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