Two polyadenylated transcripts are homologous to the methionyl-tRNA synthetase gene . The major MES1 transcript is not affected by the general control or starvation for several different amino acids, including methionine.

Biochim Biophys Acta, 1983 May 20, 740(1), 88 - 98
A petite mitochondrial DNA segment arising in exceptionally high frequency in a mit- mutant of Saccharomyces cerevisiae; Bingham CG et al.; In cultures of the mit- mutant strain Mb12 of Saccharomyces cerevisiae (carrying a mutation in the oli2 gene), 70% of the cells are petite mutants . More than 80% of the petites from Mb12 contain a particular mtDNA segment, denoted BB5, that is 880 bp long and carries a single MboI site . Thus, in cultures of Mb12, about 56% of the cells are petites containing the defective BB5 mtDNA genome, and only 30% are mit- cells containing parental Mb12 mtDNA . The BB5 mtDNA segment is also found in petites arising from the wild-type strain J69-1B (from which Mb12 was derived), but in this case mtDNA from only five out of 24 petites produced an 880 bp band after MboI digestion . Since J69-1B cultures carry a petite frequency of about 5%, approximately 1% of cells in J69-1B cultures contain the BB5 mtDNA segment . The difference between Mb12 and J69-1B cultures is reflected in the MboI digestion patterns of the respective mtDNAs . While Mb12 mtDNA contains a grossly superstoicheiometric 880 bp MboI fragment, the corresponding fragment in J69-1B mtDNA cannot be seen on stained gels, but can be readily visualized in Southern blots hybridized to a 32P-labelled DNA probe obtained from the 880 bp MboI fragment . The BB5 mtDNA segment was shown to contain the ori1 sequence (one of several very similar sequences in wild-type mtDNA thought to act as origins of replication of mtDNA) which confers the genetic property of very high suppressiveness on petites carrying this mtDNA . The efficient replication of BB5 mtDNA may contribute to its abundance in Mb12 cultures . Nevertheless, other factors must operate to influence the abundance of the BB5 mtDNA segment in cultures of different strains, the most important of which is likely to be the rate of excision of this mtDNA segment from the parental mtDNA genome.

J Biol Chem, 1983 May 10, 258(9), 5614 - 7
Calcium transport driven by a proton motive force in vacuolar membrane vesicles of Saccharomyces cerevisiae; Ohsumi Y et al.; Vacuolar membrane vesicles of Saccharomyces cerevisiae accumulate Ca2+ ion in the presence of ATP, not in the presence of ADP or adenyl-5'-yl imidodiphosphate . Calcium transport showed saturation kinetics with a Km value of 0.1 mM and optimal pH of 6.4 . Ca2+ ion incorporated in the vesicles was exchangeable and released completely by a protonophore uncoupler, 3,5-di-tert-butyl-4-hydroxybenzilidenemalononitrile (SF6847), or calcium-specific ionophore, A23187 . The transport required Mg2+ ion but was inhibited by Cu2+ or Zn2+ ions, inhibitors of H+-ATPase of the vacuolar membrane . The transport activity was sensitive to the H+-ATPase inhibitor N,N'-dicyclohexylcarbodiimide, but not to oligomycin or sodium vanadate . SF6847 or nigericin blocked Ca2+ uptake completely, but valinomycin stimulated it 1.35-fold . These results indicate that an electrochemical potential difference of protons is a driving force for this Ca2+ transport . The ATP-dependent formation of the deltapH in the vesicles and its partial dissipation by CaCl2 were demonstrated by fluorescence quenching of quinacrine . This Ca2+ uptake by vacuolar membrane vesicles is suggested to be catalyzed by a Ca2+/H+ antiport system.

FEBS Lett, 1983 May 8, 155(2), 225 - 9
Ty1 extrachromosomal circular copies in Saccharomyces cerevisiae; Ballario P et al.; The eukaryotic transposable element Ty1 is present in about 20-30 integrated copies per yeast aploid genome, variably localized in different strains . Here, we report the presence in yeast of circular extrachromosomal molecules homologous to Ty1, 6 kilobases in size (the same as integrated copies) present in about 1 circular copy/250-300 cells . This finding shows another analogy between eukaryotic-transposable elements and the pro-viral integrative form of retroviruses.

Eur J Biochem, 1983 May 2, 132(2), 235 - 40
Mitochondrial polypeptide elongation factor EF-Tu of Saccharomyces cerevisiae . Functional and structural homologies to Escherichia coli EF-Tu; Piechulla B et al.; The polypeptide elongation factor EF-Tu was isolated from a mitochondrial 100 000 x g supernatant of the yeast Saccharomyces cerevisiae and purified over 880-fold by DEAE-Sephadex chromatography and gel filtration . The factor efficiently replaces bacterial EF-Tu in a phenylalanine polymerizing cell-free system of Escherichia coli, it binds GDP and it protects phenylalanyl-tRNA against hydrolysis of the ester bond in the presence of 10 mM GTP . The polymerizing activity of the mitochondrial factor is inhibited to 90% by 50 microM N-ethylmaleimide and to 50% by 2.5 microM kirromycin . The purified factor contains two major polypeptides of apparent molecular weights 48 000 and 34 000 . Antibodies raised against the 48 000-Mr protein react with EF-TuE . coli, as revealed by immune blotting and by the inhibition of phenylalanine polymerization . No reaction was observed between anti-(34 000-Mr) and 48 000-Mr protein or EF-TuE . coli . The 48 000-Mr protein has the same isoelectric point (pI = 6.2) and a content of cysteine and basic amino acids similar to the bacterial EF-Tu . It is concluded that the 48 000-Mr protein is the analogue to EF-TuE . coli, and that yeast mitochondrial EF-Tu is functionally and structurally more related to bacterial EF-Tu than cytosolic EF-1 of the same cell.

J Bacteriol, 1983 May, 154(2), 1013 - 4
Effect of carbon source on lysine-mediated inhibition of postexponential growth of Saccharomyces cerevisiae; Watson TG; Lysine-mediated inhibition of postexponential growth in Saccharomyces cerevisiae occurred when glucose, fructose, or maltose, but not lactate, pyruvate, or ethanol, was used as the carbon source . Arginine starvation is not responsible for the inhibitory effect, since neither the intracellular pool of glucose-grown (inhibited) cells nor that of lactate-grown (noninhibited) cells contained arginine.

Cell, 1983 May, 33(1), 203 - 10
Genes that act before conjugation to prepare the Saccharomyces cerevisiae nucleus for caryogamy; Dutcher SK et al.; Mutations in four nuclear genes, kar1 cdc4, 28, and 37, block or impair nuclear fusion during conjugation of Saccharomyces cerevisiae . Mutations in all four genes are recessive for the caryogamy defect; in matings between diploid cells both of which are heterozygous for any one of the four mutations (-/+ X -/+), caryogamy occurs with normal proficiency . However, mutations in all four genes are "nuclear dominant"; that is, both parent nuclei must contribute one wild-type allele of each gene for successful caryogamy . In order to discriminate between two possible models to explain nuclear dominance, we have examined the caryogamy proficiency of mutant nuclei after they had passed through a heterocaryotic cytoplasm . The kar1, cdc28, and cdc37 caryogamy defects are all phenotypically suppressed in this experiment (cdc4 could not be tested) . We conclude from our results that the KAR1, CDC28, and CDC37 gene products can diffuse between nuclei in a heterocaryon and that they probably perform their function for caryogamy prior to cell fusion . One simple model consistent with the roles of CDC28 and CDC37 in mitosis as well as in caryogamy is that these gene products are structural components of the nucleus that must be built into it during one cell cycle in order to permit successful caryogamy at the next G1.

J Gen Microbiol, 1983 May, 129 (Pt 5), 1589 - 91
Alpha-factor enhancement of hybrid formation by protoplast fusion in the yeast Saccharomyces cerevisiae; Curran BP et al.; Two a strains of Saccharomyces cerevisiae, carrying complementary genetic markers, were arrested for 3 h with alpha-factor . These were then protoplasted, prior to fusion using polyethylene glycol . The number of viable fusion products was enhanced by a factor of 20 as compared with unarrested controls.

Mol Cell Biol, 1983 May, 3(5), 871 - 80
SAD mutation of Saccharomyces cerevisiae is an extra a cassette; Kassir Y et al.; Sporulation of Saccharomyces cerevisiae ordinarily requires the a1 function of the a mating type locus . SAD is a dominant mutation that allows strains lacking a1 (MAT alpha/MAT alpha and mata1/MAT alpha diploids) to sporulate . We provide functional and physical evidence that SAD is an extra cassette in the yeast genome, distinct from those at HML, MAT, and HMR . The properties of SAD strains indicate that the a cassette at SAD produces a limited amount of a1 product, sufficient for promoting sporulation but not for inhibiting mating and other processes . These conclusions come from the following observations . (i) SAD did not act by allowing expression of HMRa: mata1/MAT alpha diploids carrying SAD and only alpha cassettes at HML and HMR sporulated efficiently . (ii) SAD acted as an a cassette donor in HML alpha HMR alpha strains and could heal a mata1 mutation to MATa as a result of mating type interconversion . (iii) The genome of SAD strains contained a single new cassette locus, as determined by Southern hybridization . (iv) Expression of a functions from the SAD a cassette was limited by Sir: sir- SAD strains exhibited more extreme phenotypes than SIR SAD strains . This observation indicates that SAD contains not only cassette information coding for a1 (presumably from HMRa) but also sites for Sir action.

Mol Cell Biol, 1983 May, 3(5), 803 - 10
Efficient production of a ring derivative of chromosome III by the mating-type switching mechanism in Saccharomyces cerevisiae; Klar AJ et al.; The mating-type switches in the yeast Saccharomyces cerevisiae occur by unidirectional transposition of replicas of unexpressed genetic information, residing at HML or HMR, into the mating-type locus (MAT) . The source loci, HML and HMR, remain unchanged . Interestingly, when the HM cassettes are expressed, as in marl strains, the HML and HMR cassettes can also efficiently switch, apparently by obtaining genetic information from either of the other two cassettes (Klar et al., Cell 25:517-524, 1981) . We have isolated a novel chromosome III rearrangement in heterothallic (marl ho) strains, which is also produced efficiently in marl HO cells, presumably the consequence of a recombination event between HML and HMR . The fusion results in the loss of sequences which are located distal to HML and to HMR and produces a ring derivative of chromosome III . Cells containing such a ring chromosome are viable as haploids; apparently, no essential loci are located distal to the HM loci . The fusion cassette behaves as a standard HM locus with respect to both regulation by the MAR/SIR control and its role in switching MAT.

Proc Natl Acad Sci U S A, 1983 May, 80(10), 2824 - 8
In vitro nonsense suppression in {psi+} and {psi-} cell-free lysates of Saccharomyces cerevisiae; Tuite MF et al.; An homologous in vitro assay for yeast nonsense suppressors was used to examine the effect of the cytoplasmically inherited genetic determinant {psi} on the efficiency of in vitro nonsense suppression . The efficiency of all three types of yeast tRNA-mediated nonsense suppressor (ochre, amber, and UGA) is much greater in cell-free lysates prepared from a sup+ {psi+} strain than in lysates prepared from an isogeneic sup+ {psi-} strain . Lysates prepared from a {psi-} strain, into which the {psi+} determinant was reintroduced by kar1-mediated cytoduction, support efficient suppression . Evidence is also presented that {psi-} lysates contain an inhibitor of in vitro nonsense suppression.

Proc Natl Acad Sci U S A, 1983 May, 80(9), 2437 - 41
Primary structure of the Saccharomyces cerevisiae gene for methionyl-tRNA synthetase; Walter P et al.; The sequence of a 5-kilobase DNA insert containing the structural gene for yeast cytoplasmic methionyl-tRNA synthetase has been determined and a unique open reading frame of 2,253 nucleotides encoding a polypeptide chain of 751 amino acids (Mr, 85,500) has been characterized . The data obtained on the purified enzyme (subunit size, amino acid composition, and COOH-terminal sequence) are consistent with the gene structure . The protein sequence deduced from the nucleotide sequence reveals no obvious internal repeats . This protein sequence shows a high degree of homology with that of Escherichia coli methionyl-tRNA synthetase within a region that forms the putative methionyl adenylate binding site . This strongly suggests that both proteins derive from a common ancestor.

J Bacteriol, 1983 May, 154(2), 612 - 22
Maintenance of the 2 microns circle plasmid in populations of Saccharomyces cerevisiae; Futcher AB et al.; The 2 microns circle plasmid is maintained at high frequencies in populations of yeast cells . To find out how the plasmid is maintained, three forces were measured: the selective advantage or disadvantage conferred by 2 microns circles, the rate of generation of {Cir0} cells, and the rate of illegitimate transfer of 2 microns circles from cell to cell . It was found that under the conditions used, 2 microns circles confer a selective disadvantage of about 1%, that {Cir0} cells are generated at the rate of 7.6 x 10(-5) per {Cir+} cell per generation, and that illegitimate transfer of 2 microns circles occurs at a rate less than 10(-7) per recipient cell per generation . The most likely explanation of 2 microns circle maintenance is that the plasmid is sexually transmitted at such a rate that it spreads through populations despite selection against it.

Mol Cell Biol, 1983 May, 3(5), 922 - 30
Chitin synthesis and localization in cell division cycle mutants of Saccharomyces cerevisiae; Roberts RL et al.; Growth of Saccharomyces cerevisiae cell cycle mutants cdc3, cdc4, cdc7, cdc24, and cdc28 at a nonpermissive temperature (37 degrees C) resulted in increased accumulation of chitin relative to other cell wall components, as compared with that observed at a permissive temperature (25 degrees C) . Wild-type cells showed the same chitin/carbohydrate ratio at both temperatures, whereas mutants cdc13 and cdc21 yielded only a small increase in the ratio at 37 degrees C . These results confirm and extend those reported by B . F . Sloat and J . R . Pringle (Science 200:1171-1173, 1978) for mutant cdc24 . The distribution of chitin in the cell wall was studied by electron microscopy, by specific staining with wheat germ agglutinin-colloidal gold complexes . At the permissive temperature, chitin was restricted to the septal region in all strains, whereas at 37 degrees C a generalized distribution of chitin in the cell wall was observed in all mutants . These results do not support a unique interdependence between the product of the cdc24 gene and localization of chitin deposition; they suggest that unbalanced conditions created in the cell by arresting the cycle at different stages result in generalized activation of the chitin synthetase zymogen . Thus, blockage of an event in the cell cycle may lead to consequences that are not functionally related to that event under normal conditions.

J Biol Chem, 1983 Apr 25, 258(8), 4754 - 8
Biosynthetic labeling of diphthamide in Saccharomyces cerevisiae; Dunlop PC et al.; Diphthamide, the post-translational amino acid derivative in the diphtheria toxin-modification site of protein synthesis elongation factor 2 (EF-2), has the proposed structure 2-{3-carboxyamido-3-(trimethylammonio)propyl}histidine (Van Ness, B . G., Howard, J . B., and Bodley, J . W . (1980) J . Biol . Chem . 255, 10710-10716) . The identification of the biosynthetic precursors of diphthamide would provide a means of evaluating its proposed structure and determining if the amino acid occurs in proteins other than EF-2 . Toward this end, yeast were grown on potential radiolabeled precursors and the resulting radiolabeled protein was hydrolyzed in acid . The acid hydrolysates were subjected to amino acid analysis with a program optimized to resolve diphthine, the acid hydrolysis product of diphthamide, from the common amino acids . Radiolabel from {beta-3H}histidine, {alpha-3H}methionine, and {Me-3H} methionine was found to be incorporated into diphthine in a molar ratio of 1:1:3 while that of {35S}methionine was not incorporated . These results are in accord with the proposed structure of diphthamide and suggest that in its biosynthesis the backbone and 3 methyl groups of methionine are added to a histidine residue in the peptide chain of EF-2 . These labeling experiments show that diphthine (diphthamide) constitutes approximately 6 X 10(-6) mol fraction of the total amino acids in yeast protein hydrolysates . Estimates of the amount of diphthamide present in the diphtheria toxin-modification site of EF-2 indicate that it constitutes from 4.5 to 9 X 10(-6) mol fraction of the total amino acids in yeast protein . The present evidence suggests that diphthamide occurs only in EF-2.

Nucleic Acids Res, 1983 Apr 25, 11(8), 2287 - 302
Expression in Saccharomyces cerevisiae of human interferon-alpha directed by the TRP1 5' region; Dobson MJ et al.; The complete 5' flanking region of the yeast TRP1 gene encoding N-(5'-phosphoribosyl)- anthranilate isomerase, a nonabundant protein, has been cloned and the nucleotide sequence data has been extended from -102 to -440 . The CT block--CAAG structure common to all efficiently expressed yeast genes is altered in the 5' region of TRP1 and a sequence postulated to be involved in general amino acid regulation is absent . There are two possible TATA boxes at -224 and -262 . TRP1, in common with HIS3, HIS4 and TRP5 has a region of dyad symmetry upstream of the coding sequence which may play a role in initiation of transcription . The relative efficiency of gene expression directed by the complete 5' TRP1 region was assessed by comparison with that of PGK by inserting a cDNA for a human interferon-alpha downstream of their respective 5' regions . The respective interferon yields indicate that their in vivo expression capabilities are a function of their 5' regions.

J Biol Chem, 1983 Apr 25, 258(8), 5238 - 47
Repeated DNA sequences upstream from HIS1 also occur at several other co-regulated genes in Saccharomyces cerevisiae; Hinnebusch AG et al.; The HIS1 gene of Saccharomyces cerevisiae encodes ATP phosphoribosyltransferase, the first enzyme in the pathway of histidine biosynthesis . We have cloned this gene by complementation of a his1 auxotroph . The HIS1 coding region was localized within the cloned segment by assay of subcloned fragments for their ability to complement a his1 auxotroph . We determined the DNA sequence of the HIS1 region defined by this complementation test . S1 nuclease and exonuclease VII mapping of the 5' and 3' termini of HIS1 mRNA reveal considerable heterogeneity at both ends of the transcript, especially the 5' end which displays 13 different termini that span a 110-base pair region . Northern analysis shows that derepression of HIS1 enzyme activity under conditions of amino acid deprivation can be accounted for by an increase in the steady state level of HIS1 mRNA . There are no large differences between the relative levels of HIS1 mRNA molecules with different 5' termini in repressed and derepressed cells . In the DNA sequence upstream from the 5' termini of HIS1 mRNA we have found four closely related copies of a 9-base pair sequence . This sequence is also repeated in the 5' noncoding regions of HIS4, HIS3, and TRP5 . Closely related sequences are not found flanking a number of other yeast genes, suggesting that the repeated sequence plays a role in the regulation of amino acid biosynthetic genes subject to the general amino acid control.

Anal Biochem, 1983 Apr 15, 130(2), 469 - 70
Measurement of invertase activity of cells of Saccharomyces cerevisiae; Vitolo M et al.; The determination of the invertase activity of intact yeast cells presents a critical point, that is, the blockage of the enzyme action at a given moment . In this paper seven blockage methods were compared: the addition of 0.010 M sodium hydroxide solution, addition of 0.010 M sodium carbonate solution, addition of 0.010 M sodium carbonate solution followed by centrifugation (9750g; 10 min), immersion of the reacting mixture in a boiling water bath, immersion of the mixture in a -15 degrees C bath, filtration through a Millipore membrane, and addition of the first Somogyi's reagent followed by immersion in a boiling water bath . Only the last two methods lead to a rapid and effective blockage of the invertase activity.

Biochem Biophys Res Commun, 1983 Apr 15, 112(1), 47 - 54
Stereochemically distinct roles for sterol in Saccharomyces cerevisiae; Pinto WJ et al.; Cholesterol, (E)- but not (Z)-17(20)-dehydrocholesterol, 5 alpha-cholestan-3 beta-ol, sitosterol, and certain other sterols lacking a 24 beta-methyl group will replace (spare) most but not all of the 24 beta-methylsterol which has recently been found to be absolutely necessary for growth of oxygen-deprived wild type Saccharomyces cerevisiae in the presence of 2,3-iminosqualene . The results imply the existence of two stereochemically distinct roles for sterol in this organism . One of them (perhaps regulatory) requires, whereas the other (probably playing the so-called "bulk" membranous role) does not require the presence of the 24 beta-methyl group . The latter function, for which most of the sterol is needed, can be performed by various 24-alkyl- and 24-desalkylsterols.

J Biol Chem, 1983 Apr 10, 258(7), 4472 - 6
Stereochemical specificity for sterols in Saccharomyces cerevisiae; Pinto WJ et al.; When sterol biosynthesis in oxygen-deprived wild type Saccharomyces cerevisiae was prevented by the presence of 2,3-iminosqualene, an inhibitor of 2,3-oxidosqualene cyclase, an absolute requirement for a sterol with a 24 beta-methyl group was found . Neither the configuration nor the size of the alkyl group at C-24 could be altered . For instance, while 24 beta-methylcholesterol (22-dihydrobrassicasterol) permitted good growth, contrary to earlier work without the inhibitor no growth at all resulted from the presence of cholesterol or its 24 alpha-methyl-, 24 alpha-ethyl-, or 24 beta-ethyl derivatives (campesterol, sitosterol, and clionasterol, respectively) . The only sterol lacking a 24 beta-methyl group which allowed growth was desmosterol (24-dehydro-cholesterol), but desmosterol was metabolized to 24 beta-methylcholesterol by C1-transfer and reduction . When cholesterol supported growth in the absence of the inhibitor, small amounts of endogenously synthesized 24 beta-methylsterols (ergosterol and 22-dihydroergosterol) were identified . This previously unrecognized absolute specificity for both chirality and bulk at C-24 suggests the involvement of protein binding in at least one of the roles which sterol plays in this single-celled eukaryote.

Gene, 1983 Apr, 22(1), 31 - 9
Saccharomyces cerevisiae actin--Escherichia coli lacZ gene fusions: synthetic-oligonucleotide-mediated deletion of the 309 base pair intervening sequence in the actin gene; Larson GP et al.; Plasmids carrying gene fusions between the yeast (Saccharomyces cerevisiae) actin gene and an initiation-defective Escherichia coli lacZ (beta-galactosidase) gene have been constructed . Expression of beta-galactosidase in such fusion plasmids depends on transcription of the actin gene, and is possible only after the RNA-splicing machinery has removed from the primary RNA transcript the 309-bp intervening sequence (IVS) interrupting the actin coding region . Mutants deleting the actin IVS were constructed via synthetic oligonucleotide-mediated in vitro mutagenesis of the actin-beta-galactosidase fusion plasmid . A 17-base synthetic oligonucleotide was used to generate a 309-bp deletion which precisely removed the actin IVS . A partial deletion mutant was also constructed in which 272-bp, starting at the 5' end of the actin IVS, and including the 5' splice junction signal, were deleted . Both the complete and partial IVS-deletion mutants were transformed into yeast hosts . However, the partial deletion resulted in a greater than 98% reduction in beta-galactosidase activity . The precise deletion of the actin IVS did not reduce the levels of beta-galactosidase activity as compared with the parental fusion plasmid containing the intact IVS.

Mol Cell Biol, 1983 Apr, 3(4), 580 - 6
Fusion of the Saccharomyces cerevisiae leu2 gene to an Escherichia coli beta-galactosidase gene; Martinez-Arias AE et al.; The promoter and translation initiation region of the Saccharomyces cerevisiae leu2 gene was fused to the Escherichia coli beta-galactosidase gene . This fusion located the control region of the leu gene and orientated its direction of expression . When the fusion was placed into yeast cells, beta-galactosidase was expressed under the same regulatory pattern as the original leu2 gene product: its synthesis was repressed in the presence of leucine and threonine . Sensitive chromogenic substrates for beta-galactosidase were used to detect expression in isolated colonies growing on agar medium . Mutant yeast cells with increased beta-galactosidase activity were identified by the color of the colonies they formed . One class of mutants obtained appeared to affect ars1 plasmid maintenance, and another class appeared to affect beta-galactoside uptake.

Genetika, 1983 Apr, 19(4), 532 - 40
{Genetic effects of the decay in Saccharomyces cerevisiae yeast cells of the radionuclide products of nuclear fuel fission . II . Lethal mutagenic effects and the nature of mutations induced by an equilibrium mixture of 90Sr-90Y and 89Sr}; Gracheva LM et al.; The lethal and mutagenic effects and the nature of mutations induced by 90Sr-90Y and 89Sr in cells of the yeast Saccharomyces cerevisiae were studied . The lethal efficiency was determined for 89Sr as (7,6 +/- 1,05) X 10(-5) decay-1, for 90Sr-90Y-(3,3 +/- 1,6) X 10(-4) decay-1 . The mutagenic efficiency for ade1 and ade2 genes was determined for 89Sr as (8,3 +/- 2,5) X 10(-9) decay-1, for 90Sr-90Y-(2,9 +/- 1,5) X 10(-8) decay-1 . For ade2 locus, the spectrum of mutations induced by 89Sr was a follows: one deletion, 17% of frameshifts and 83% of base pair substitutions--51% of transversions, 22% of GC-AT transitions and 10% of AT-GC transitions . The data of the present work suggest that 90Sr-90Y and 89Sr are very efficient physical mutagens . The relative mutagenic efficiency (RME) was estimated for radionuclides studied.

Mol Cell Biol, 1983 Apr, 3(4), 654 - 61
Killer systems in Saccharomyces cerevisiae: three distinct modes of exclusion of M2 double-stranded RNA by three species of double-stranded RNA, M1, L-A-E, and L-A-HN; Wickner RB; M1 and M2 double-stranded RNAs (dsRNAs) code for the K1R1 and K2R2 killer toxin and resistance functions, respectively . Natural variants of a larger dsRNA (L-A) carry various combinations of the {EXL}, {HOK}, and {NEX} genes, which affect the K1 and K2 killer systems . Other dsRNAs, the same size as L-A, called L-B and L-C, are often present with L-A . We show that K1 killer strains have {HOK} and {NEX} but not {EXL} on their L-A (in disagreement with Field et al., Cell 31:193-200, 1982) . These strains also carry other L-size molecules detectable after heat-curing has eliminated L-A . The exclusion of M2 dsRNA observed on mating K2 strains with K1 strains is due to the M1 dsRNA (not the L-A dsRNA as claimed by Field et al.) in the K1 strains . Four independent mutants of a {KIL-k2} {NEX-o} {HOK-o} strain were selected for resistance to {EXL} exclusion of M2 ({EXLR} phenotype) . The {EXLR} phenotype showed non-Mendelian inheritance in each case, and these mutants had simultaneously each acquired {HOK} . The mutations were located on L-A and not on M2, and did not confer resistance to M1 exclusion of M2.

Mol Cell Biol, 1983 Apr, 3(4), 570 - 9
Comparative analysis of the 5'-end regions of two repressible acid phosphatase genes in Saccharomyces cerevisiae; Thill GP et al.; The nucleotide sequence of 5'-noncoding and N-terminal coding regions of two coordinately regulated, repressible acid phosphatase genes from Saccharomyces cerevisiae were determined . These unlinked genes encode different, but structurally related polypeptides of molecular weights 60,000 and 56,000 . The DNA sequences of their 5'-flanking regions show stretches of extensive homology upstream of, and surrounding, a "TATA" sequence and in a region in which heterogeneous 5' ends of the p60 mRNA were mapped . The predicted amino acid sequences encoded by the N-terminal regions of both genes were confirmed by determination of the amino acid sequence of the native exocellular acid phosphatase and the partial sequence of the presecretory polypeptide synthesized in a cell-free protein synthesizing system . The N-terminal region of the p60 polypeptide was shown to be characterized by a hydrophobic 17-amino acid signal polypeptide which is absent in the native exocellular protein and thought to be necessary for acid phosphatase secretion.

Mol Cell Biol, 1983 Apr, 3(4), 562 - 9
RNA and homology mapping of two DNA fragments with repressible acid phosphatase genes from Saccharomyces cerevisiae; Andersen N et al.; Two EcoRI restriction fragments carrying Saccharomyces cerevisiae repressible acid phosphatase genes were analyzed . Transcripts were mapped by restriction endonuclease cleavage of glyoxal-stabilized R-loops and by gel blot hybridizations to cDNA . Homology between the two fragments was examined by gel blots and heteroduplex analysis . Each fragment carried a region of about 1.5 kilobases that coded for a repressible acid phosphatase, and these regions showed homology to one another . In addition, one fragment carried a second region of somewhat lower homology that probably codes for the so-called constitutive acid phosphatase.

Mutat Res, 1983 Apr, 117(1-2), 213 - 24
Mutagenicity of 3 structurally related epoxides, with defined stereochemical configuration, in Saccharomyces cerevisiae and in V79 Chinese hamster cells; Turchi G et al.; 3 structurally related epoxides, 3,4-epoxycyclohexene, trans-1,2,3,4-diepoxycyclohexane and trans-3,4-epoxycyclohexane-r-1,trans-2-diol (anti isomer) were tested for their ability to induce both point mutation, mitotic gene conversion and recombination in a diploid strain (D7) of the yeast Saccharomyces cerevisiae, with and without a mammalian microsomal activation system, and the formation of 6-thioguanine-resistant mutants in V79 hamster cells . Genetic effects were related to the alkylating properties of the epoxides, as measured by alkylation of 4-(p-nitrobenzyl)pyridine (NBP) . Of the 3 epoxides, only 3,4-epoxycyclohexene, characterized by the highest reactivity towards NBP, induced all genetic effects in both test systems . A marginal activity was shown by trans-1,2,3,4-diepoxycyclohexane only in the yeast . The lack of genetic activity of the anti isomer of 3,4-epoxycyclohexane-1,2-diol, in spite of the formal similarity of its functional groups with those present in mutagenic polycyclic arene epoxydiols, was attributed to the dramatic reduction of lipophilicity of the molecule.

J Bacteriol, 1983 Apr, 154(1), 304 - 11
Phosphatidylinositol biosynthesis in Saccharomyces cerevisiae: purification and properties of microsome-associated phosphatidylinositol synthase; Fischl AS et al.; The membrane-associated phospholipid biosynthetic enzyme phosphatidylinositol synthase (cytidine 5'-diphospho-1,2-diacyl-sn-glycerol:myo-inositol 3-phosphatidyltransferase, EC 2.7.8.11) was purified 1,000-fold from the microsomal fraction of Saccharomyces cerevisiae . The purification procedure included Triton X-100 solubilization of the microsomal membranes, CDPdiacylglycerol-Sepharose (Larson et al., Biochemistry 15:974-979, 1976) affinity chromatography, and chromatofocusing . The procedure resulted in the isolation of a nearly homogeneous protein preparation with an apparent minimum subunit molecular weight of 34,000, as determined by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate . Phosphatidylinositol synthase was dependent on manganese and Triton X-100 for maximum activity . The pH optimum was 8.0 . Thioreactive agents inhibited enzyme activity . The energy of activation was found to be 35 kcal/mol (146,540 J/mol) . The enzyme was reasonably stable at temperatures of up to 60 degrees C.

J Gen Microbiol, 1983 Apr, 129 (Pt 4), 1103 - 8
Synthesis of a spore-specific surface antigen during sporulation of Saccharomyces cerevisiae; Dawes IW et al.; Antisera raised against purified yeast ascospores caused agglutination of both ascospores and vegetative cells . A spore-specific activity was obtained by absorbing out anti-vegetative activity with vegetative cells . The anti-vegetative cell activity was directed against mannan, and was probably due to exposure of some spore coat mannan at the spore surface since concanavalin A and lentil lectin also caused agglutination of ascospores . The spore-specific activity was probably determined by a protein or proteins, since extraction of spores with a mixture of sodium dodecyl sulphate and dithiothreitol markedly affected their agglutination by the spore-specific serum . The spore-specific antigen was synthesized in a soluble form during sporulation several hours before the appearance of the spore surface and the pool of soluble antigen declined as the spore was assembled . Synthesis of the soluble antigen was inhibited by adding cycloheximide at all times up to its first appearance in the sporulating cell.

J Biol Chem, 1983 Mar 25, 258(6), 3608 - 14
Glucose transport activity in isolated plasma membrane vesicles from Saccharomyces cerevisiae; Franzusoff AJ et al.; Purified plasma membranes prepared from yeast cells by mechanical agitation with glass beads exhibit no detectable sugar transport activity . However, the addition of phospholipid (asolectin) liposomes to the purified plasma membranes followed by freezing, thawing, and brief sonication produces membrane vesicles which exhibit D-glucose-specific transport activity . The characteristics of zero trans, equilibrium exchange, and influx counterflow exhibited by the membrane vesicles are similar to those of intact cells.

FEBS Lett, 1983 Mar 7, 153(1), 34 - 6
The possible functional significance of phosphatidylinositol in G1 arrest of Saccharomyces cerevisiae; Dudani AK et al.; Individual phospholipids were assayed in exponentially growing and G1-arrested temperature-sensitive cell division cycle (cdc) mutants of Saccharomyces cerevisiae . It was observed that cdc28 cells which are known to arrest at 'start' when shifted to their non-permissive temperature, resulted in a 40% decrease in phosphatidylinositol (PI) level while the phosphatidylserine (PS) content was doubled in these cells . The reduced level of PI was restored in cdc4 and cdc7 mutants which are known to arrest past the 'start' . The increase in PS level in cdc28 mutant which was probably to compensate the intrinsic charging of membrane environment, was also reduced in cdc4 and cdc7 mutants . Our results demonstrate that PI may play a role in yeast cell division and growth and that the abnormalities of cdc28 could also be related to PI decrease.

Biochem J, 1983 Mar 1, 209(3), 677 - 85
Metabolism of 3-deoxy-3-fluoro-D-mannose and 4-deoxy-4-fluoro-D-mannose by Saccharomyces cerevisiae S288C; Grier TJ et al.; Incubation of Saccharomyces cerevisiae S288C with 4-deoxy-4-fluoro-D-{1-14C}-mannose resulted in the formation of three metabolites that were characterized as 4-deoxy-4-fluoro-D-{1-14C}mannose 1,6-bisphosphate, 4-deoxy-4-fluoro-D-{1-14C}-mannose 6-phosphate and GDP-4-deoxy-4-fluoro-D-{1-14C}mannose . In addition, radioactive material was incorporated into a particulate fraction composed primarily of cell-wall polysaccharides . Compared with the 4-fluoro sugar, 3-deoxy-3-fluoro-D-{1-14C}mannose was not transported into yeast cells as well, and its conversion into sugar nucleotide was much less efficient . Metabolites that were isolated after incubation with the 3-fluoro analogue were identified as 3-deoxy-3-fluoro-D-{1-14C}mannose 1,6-bisphosphate, 3-deoxy-3-fluoro-D-{1-14C}mannose 6-phosphate and GDP-3-deoxy-3-fluoro-D-{1-14C}mannose . Little radioactivity was transferred into the cell-wall fraction.

Mol Cell Biol, 1983 Mar, 3(3), 415 - 20
Evidence for a new chromosome in Saccharomyces cerevisiae; Wickner RB et al.; The current yeast map has 16 chromosomes, each originally defined by a centromere-linked gene unlinked to previously defined centromere markers . We examined four genes, cly2, KRB1, AMY2, and tsm0115, each centromere linked, but previously thought to be not on chromosomes I to XVI . We found that AMY2 is linked to cly2, and both are on chromosome II . tsm0115 is on the left arm of chromosome XVI . We confirm the earlier evidence that KRB1 is not on chromosomes I through XVI . This gene thus defines a new chromosome XVII . We also report meiotic linkage of met4 and pet8 (on chromosome XIV), confirming the connection between the petx-kex2 fragment of XIV and the centromere of XIV.

Mol Cell Biol, 1983 Mar, 3(3), 371 - 9
Role of nuclear genes in expression of a mitochondrial tRNA gene in Saccharomyces cerevisiae; Wesolowski M et al.; In yeast mitochondria, most of the isoaccepting species of tyrosyl tRNA are coded by a mitochondrial gene, tyrA . A particular isoaccepting species is coded by a second mitochondrial gene, tyrB . This gene is not expressed in certain strains of yeast which show no deficient phenotype . Genetic crosses between strains expressing or not expressing the tyrB gene demonstrate that expression is controlled by specific nuclear genes and that a mutation of the tyrA gene can be bypassed when the tyrB gene is operative.

Mutat Res, 1983 Mar, 108(1-3), 161 - 8
The intrasanguineous host mediated assay procedure using Saccharomyces cerevisiae: comparison with two other metabolic activation systems; Frezza D et al.; 3 metabolic activation systems--organ homogenates, perfused liver, and the intrasanguineous host mediated assay (IHMA)--were compared in their abilities to activate demethylnitrosamine (DMN) and induce gene conversion in Saccharomyces cerevisiae D4 . Both rats and mice were used for the organ homogenates and IHMA studies . All activation systems were able to activate DMN; where the different organs were compared, liver was more active than kidney, followed by lung . The IHMA was the most sensitive of the systems examined.

Mutat Res, 1983 Mar, 108(1-3), 109 - 20
The induction of mutation and recombination following UV irradiation during meiosis in Saccharomyces cerevisiae; Kelly SL et al.; Irradiation of yeast cultures with ultraviolet light at discrete stages during meiosis produces cyclic variations in sensitivity, i.e . cells are more sensitive to the lethal effects of UV light prior to entry into the meiotic DNA synthesis, and this corresponds to a peak of induction of point mutation . Cells become more resistant to both induced point mutation and lethality as they enter meiotic DNA synthesis, but become more sensitive again during spore formation . The induced level of intragenic recombination rises during the period of commitment to recombination to a level indistinguishable from the full meiotic level of spontaneous intragenic recombination . Induced reciprocal recombination remains above the spontaneous level up to the point of commitment to sporulation.

J Bacteriol, 1983 Mar, 153(3), 1405 - 14
Recessive mutations conferring resistance to carbon catabolite repression of galactokinase synthesis in Saccharomyces cerevisiae; Matsumoto K et al.; A total of 37 recessive mutations showing enhanced resistance to the glucose repression of galactokinase synthesis have been isolated by a selection procedure with a GAL81 gal7 double mutant . These mutations were grouped into three different complementation classes . One class, reg1, contains mutants arising from mutations at a site close to, but complementing, the gal3 locus . The reg1 mutant also showed resistance to the glucose repression of invertase synthesis but not to that of alpha-D-glucosidase . The two other classes were identified as arising from recessive mutations at the GAL82 locus and the GAL83 locus, respectively, at which various dominant mutations were isolated previously . When in a constitutive background due to the GAL81 or gal80 mutation, the GAL82 and GAL83 mutations did not show a mutually additive effect on the resistance to glucose repression of galactokinase synthesis, while the reg1 and GAL82 (or GAL83) mutations did . Based upon the specific behavior of cells with various genotypes for the above genes in response to the concentration of galactose and glucose in the medium, we propose a model involving three independent circuits for glucose signals in the regulation of the structural genes for the galactose pathway enzymes.

J Bacteriol, 1983 Mar, 153(3), 1125 - 32
Mitochondrial translation products during release from glucose repression in Saccharomyces cerevisiae; Falcone C et al.; Mitochondrial protein synthesis was studied during release from glucose repression in Saccharomyces cerevisiae cells bearing different mitochondrial genomes . The increase in the rate of the synthesis of mitochondrial translation products was analyzed during respiratory induction . Different kinetic patterns were found for strains having a different structure of mitochondrial mosaic genes, even when the nuclear background was the same . A very limited response of the synthesis of the var1 ribosomal protein to inducing conditions was observed.

Proc Natl Acad Sci U S A, 1983 Mar, 80(6), 1730 - 4
Involvement of kinases in glucose and fructose uptake by Saccharomyces cerevisiae; Bisson LF et al.; Uptake of glucose, fructose, and the nonmetabolizable analog 6-deoxyglucose was measured in wild-type Saccharomyces cerevisiae and two mutant strains, one (hxk1 hxk2) lacking both hexokinase A(P-I) and B(P-II) but containing glucokinase (and hence able to grow on glucose but not fructose) and the other (hxk1 hxk2 glk) also lacking glucokinase (and not able to grow on glucose either) . Uptake of the nonmetabolized substances (i.e., 6-deoxyglucose in all three strains, fructose in the two mutants, and glucose in the triple mutant) reached a plateau at or below the external concentration . The kinetic characteristics of uptake were determined from 5-sec incubations by plotting velocity (V) vs . velocity/substrate concentration (V/S) curves . According to such plots, in the wild-type strain uptake had two components, "high affinity uptake" with Km values of ca . 1 mM for glucose and 6 mM for fructose and "low affinity uptake" with Km values of ca . 20 and 50 mM, respectively . The double kinase mutant showed both components for glucose but only the high Km component for fructose, while the triple kinase mutant showed only high Km uptake for both glucose and fructose . Genetic analysis showed that only in strains lacking both hexokinases (hxk1 hxk2) was the low Km system for fructose absent . Low Km uptake was restored to the triple mutant by introduction of the cloned wild-type genes: HXK1 or HXK2, for fructose uptake, and HXK1, HXK2, or GLK1, for glucose uptake . A phosphoglucose isomerase mutant had both low and high Km uptake for glucose . These results indicate the presence of two types of uptake mechanism for glucose and fructose in yeast, the functioning of one of which, the low Km system, is influenced by the cognate kinases.

Mutat Res, 1983 Mar, 108(1-3), 147 - 59
Metabolic activation of cytochrome P-450/P-448 in the yeast Saccharomyces cerevisiae; Kelly D et al.; The strains D6 and JD1 of the yeast Saccharomyces cerevisiae were used to assay the genetic activity of several compounds, benzo{a}pyrene, 15,16-dihydro-11-methyl-cyclopenta{a}phenanthren-17-one, 2-naphthylamine and cyclophosphamide, which require metabolic activation by cytochromes P-450 and P-448 to produce genetically active chemical species . Cells from both strains were harvested from cultures grown in low concentrations of glucose and switched to growth in high glucose containing media . Treatments under these conditions resulted in increased sensitivity of the test systems without the presence of an exogenous S9 mix and the presence of S9 was found not to enhance this sensitivity . The yeasts used under these treatment conditions showed a P-450/P-448 type metabolism.

Biochim Biophys Acta, 1983 Feb 28, 743(1), 58 - 68
Determination of the coordination geometry at the heme iron in three cytochromes c from Saccharomyces cerevisiae and from Candida krusei based on individual 1H-NMR assignments for heme c and the axially coordinated amino acids; Senn H et al.; The 1H-NMR lines of heme c and the axial ligands in reduced and oxidized Iso-1 and Iso-2 cytochromes c from Saccharomyces cerevisiae and in cytochrome c from Candida krusei were individually assigned and the conformation of the coordination sphere of the heme iron was investigated with the use of proton-proton Overhauser enhancement measurements and circular dichroism spectroscopy . The coordination geometry of the axial methionine and the axial histidine and the electronic structure of the heme were found to be closely similar in these yeast cytochromes c and in mammalian cytochromes c . In particular, R chirality at the sulfur atom of the iron-bound methionine was observed in both groups of proteins . Additional nuclear Overhauser enhancement studies of the spatial arrangement relative to the heme group of amino acid side-chains in the heme crevice of yeast ferrocytochromes c showed that the conformational homologies extend beyond the immediate coordination sphere of the heme iron . These data provide a conformational basis for observations on the functional properties of cytochromes c from yeast and mammalian species, which were reported previously by other groups.

J Biol Chem, 1983 Feb 25, 258(4), 2193 - 201
The isolation, characterization, and sequence of the pyruvate kinase gene of Saccharomyces cerevisiae; Burke RL et al.; The Saccharomyces cerevisiae gene encoding the glycolytic enzyme pyruvate kinase has been isolated by complementation of a pyk mutant with DNA from a wild type yeast genomic library . Pyruvate kinase enzyme activity is 20-fold higher in the transformant compared to the parental strain and is glucose inducible . The cloned gene has been localized by hybridization of DNA fragments to yeast poly(A+) RNA and by complementation of the mutant defect with select subclones . A DNA sequence of 2885 nucleotides encoding a protein of 499 amino acids is reported . A polypeptide chain of 34 residues of the deduced yeast amino acid sequence closely resembles a peptide sequence at the ADP binding site of bovine muscle pyruvate kinase . The 5' end of the pyruvate kinase mRNA has been mapped and starts within the DNA sequence CAAG at -38 to -27 nucleotides upstream from the first ATG . We note that the sequence PyAAPu in this region appears to be a common consensus site for yeast RNA polymerase II transcriptional starts.

Nucleic Acids Res, 1983 Feb 11, 11(3), 789 - 800
Sequence of a Drosophila DNA segment that functions in Saccharomyces cerevisiae and its regulation by a yeast promoter; Henikoff S et al.; We have determined the complete DNA sequence of a segment derived from Drosophila melanogaster that complements a yeast adenine-8 (ade8) mutation . This sequence, in conjunction with transcriptional analysis, predicts that a 206 amino acid sequence accounts for complementation . We have ligated a 1.5 kb promoter segment of the yeast alcohol dehydrogenase I (ADHI) gene just upstream of the open reading frame . This construction results in ADE8+ function that responds to glucose and ethanol in a manner similar to that for the chromosomal ADHI gene, as determined by a simple quantitative color assay for ADE8+ function.

Anal Biochem, 1983 Feb 1, 128(2), 294 - 301
Purification of DNA-dependent RNA polymerase II from Saccharomyces cerevisiae; Bitter GA; A rapid procedure for the purification of RNA polymerase II from Saccharomyces cerevisiae is described . Total RNA polymerase activity was solubilized from whole cells by sonication in 0.32 M (NH4)2SO4 and RNA polymerase II purified by polyethylenimine fractionation, ammonium sulfate precipitation, and chromatography on DEAE-cellulose, DEAE-Sephadex, and phosphocellulose . The procedure may be completed in 2.5 days and the resultant enzyme is judged to be greater than 90% pure.

Eur J Biochem, 1983 Feb 1, 130(2), 235 - 9
Modified plasma-membrane ATPase in mutants of Saccharomyces cerevisiae; Ulaszewski S et al.; Mutations affecting the plasma membrane ATPase of Saccharomyces cerevisiae were obtained by selecting mutants resistant to Dio-9 . In a plasma-membrane-enriched fraction of the mutant MG2130, the ATPase activity was resistant to vanadate (50% inhibition by 26 microM in the mutant compared to 1.3 microM in the parental strain) . Several catalytic properties of the membrane-bound ATPase were modified by 60-120% in the mutant which had a higher Km for MgATP and was more heatstable, less sensitive to mercurials, and more stimulated by monovalent cations than the parental type . A single mutation is responsible for the phenotypes of four independent allelic mutants . Resistance to Dio-9 in vivo and resistance to vanadate in vitro segregated together in three tetrads issued from a cross between the wild type and mutant . The mutation is semi-dominant as shown by expression of the mutant phenotype in a heterozygous diploid resulting from the cross between the wild type and mutant . It is concluded that the pma locus, affected by these mutations, is the structural gene either for the 100000-Mr subunit of plasma membrane ATPase or for a protein which tightly controls the conformation of the plasma-membrane ATPase within the membrane.

Mutat Res, 1983 Feb, 107(2), 249 - 64
Disomic and diploid meiotic products in Saccharomyces cerevisiae . Effect of vincristine, vinblastine, adriamycin, bleomycin, mitomycin C and cyclophosphamide; Sora S et al.; The effect of some antineoplastic drugs on the induction of disomic and diploid meiotic products in Saccharomyces cerevisiae was evaluated . Vincristine and vinblastine turned out not to be effective on any of the four genetic phenomena studied (sporulation, recombination, disomic induction and diploid induction) . Adriamycin showed only slight activity in inducing diploids, particularly during the first meiotic division . Cyclophosphamide was active on both phenomena leading to the formation of disomic and diploid spores . Mitomycin C and bleomycin were effective on the induction of diploids . In all of these inductions, the origin of diploids was due to failure of the second meiotic division . No significant effects were found on recombination frequency . As a general conclusion, one may assume that formation of aneuploid and diploid gametes are two distinct phenomena, not necessarily correlated from the point of view of the mechanism and of the specificity of induction.

J Biol Chem, 1983 Jan 25, 258(2), 854 - 8
Binding of proteins from the large ribosomal subunits to 5.8 S rRNA of Saccharomyces cerevisiae; Lee JC et al.; Specific binding of purified proteins from the large ribosomal subunits of Saccharomyces cerevisiae to 5.8 S rRNA was examined by three different methods: nitrocellulose membrane filtration, sucrose density gradient centrifugation, and RNA-Sepharose column chromatography . RNA-protein complex formation was proportional to the amount of proteins added to the reaction mixture . The binding of proteins to the RNA could be saturated . Such RNA-protein complexes were isolated on sucrose density gradients . Protein species present in these complexes were isolated, iodinated, and analyzed by two-dimensional polyacrylamide gel electrophoresis . Eleven proteins, L13, L14, L17, L19, L21, L24, L25, L29, L30, L33, and L39, were identified . By comparison, only six proteins interacted with the 5.8 S rRNA-Sepharose under similar ionic conditions . They were proteins L14, L21, L24, L27, L29, and L30 . To better characterize these binding proteins, the interaction of individual proteins with 5.8 S rRNA was studied by nitrocellulose membrane filtration . Proteins L14, L19, L21, L29, L33, and L39 were observed to bind individually with 5.8 S rRNA . Binding of each protein to the RNA could be saturated . The apparent association constants (K'a), measured at 4 degrees C and in 30 mM Tris-HCl, pH 7.4, 20 mM MgCl2, 330 mM KCl, and 6 mM beta-mercaptoethanol, ranged from 1.05 to 3.70 X 10(6) M-1.

J Biol Chem, 1983 Jan 10, 258(1), 610 - 5
Assembly of the mitochondrial membrane system . Characterization of the oxi2 transcript and localization of its promoter in Saccharomyces cerevisiae D273-10B; Thalenfeld BE et al.; The oxi2 gene of yeast mitochondrial DNA was previously shown to code for subunit 3 of cytochrome oxidase (Thalenfeld, B.E., and Tzagoloff, A . (1980) J . Biol . Chem . 255, 6173-6180) . In Saccharomyces cerevisiae D273-10B, a 3.6-kilobase (kb) transcript has been mapped to the oxi2 region of mitochondrial DNA . This transcript, presumed to be the messenger RNA of subunit 3, has been characterized by Northern hybridization analysis and by S1 nuclease mapping . The 3.6-kb transcript has a 5' untranslated leader of 490 nucleotides followed by a 807-nucleotide long coding sequence and a 3' extension of approximately 2450 nucleotides . The nucleotide sequence of the coding region in the 3.6-kb transcript is identical with the gene sequence, thus excluding the presence of introns in the oxi2 gene . Analysis of mitochondrial RNA in cytoplasmic petite mutants containing the oxi2 gene, but with varying lengths of flanking sequences, suggest the presence of a common promoter for oxi2 and the upstream valine tRNA . The promoter has been mapped to a 400-nucleotide long region located on the 5' side of the tRNA gene . Generation of the mature subunit 3 mRNA must, therefore, involve the excision of the tRNA from the primary transcript.

J Biol Chem, 1983 Jan 10, 258(1), 119 - 24
Purification and properties of glutamine synthetase from Saccharomyces cerevisiae; Mitchell AP et al.; We have purified glutamine synthetase over 130-fold from Saccharomyces cerevisiae . The enzyme exhibits a Km for glutamate of 6.3 mM and a Km for ATP of 1.3 mM in the biosynthetic reaction, with a pH optimum from 6.1 to 7.0 . Ten to twelve 43,000 molecular weight subunits comprise the active enzyme of 470,000 molecular weight . Rabbit antibodies prepared against the purified enzyme were used to show that induction of enzyme activity correlates with de novo synthesis of the enzyme subunit.

Mol Gen Genet, 1983, 191(1), 59 - 65
Induction of mating type interconversion in a heterothallic strain of Saccharomyces cerevisiae by DNA damaging agents; Schiestl R et al.; Mating type interconversion of the yeast, Saccharomyces cerevisiae, is an example of a directed genome rearrangement leading to a change in gene expression and in the differentiation state of a cell . In heterothallic haploid cells this switching of the mating type from a to alpha or vice versa, which is accomplished by an intrachromosomal gene conversion mechanism, is a rare event, happening about once per 10(6) cells per generation . Those cells that have changed their mating type can be trapped as diploid colonies by making them mate with tester cells possessing complementary markers . We found that treating haploids with UV light or with chemical carcinogens before they could mate resulted in a significant and dose-dependent enhancement of the numbers of diploid colonies . By genetic as well as by DNA hybridization analyses, these diploid clones were proved to be descendants of haploids which had changed their mating type by the bona fide gene conversion process . Thus, the DNA damaging agents had caused the induction of a directed gene rearrangement . It is suggested that induction of genome rearrangements might be part of a general response to DNA damage, at least in yeast cells . If similar responses also took place in cell populations constituting multicellular organisms, induced gene rearrangements and a generally enhanced mobility of the genome as a consequence of DNA damage might play a determining role in chemical and radiation-induced carcinogenesis.

Radiat Environ Biophys, 1983, 22(4), 269 - 80
Cellular radiation effects and hyperthermia cell cycle kinetics of radiation sensitive mutants of Saccharomyces cerevisiae after X-irradiation and hyperthermia; Fingerhut R et al.; Radiosensitive mutants rad2, rad9, and rad51 of Saccharomyces cerevisiae were X-irradiated with 120 Gy or 60 Gy, heated at 50 degrees C for 30 min or treated with a combination of both and incubated in nutrient medium at 30 degrees C . Cell number, percentage of budding cells, and cell cycle progression were determined in 45-min intervals . Cell cycle kinetics were investigated by flow cytofluorometry . Hyperthermia leads mainly to a lengthening of G1, whereas X-rays arrest cells of the rad2 and rad9 mutant in G2 and the rad51--mutant additionaly in a state with DNA contents above G2 . Cell division delay is influenced by oxygen in all strains but to a lesser extent in the rad2 mutant . The effect of the combined treatment appears to be merely additive in the rad2 and rad9 mutant while the rad51 mutant is sensitized to X-irradiation by hyperthermia . No selective action of hyperthermia on hypoxic cells was found.

Int J Biochem, 1983, 15(11), 1373 - 7
Mannan structure analysis of the fragile Saccharomyces cerevisiae mutant VY1160; Maerkisch U et al.; The fragile Saccharomyces cerevisiae mutant VY1160 has a cell-wall mannan with a lower molecular weight and a higher nitrogen content than the parental S288C strain . More mannobiose but less mannotriose and mannotetraose were found in O-glycosidically-bound oligosaccharides in the mutant . Acetolysis analysis of the alkali-stable mannan also showed a reduction of mannotetraose and an increase of the mannobiose fractions . Methylation analysis of total mannan, or side chains recovered after acetolysis, showed that in the mutant the amount of alpha(1-3)-linked mannose units prevailed over alpha(1-2)-linked residues.

Annu Rev Microbiol, 1983, 37, 623 - 60
Cell interactions and regulation of cell type in the yeast Saccharomyces cerevisiae; Sprague GF Jr et al.; Examination of the control of cell type in yeast at the molecular level and understanding of the biochemical basis of the cell-cell interactions involved in the mating process are clearly entering an extremely productive and exciting period . The tools and opportunities are now available to answer fundamental questions with regard to the mechanism of differential gene expression in eukaryotic cells by using cloned a-specific, alpha-specific, and haploid-specific genes as the probes . Basic questions concerning eukaryotic chromosome structure and organization can be addressed by elucidating the properties of the SIR/MAR regulators and their mode of action . Furthermore, the availability both of cloned MAT, HML, and HMR regions and of the HO gene will provide the material for unravelling the enzymology of the DNA transposition that occurs during mating type interconversion . The isolation of the structural genes for the pheromones and mutations that block pheromone production will provide useful information on how extracellular hormones are synthesized, processed, and secreted by eukaryotic cells . Moreover, the apparent mode of action of the phermonones through cyclic AMP as an intracellular "second messenger," and the genetic and biochemical tractability of yeast cells, may allow tracing of the entire pathway of hormonal regulation of a eukaryotic cell division cycle . These and other studies of the developmental biology of yeast cells will provide more important insights into fundamental aspects of the genetic control of developmental processes in eukaryotic cells.

Microbios, 1983, 38(151), 33 - 41
Sugar uptake during early ascosporulation in Saccharomyces cerevisiae; Ota A; The uptake rates of several sugars were examined in cells of Saccharomyces cerevisiae during early ascosporulation . The pH optimum of glucose uptake was about 7 both in the vegetative cells and in the cells after transfer to the sporulation medium . The glucose uptake rate of the cells 3 h after transfer to sporulation medium was about two-fold higher than that of vegetative cells . Uptake rates of glucose, mannose, and galactose increased during the first 3 h and then decreased slightly . The uptake rate of lactose increased throughout the 4 h period of observation, and the uptake rate of 2-deoxy-glucose was constant but at a very low level . Uptake rates of xylose and rhamnose, using the 3,5-dinitrosalicyclic acid method, were below detection levels.

Mol Gen Genet, 1983, 191(3), 407 - 12
Hypoxanthine: guanine phosphoribosyltransferase mutants in Saccharomyces cerevisiae; Woods RA et al.; Yeast mutants lacking activity of the enzyme hypoxanthine:guanine phosphoribosyltransferase (H:G-PRT) have been isolated by selecting for resistance to 8-azaguanine in a strain carrying the wild type allele, ade4%, of the gene coding for amidophosphoribosyltransferase (PRPPAT), the first enzyme of de novo purine synthesis . The mutants excrete purines and are cross-resistant to 8-azaadenine . They are recessive and represent a single complementation group, designated hpt1 . Ade4-su, a prototrophic allele of ade4 with reduced activity of PRPPAT, is epistatic to hpt1, suppressing purine excretion and resistance to azaadenine but not resistance to azaguanine . The genotype ade2hpt1 does not respond to hypoxanthine . Hpt1 complements and is not closely linked to the purine excreting mutants pur1 to pur5 . Hpt1 and pur6, a regultory mutant of PRPPAT, are also unlinked but do not complement, suggesting a protein-protein interaction between H:G-PRT and PRPPAT . Mycophenolic acid (MPA), an inhibitor of de novo guanine nucleotide synthesis, inhibits the growth of hpt1 and hpt1+ . Xanthine allows both genotypes to grow in the presence of MPA whereas guanine only allows growth of hpt1+ . Activity of A-PRT, X-PRT and H:G-PRT is present in hpt+ . Hpt1 lacks activity of H:G-PRT but has normal A-PRT and X-PRT.

Mol Gen Genet, 1983, 191(2), 314 - 8
Cyclic variations in sensitivity to X-irradiation during meiosis in Saccharomyces cerevisiae; Kelly SL et al.; Cyclic variation in mutation induction and lethality was found following X-irradiation during meiosis in Saccharomyces cerevisiae . An enhanced mutagenic response was found in meiotic G1 phase cells in comparison to cells later in meiosis, similar to the response shown during mitosis, but meiotic G1 phase cells appeared more resistant to the lethal effects of X-irradiation than mitotic G1 phase cells . Resistance to the lethal effects of X-rays was found during meiotic DNA synthesis in the strain SK1, which may indicate the operation of a sister-chromatid exchange repair mechanism . A difference was found between gene conversion which appeared to be at a maximum by the end of meiotic DNA synthesis and reciprocal recombination, which could be induced up to prophase I.

Mol Gen Genet, 1983, 191(1), 31 - 8
Regulation of expression of the galactose gene cluster in Saccharomyces cerevisiae . Isolation and characterization of the regulatory gene GAL4; Hashimoto H et al.; The GAL4 gene positively regulating the expression of the gene cluster GAL7-GAL10-GAL1 in the yeast Saccharomyces cerevisiae was isolated for its ability to suppress a recessive mutation in that gene . When the isolated gene was incorporated into a multi-copy plasmid, the GAL cluster genes in the host chromosome partially escaped the normal control; a yeast that harbors the plasmid bearing the GAL4 gene synthesized the galactose-metabolizing enzymes encoded by the GAL cluster genes at a low but significant level in the absence of galactose . If the GAL7 gene was amplified along with GAL4 on the multi-copy plasmid, the constitutive synthesis of Gal-1-P uridylyl transferase encoded by GAL7 was further pronounced and the enzyme activity reached the level of the fully induced wild-type yeast . Such an escape synthesis of the GAL enzymes was not detected if GAL4 or both GAL4 and GAL7 were carried by a single-copy plasmid . The results suggest that the escape synthesis of GAL enzymes observed in the GAL4-amplified yeast was a consequence of overproduction of the GAL4 protein . The GAL80 gene negatively regulating the GAL cluster genes was also isolated, and when amplified together with GAL4, no escape synthesis of the GAL enzymes was observed, suggesting that the balanced synthesis of two regulatory proteins was essential to maintain the repressed state of the GAL cluster genes.

Mol Gen Genet, 1983, 191(1), 17 - 21
The isolation and genetic analysis of sporulation-deficient mutants in Saccharomyces cerevisiae; Tsuboi M; Sporulation-deficient mutants were isolated from a homothallic strain of Saccharomyces cerevisiae . Sporulation was induced in these mutants by procedures to sporulate the products of protoplast fusion between mutants and wild-type strains . Spores formed in this way were crossed to wild-type strains in order to analyze them genetically . Twenty-three genes essential to sporulation were identified by tetrad analysis and complementation tests . Gene symbols spoT1 to spoT23 were tentatively assigned to them . These mutants fell into four classes by examination of premeiotic DNA synthesis and meiotic nuclear division: (i) Premeiotic DNA synthesis did not occur (spoT1 - spoT11); (ii) premeiotic DNA synthesis occurred but meiosis I did not occur (spoT12 - spoT15); (iii) meiosis II did not occur (spoT16 - spoT18); (iv) meiosis II occurred but mature spores were not formed (spoT19 - spoT23) . Genes spoT4, spoT8, spoT20, and spoT23 were mapped on chromosomes IV, II, XVI and XI, respectively . SpoT18-1 was a UAG nonsense mutation.

Mol Gen Genet, 1983, 190(3), 413 - 6
Analysis of mutagenic DNA repair in a thermoconditional repair mutant of Saccharomyces cerevisiae . II . Influence of cycloheximide on UV-irradiated exponentially growing rev2ts cells; Siede W et al.; The time course of REV2 dependent recovery from prelethal UV damage and UV-induced locus-specific reversion of the his5-2 allele was determined in temperature-shift experiments by use of a thermoconditional allele of the rev2 gene (rad5-8, rev2ts) . In UV-irradiated, exponentially growing rev2ts cells the REV2 dependent repair activity persists for up to 8 h at permissive temperature (23 degrees C), while the REV2 dependent mutagenic process is mostly completed within 2 h . The REV2 dependent process in exponentially growing cells is highly impaired by inhibition of protein synthesis . However, a REV2 dependent repair activity independent of de novo synthesis is detectable, even in the presence of up to 200 micrograms/ml cycloheximide, a response not found in stationary phase cells . Thus, the REV2 dependent process seems to be partially constitutive in exponentially growing cells . Additionally, exponentially growing rev2ts cells were considerably more UV-sensitive at restrictive temperature (36 degrees C) than were stationary phase cells.

Carcinogenesis, 1983, 4(7), 851 - 6
N-methyl-N'-nitro-N-nitrosoguanidine induced genetic change during the meiotic cell cycle in Saccharomyces cerevisiae: an absence of S-phase specificity; Kelly SL et al.; In the yeast Saccharomyces cerevisiae enhanced nuclear mutagenesis was found during meiotic DNA synthesis after treatment with N-methyl-N'-nitrosoguanidine (MNNG), but not during meiotic DNA synthesis . Further experiments found no significant variation in the mol . wt . of DNA from control or MNNG-treated samples during meiosis after alkaline sucrose gradient analysis . Using tritiated MNNG no variation in uptake of MNNG by meiotic cells was found, but enhanced binding of label from {3H}MNNG to mitochondrial DNA was detected during meiotic DNA synthesis when mitochondrial DNA synthesis occurred . In contrast, no enhanced binding of label from {3H}MNNG to the nuclear DNA during meiotic DNA synthesis was found.

Mol Gen Genet, 1983, 189(2), 256 - 62
Repair of gamma-ray induced DNA strand breaks in the radiation-sensitive mutant rad18-2 of Saccharomyces cerevisiae; Mowat MR et al.; The repair of gamma-ray induced DNA single and double-strand breaks was looked at in wild type and rad18-2 strains of the yeast Saccharomyces cerevisiae using sucrose gradient centrifugation . It was found that rad18-2 diploid cells could repair single and double-strand breaks induced by gamma-rays . It was also found that rad18-2 cells experienced a breakup of their DNA during post-irradiation incubation to a size smaller than seen in cells just receiving irradiation . This breakup of DNA in rad18-2 cells is not degradation due to cell death since wild type cells irradiated to similar low survival levels do not show this breakup of DNA with 8 h incubation . The breakup of DNA in rad18-2 cells is not due to replication gaps being formed by synthesis on a damaged template since treatment of rad18-2 a mating type cells with alpha factor, to prevent initiation of DNA synthesis, does not prevent breakup of the DNA.

Nauchnye Doki Vyss Shkoly Biol Nauki, 1983, (3), 25 - 8
{Effect of the photo-induced synthesis of serotonin on the photoreactivation of Saccharomyces cerevisiae yeasts}; Strakhovskaia MG et al.; The photoreactivation of yeast (Saccharomyces cerevisiae) cells irradiated by far-UV-light has been studied . It has been shown that the near-UV-induced (334, 365 nm) synthesis of serotonin in yeast is an underlying cause of its photoreactivation action spectrum change . The effect of serotonin is due to its binding to far-UV-irradiated DNA (pyrimidine dimers) making them unavailable to the photoreactivating enzyme . The data obtained support the idea that serotonin affects the repair enzymes which are engaged in an elimination of lethal photoproducts from DNA.

Mikrobiologiia, 1983 Jan-Feb, 52(1), 68 - 72
{Toxic effect of cysteine on cells of Saccharomyces cerevisiae growing on media of various compositions}; Damberg BE et al.; The oxidative processes in Saccharomyces cerevisiae were shown to be more susceptible to the toxic effect of cysteine as compared to the glycolytic processes . When cysteine was added to a suspension of resting cells, the activity of catalase and oxygen uptake decreased, particularly in the yeast was grown in a medium preventing catabolyte repression . As was demonstrated using disc-electrophoresis in acrylamide gel, cysteine decreased the activity of catalases T and A.

Proc Natl Acad Sci U S A, 1983 Jan, 80(2), 510 - 4
Mutations in PEP4 locus of Saccharomyces cerevisiae block final step in maturation of two vacuolar hydrolases; Zubenko GS et al.; The biosynthesis of carboxypeptidase Y (EC 3.4.16.1) and proteinase A (EC 3.4.23.6) in yeast cells involves a series of posttranslational events, the last of which is dependent upon a function supplied by the PEP4 gene . Because pep4 mutations result in a 90-95% reduction in the levels of activity of at least three additional vacuolar hydrolases, it is likely that these, and perhaps all, yeast vacuolar hydrolases are synthesized as inactive precursors, which mature by a common mechanism that depends upon a function supplied by the PEP4 gene . The pep4 mutation shows an apparent gene dosage effect on levels of activity of proteinases A and B but not on the level of activity of carboxypeptidase Y . This effect appears to come about because the maturation machinery is capable of discriminating among these hydrolase precursors.

Genetika, 1983, 19(1), 49 - 57
{Intragenic mitotic recombination induced by ultraviolet and gamma rays in radiosensitive mutants of Saccharomyces cerevisiae yeasts}; Zakharov IA et al.; The effect of UV- and gamma-irradiation on the survival and intragenic mitotic recombination (gene conversion) of 5 radiosensitive mutants was studied in comparison with the wild type . The level of spontaneous conversion was similar for RAD, rad2 and rad15, mutations xrs2 and xrs4 increasing and rad54 significantly decreasing it . The frequency of conversion induced by UV-light was greater in rad2, rad15 and xrs2 mutants and lower in xrs4, as compared to RAD . Gamma-irradiation caused induction of gene conversion with an equal frequency in RAD, rad2, rad15 . Xrs2 and xrs4 mutations slightly decreased gamma-induced conversion . In rad54 mutant, UV-and gamma-induced conversion was practically absent . In the wild type yeast, a diploid strain is more resistant than a haploid, whereas in rad54 a diploid strain has the same or an increased sensitivity, as compared to a haploid strain (the "inverse ploidy effect") . This effect and also the block of induced mitotic recombination caused by rad54 indicate the presence in the yeast Saccharomyces cerevisiae of repair pathways of UV- and gamma-induced damages acting in diploid cells and realised by recombination . The data obtained as a result of many years' investigation of genetic effects in radiosensitive mutants of yeast are summarised and considered.

Genetika, 1983, 19(1), 26 - 32
{Disorder of the repair of DNA double-stranded breaks in radiosensitive mutants of Saccharomyces cerevisiae yeasts}; Vishnevetskaia OIu et al.; DNA double-strand break repair and restoration of viability in X-irradiated diploid yeast cells homozygous for rad50, rad51, rad52, rad55 mutations were studies under conditions of keeping the cells in non-nutrient medium, after irradiation . All the cells were synchronized at the G1 stage of the cell cycle . In contrast to the wild-type yeast, this group of mutants are unable to repair DNA double-strand breaks and do not enhance viability, when kept in non-nutrient medium after irradiation.

Eur J Cell Biol, 1983 Jan, 29(2), 121 - 5
Analysis of anaphase B in saccharomyces cerevisiae using a monoclonal antibody against yeast tubulin; King SM et al.; In Saccharomyces cerevisiae the mitotic spindle is composed of approximately 10 continuous microtubules, that pass uninterrupted between the 2 spindle pole bodies (SPBs), and up to 34 discontinuous microtubules, 17 of which arise from each SPB and terminate in the nucleoplasm {7, 12} . We have previously shown that during spindle elongation (anaphase B) the number of continuous microtubules decreases rapidly until at a length of 4 microns only 1 remains {8} . Further elongation from 4 to 8 microns is achieved with only a single continuous microtubule . Here we have extended our analysis of spindle elongation by investigating the timing of this event by indirect immunofluorescence microscopy using a monoclonal antibody to yeast tubulin . From the frequency distribution of spindle lengths in exponential phase cultures of the haploid wild type strain A364A, the time in mitosis and the velocity of spindle elongation have been calculated . For the first 55 min of mitosis spindle elongation proceeds at a slow rate, 0.36 microns/min . There is then a marked increase to a maximal value of 4.16 microns/min . Correlated with data from previous ultrastructural studies on the decrease in continuous microtubule number during anaphase B, this 12-fold increase in velocity is shown to occur at the single microtubule stage . These experiments provide further evidence to suggest that the role of the spindle is to regulate the rate of anaphase B which is actively achieved by an, as yet, unknown motive force generating system . Antibody staining has also allowed the terminal phenotypes of 2 diploid cell division cycle (cdc) mutants to be more accurately described . This has shown that, although arrested synchronously as judged by phase contrast microscopy, these strains do in fact exhibit considerable heterogeneity in spindle length.

Biokhimiia, 1983 Jan, 48(1), 62 - 8
{Some properties of two forms of alpha-glucosidase from Saccharomyces cerevisiae-II}; Krakenaite RP et al.; The amino acid composition of two forms of alpha-glucosidase from the yeast Saccharomyces cerevisiae-II was established and the values of Km, V, kcat and kcat/Km for maltose, maltotriose and p-nitrophenyl-alpha-D-glucopyranoside (PNPG) were determined . PNPG possessed a much higher affinity for the enzyme as compared to sucrose, maltose and maltotriose . The value of V decreased in the following order: PNPG greater than sucrose greater than maltose greater than greater than maltotriose . No differences between the kinetic parameters of individual forms of alpha-glucosidase were observed . Glucose, fructose and methyl-alpha-glucoside act as competitive inhibitors . The two forms of alpha-glucosidase under study have an identical pH optimum and thermal stability.

Mol Gen Genet, 1983, 191(3), 339 - 46
An insertion mutation associated with constitutive expression of repressible acid phosphatase in Saccharomyces cerevisiae; Toh-e A et al.; The PHO83 mutation in Saccharomyces cerevisiae, which had been detected on the basis of constitutive production of repressible acid phosphatase and mapped at the end of the PHO5 locus, was analysed by Southern hybridization with cloned DNA fragments of the PHO5 gene as probe . It was shown that this mutant has a DNA insertion of about 6 kilobase pairs, probably in the 5'-noncoding region of the PHO5 gene . Production of repressible acid phosphatase by the PHO83 mutant is partially independent of the function of the PHO2 and PHO4 genes, the positive regulatory genes whose functions are indispensable for PHO5 expression . PHO83 mutants are constitutive in a and alpha cells, either haploid or diploid, but not in non-mating cells, MATa/MAT alpha or a certain sterile mutation . These observations strongly suggest that the PHO83 mutation is caused by insertion of a Ty element in the 5'-noncoding region of the PHO5 gene.

EMBO J, 1983, 2(7), 1049 - 54
A nuclear mutation prevents processing of a mitochondrially encoded membrane protein in Saccharomyces cerevisiae; Pratje E et al.; Subunit II of cytochrome oxidase is encoded by the mitochondrial OXI1 gene in Saccharomyces cerevisiae . The temperature-sensitive nuclear pet mutant ts2858 has an apparent higher mol . wt . subunit II when analyzed on lithium dodecylsulfate (LiDS) polyacrylamide gels . However, on LiDS-6M urea gels the apparent mol . wt . of the wild-type protein exceeds that of the mutant . Partial revertants of mutant ts2858 that produce both the wild-type and mutant form of subunit II were isolated . The two forms of subunit II differ at the N-terminal part of the molecule as shown by constructing and analyzing nuclear ts2858 and mitochondrial chain termination double mutants . The presence of the primary translation product in the mutant and of the processed form in the wild-type lacking 15 amino-terminal residues was demonstrated by radiolabel protein sequencing . Comparison of the known DNA sequence with the partial protein sequence obtained reveals that six of the 15 residues are hydrophilic and, unlike most signal sequences, this transient sequence does not contain extended hydrophobic parts . The nuclear mutation ts2858 preventing post-translational processing of cytochrome oxidase subunit II lies either in the gene for a protease or an enzyme regulating a protease.

J Bacteriol, 1983 Jan, 153(1), 345 - 9
Isolation of Saccharomyces cerevisiae TRP3; Paluh JL et al.; Several plasmids, isolated from two plasmid pools, complemented a Saccharomyces cerevisiae trp3 mutant with defective indole-3-glycerol-phosphate synthase activity . Restriction mapping indicated that a 1.2-kilobase StuI segment was common to all complementing plasmids . Southern blot hybridization established that a cloned 5.2-kilobase BamHI fragment was derived intact from chromosomal DNA . A yeast trp3 mutant transformed with trp3-complementing plasmids contained approximately 40-fold elevated indole-3-glycerol-phosphate synthase activity . These plasmids also complemented an Escherichia coli trpC mutant, and transformants exhibited enzyme activity . Yeast trp3 is therefore associated with a 1.2-kilobase StuI DNA segment.

FEBS Lett, 1982 Dec 27, 150(2), 329 - 31
Does a cyclic AMP-dependent phosphorylation initiate the transfer of trehalase from the cytosol into the vacuoles in Saccharomyces cerevisiae?
Wiemken A, Schellenberg M.
Trehalase activity in a yeast protoplast lysate increased approximately 40-times upon preincubation with cAMP and ATP . The activity present without the preincubation could all be sedimented at 8000 x g, for 10 min confirming the previously reported localization of the active trehalase (Ta) in the vacuoles . Virtually all the trehalase activity newly formed upon the preincubation, however, was found in the soluble fraction, indicating that a trehalase-zymogen (Tz) is located in the cytosol . This raises the possibility that a cAMP-dependent phosphorylation not only transforms Tz to Ta but also initiates the transfer of trehalase from the cytosol into the vacuoles.

C R Seances Acad Sci III, 1982 Dec 20, 295(13), 787 - 90
{Demonstration of maturation promoting factor activity in Saccharomyces cerevisiae}; Weintraub H et al.; Microinjection of a crude yeast extract into fully grown immature Xenopus laevis oocytes induces the reinitiation of meiotic maturation as does Xenopus laevis oocyte MPF (Maturation Promoting Factor) by itself . Using several strains of yeast exhibiting temperature sensitive cell cycle block (cdc), it was observed that yeast extract exhibits MPF activity only when obtained during the time interval from the G2 period to the metaphase . This observation stresses eucaryotic ubiquity of MPF.

Nucleic Acids Res, 1982 Dec 11, 10(23), 7791 - 808
The primary structure of the Saccharomyces cerevisiae gene for 3-phosphoglycerate kinase; Hitzeman RA et al.; The DNA sequence of the gene for the yeast glycolytic enzyme, 3-phosphoglycerate kinase (PGK), has been obtained by sequencing part of a 3.1 kbp HindIII fragment obtained from the yeast genome . The structural gene sequence corresponds to a reading frame of 1251 bp coding for 416 amino acids with no intervening DNA sequences . The amino acid sequence is approximately 65 percent homologous with human and horse PGK protein sequences and is in general agreement with the published protein sequence for yeast PGK . As for other highly expressed structural genes in yeast, the coding sequence is highly codon biased with 95 percent of the amino acids coded for by a select 25 codons (out of 61 possible) . Besides structural DNA sequence, 291 bp of 5'-flanking sequence and 286 bp of 3'-flanking sequence were determined . Transcription starts 36 nucleotides upstream from the translational start and stops 86-93 nucleotides downstream from the translational stop . These results suggest a non-polyadenylated mRNA length of 1373 to 1380 nucleotides, which is consistent with the observed length of 1500 nucleotides for polyadenylated PGK mRNA . A sequence TATATATAAA is found at 145 nucleotides upstream from the translational start . This sequence resembles the TATAAA box that is possibly associated with RNA polymerase II binding.

Biochemistry, 1982 Dec 7, 21(25), 6397 - 403
Purification and characteristics of a mitochondrial endonuclease from the yeast Saccharomyces cerevisiae; von Tigerstrom RG; Saccharomyces cerevisiae contains a membrane-bound mitochondrial nuclease . The enzyme was purified nearly 500-fold from sphaeroplasts of the organism by differential centrifugation, differential solubilization, heparin-agarose chromatography, and gel filtration . A final specific activity of 98 mumol min-1 (mg of protein)-1 was obtained . The enzyme required further purification to achieve homogeneity . Two peaks of activity were obtained after gel filtration with apparent molecular weights of 140000 and 57000 . Otherwise, these two components have nearly identical characteristics . Without detergent the enzyme is insoluble and has very low activity . Zwittergent 3-14 or Triton X-100 in the presence of KCl could be used to solubilize and activate the enzyme . A number of other detergents were much less effective in solubilizing or activating the nuclease . The enzyme requires Mg2+ for activity, and this can be replaced to some degree by Mn2+ but not by Ca2+ or Zn2+ . It is most active at pH 6.5-7.0 and degrades the substrate to small oligonucleotides with 5'-phosphate ends . The relative rates of hydrolysis were 100 for poly(A), 31 for ssDNA, 19 for RNA, 2.1 for dsDNA, and less than or equal to 0.2 for poly(C) . Under the assay conditions used the enzyme appears to constitute about 90% of the total nuclease activity of the cell . The enzyme is unstable, especially at neutral and alkaline pH.

Genetics, 1982 Dec, 102(4), 653 - 64
Recessive UAA suppressors of the yeast Saccharomyces cerevisiae; Ono BI et al.; Recessive lysine-independent revertants were isolated from a psi + haploid strain of the yeast Saccharomyces cerevisiae containing one of the leucine-inserting UAA suppressors, SUP29, and various UAA mutations including lys1-1 . The majority of the revertants were found to have recessive suppressors in addition to the pre-existing SUP29 mutation . The recessive suppressors were able to suppress only a very limited number of UAA mutations, and none of the UAG mutations thus far examined . The recessive inefficient UAA suppressors were assigned to three complementation groups, sup111, sup112, and sup113 . A high incidence of gene conversion was observed for an allele of sup111 . An antisuppressor acting on sup111, but not detectably on SUP29, was inadvertently obtained during the course of the study . Interactions between SUP29, sup111 and the antisuppressor asu12 were studied.

Genetics, 1982 Dec, 102(4), 679 - 90
Genetic properties of mutations at the PEP4 locus in Saccharomyces cerevisiae; Zubenko GS et al.; Yeast cells that inherit mutations at the PEP4 locus exhibit a pronounced phenotypic lag in the expression of the mutant phenotype imparted by these mutations . This lag appears to extend to all of the enzymes that are affected by the pep4-3 mutation . For at least two of the enzymatic activities, phenotypic lag shows mitotic cosegregation . Phenotypic lag is found for meiotic progeny and for mitotic segregants from heterokaryons . The phenotypic lag in the expression of the carboxypeptidase Y deficiency is abolished by nonsense mutations in either PRC1, the structural gene for carboxypeptidase Y, or PRB1, the structural gene for proteinase B . Models to explain these observations are proposed.

Genetics, 1982 Dec, 102(4), 665 - 77
PEP4 gene function is required for expression of several vacuolar hydrolases in Saccharomyces cerevisiae; Jones EW et al.; The pep4-3 mutation results in a 90-95% reduction in the levels of five vacuolar hydrolases in yeast, including proteinases A and B, carboxypeptidase Y, RNase(s) and the repressible alkaline phosphatase . The mutation is without effect on two secreted glycoproteins, on an enzyme of the vacuolar membrane, and on a proteinase located outside of the vacuole . Mutations at the PEP4 locus exhibit a dosage effect on the levels of some, but not all, of the enzymes whose expression requires the function of the gene.

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