Mol Cell Biol, 1985 Jul, 5(7), 1743 - 9 Deletion analysis identifies a region, upstream of the ADH2 gene of Saccharomyces cerevisiae, which is required for ADR1-mediated derepression; Beier DR et al.; Deletion analysis was used to identify sequences upstream of the ADH2 gene of Saccharomyces cerevisiae that are required for its regulation . 5' and 3' internal deletions of the ADH2 control region were created in vitro, and the fragments were ligated adjacent to the ADH1 promoter and structural gene . Hybrid genes with 3' deletions extending from -119 to -216 (the start site of ADH2 transcription is designated +1) were fully repressed and derepressed to high levels . Hybrid genes with 3' deletions extending from -119 to -257 were repressed but failed to significantly derepress . Hybrid genes lacking the -216 to -257 region also failed to respond to ADR1-5c, a mutant allele of the unlinked regulatory gene ADR1, which confers constitutive expression on ADH2 . This implies that the region between these deletion endpoints, which includes a 22-base-pair sequence of dyad symmetry, is required for efficient derepression of an adjacent promoter . Internal deletions extending in the 3' direction from position -1141 confirmed these results . Deletion mutants lacking the region -1141 to -259 were normally regulated, whereas deletions extending from -1141 to -115 were not derepressible . These results support the hypotheses that the ADH2 promoter may normally be in an inactive conformation in the yeast chromosome and that derepression of ADH2 requires positive activation mediated through an upstream activation sequence located between 216 and 257 base pairs 5' to the start site of ADH2 transcription . No evidence for a DNA sequence mediating repression was obtained. Biochimie, 1985 Jul-Aug, 67(7-8), 717 - 23 Synthetic oligodeoxyribonucleotides as tools in molecular genetics: the characterization of the CYC1 (iso-1-cytochrome c encoding) locus of Saccharomyces cerevisiae; Smith M; The use of synthetic oligodeoxyribonucleotides as tools for the isolation, characterization and mutagenesis of eukaryote genes has played a major role in the molecular definition of the CYC1 locus of Saccharomyces cerevisiae, which is the structural gene for the apoprotein of iso-1-cytochrome c . Thus, the possibility of using a synthetic oligodeoxyribonucleotide as a probe to identify and monitor the isolation of a specific gene was first established by model studies which defined the melting temperatures (TmS) for duplexes of oligonucleotides of different lengths and base compositions . This led to the isolation of the CYC1 locus using a synthetic 13 nucleotide probe . A more convenient strategy for determination of DNA sequences by the Sanger method was provided by using synthetic oligodeoxyribonucleotides as primers with denatured double-strand plasmid DNA as template . By this means, the sequence of the CYC1 locus was determined by "walking" along the gene without isolating restriction fragments of the DNA or separating DNA strands . Synthetic oligodeoxyribonucleotides, used as primers for reverse transcriptase with mRNA as template, were also used to precisely define the 5'- and 3'-ends of the iso-1-cytochrome c mRNA . Yet another application of synthetic oligodeoxyribonucleotides is their use as specific mutagens after in vitro incorporation into double-stranded DNA . In the case of iso-1-cytochrome c, this mutagenic strategy is being used to define the role of conserved amino-acids in cytochrome function.(ABSTRACT TRUNCATED AT 250 WORDS) Can J Microbiol, 1985 Jul, 31(7), 654 - 6 Comparison of the mitochondrial endonucleases from Neurospora crassa and Saccharomyces cerevisiae; von Tigerstrom RG et al.; The endonucleases from Neurospora crassa and Saccharomyces cerevisiae are not closely related antigenically . They also differ with respect to their activity at pH 8, their degree of hydrophobicity, and their sensitivity to elevated temperatures . However, the two nucleases have similar specific activities, are inhibited by EDTA, and have nearly identical substrate specificities . Since the enzymes also have the same mode of action and intracellular location, these similarities may indicate that they have the same physiological role despite their structural differences. Biochemistry, 1985 Jun 18, 24(13), 3332 - 7 Structure-activity relationships in the dodecapeptide alpha-factor of Saccharomyces cerevisiae: position 6 analogues are poor inducers of agglutinability; Baffi RA et al.; Five des-Trp1,Cha3,X6 analogues of alpha-factor, where X = Ala, Val, Ile, Nle, or D-Leu and X = Leu in the natural alpha-factor sequence, were prepared by solution-phase techniques utilizing isobutyl chloroformate or 1-hydroxybenzotriazole accelerated active esters as the coupling agents . Purification to 98% or greater homogeneity was accomplished by high-performance liquid chromatography on a reversed-phase muBondapak C18 column with methanol/water/trifluoroacetic acid as the mobile phase . Three of the synthesized analogues (X6 = Val, Ile, Nle) induced morphogenesis and increased agglutinability in a cells . These substitutions demonstrate that a gamma-branched side chain at position 6 is not essential for biological activity . All of the active analogues induced morphogenesis at lower concentrations than they induced enhanced agglutinability . These results and other structure-activity relationships {Baffi, R . A., Shenbagamurthi, P., Terrance, K., Becker, J . M., Naider, F., & Lipke, P . (1984) J . Bacteriol . 158, 1152-1156} indicate that the agglutination and morphological responses to alpha-factor can be varied independently . Replacement of Leu6 with Ala or D-Leu resulted in inactive analogues that were not antagonistic for alpha-factor activity . Cell-mediated hydrolysis experiments indicated that the biological activities of the alpha-factor analogues are independent of their rates of degradation . All position 6 analogues were hydrolyzed more slowly than the parent compound, suggesting that the enzyme which degrades alpha-factor is highly specific for the native structure. Nucleic Acids Res, 1985 Jun 11, 13(11), 3791 - 804 Size and position of intervening sequences are critical for the splicing efficiency of pre-mRNA in the yeast Saccharomyces cerevisiae; Klinz FJ et al.; The size of the 309 bp actin gene intron of the yeast Saccharomyces cerevisiae was enlarged by inserting DNA fragments of different lengths and sequence . Enlarging the intron above 551 bp, the largest known yeast intron, led to a decrease in splicing efficiency . The effect on transcript splicing was dependent on the length of the inserted fragments rather than their sequence . It was also observed that insertion of the actin gene intron into different regions of the normally unsplit yeast YP2 gene, significantly influenced the efficiency of splicing of the resulting transcripts . The splicing efficiency of splicing of with the increase of the distance between the mRNA cap site and the intervening sequence. J Biol Chem, 1985 Jun 10, 260(11), 6960 - 5 A comparative study of the characteristics of eIF-2 and eIF-2-ancillary factor activities from yeast Saccharomyces cerevisiae and rabbit reticulocytes; Ahmad MF et al.; The characteristics of yeast eukaryotic initiation factor 2 (eIF-2) and Co-eIF-2A have been studied and compared with those of the corresponding factors from rabbit reticulocytes . 1) Unlike eIF-2r, purified eIF-2y did not contain bound GDP . 2) Purified eIF-2y preparation contained GTPase activity and dephosphorylated GTP to GDP . 3) An anti-eIF-2r preparation which predominantly precipitated the gamma-subunit (Mr 54,000) of eIF-2r also precipitated the larger subunit (Mr 54,000) of eIF-2y . 4) Unlike eIF-2r, ternary complex formation by eIF-2y was not inhibited by Mg2+ . 5) Both Co-eIF-2A20y and Co-eIF-2r significantly enhanced Met-tRNAf binding to eIF-2y and, again, Mg2+ did not have any effect on this stimulated Met-tRNAf binding to eIF-2y . 6) Both Co-eIF-2A20y and Co-eIF-2r were similarly effective in stimulating Met-tRNAf binding to eIF-2r in the absence of Mg2+ . However, in the presence of Mg2+, Co-eIF-2A20y was significantly less effective than Co-eIF-2r as Co-eIF-2A20y did not promote displacement of GDP from eIF-2r X GDP . 7) eIF-2y bound {3H}GDP and this binding was significantly enhanced in the presence of Mg2+ . Also, {3H}GDP in the preformed eIF-2y X {3H}GDP complex was rapidly exchanged with exogenously added unlabeled GDP in the presence of Mg2+ . Co-eIF-2A20y had no effect on GDP binding to eIF-2y nor on GDP exchange reactions . 8) Reticulocyte heme-regulated protein synthesis inhibitor, which phosphorylated almost completely (in excess of 80%) the alpha-subunit (Mr 38,000) of eIF-2r, also phosphorylated similarly the smaller subunit (Mr 36,000) of eIF-2y . However, such phosphorylation had no significant effect on ternary complex formation, GDP binding, and GDP exchange reactions. J Biol Chem, 1985 Jun 10, 260(11), 6955 - 9 Purification and properties of eukaryotic initiation factor 2 and its ancillary protein factor (Co-eIF-2A) from yeast Saccharomyces cerevisiae; Ahmad MF et al.; Two peptide chain initiation factor activities, eIF-2y and Co-eIF-2A20y, were purified from the high speed supernatant fraction of the yeast Saccharomyces cerevisiae and their properties were studied . 1) In sodium dodecyl sulfate-polyacrylamide gels, purified eIF-2y showed two major polypeptide bands corresponding to molecular weights of 54,000 and 36,000 . The Mr 54,000 band was significantly more intense than the Mr 36,000 band, indicating the possible presence of two polypeptides of equal molecular weight in this band . The molecular weight of eIF-2y, determined using a density gradient centrifugation method, was approximately 140,000 . 2) In sodium dodecyl sulfate-polyacrylamide gel, purified Co-eIF-2A20y showed a single polypeptide band corresponding to a molecular weight of 20,000 . A similar molecular weight for Co-eIF-2A20y was also found using a density gradient centrifugation method . 3) In partial reactions, eIF-2y bound Met-tRNAf in the presence of Mg2+ . The reaction required GTP . Co-eIF-2A20y stimulated Met-tRNAf binding to eIF-2y (2-3-fold) and also rendered the complex stable to 3 X 10(-5) M aurintricarboxylic acid . 4) This Co-eIF-2A20y activity was heat-labile and N-ethylmaleimide-insensitive . 5) Antibodies were prepared by injecting rabbits with homogeneous Co-eIF-2A20y . Such anti-Co-eIF-2A20y inhibited (60%) protein synthesis in a yeast cell-free protein synthesizing system and completely blocked Co-eIF-2A20y stimulation of Met-tRNAf . 40 S initiation complex formation . Protein synthesis inhibition by anti-Co-eIF-2A20y was almost completely reversed by preincubation of the antibodies specifically with homogeneous Co-eIF-2A20y. Eur J Biochem, 1985 Jun 3, 149(2), 401 - 4 Mutations affecting the enzymes involved in the utilization of 4-aminobutyric acid as nitrogen source by the yeast Saccharomyces cerevisiae; Ramos F et al.; We present genetic evidence for the enzymes 4-aminobutyrate: 2-oxoglutarate aminotransferase (EC 2.6.1.19) and succinate-semialdehyde dehydrogenase {NAD(P)+} (EC 1.2.1.16) constituting the functional pathway for the utilization of 4-aminobutyric acid as a nitrogen source by Saccharomyces cerevisiae . We show that the pathway is induced by 4-aminobutyric acid and that the presence of the pathway enzymes probably requires the integrity of a positive control element. Genetika, 1985 Jun, 21(6), 914 - 8 {Molecular nature of the direct mutations induced by 4-nitro-quinoline-N-oxide and 3-methyl-4-nitroquinoline-N-oxide in the ADE2 gene of Saccharomyces cerevisiae}; Kasinova GV et al.; The lethal and mutagenic effects and the nature of forward mutations in ADE2 gene induced by highly carcinogenic agent 4-nitroquinoline-N-oxide (4NQO) and its noncarcinogenic analogue 3-methyl-4-nitroquinoline-N-oxide (3M4NQO) have been examined in Saccharomyces cerevisiae . It is shown that 3M4NQO is more toxic than 4NQO . Both are very efficient mutagens: the mutagenic efficiency for ADE1 and ADE2 genes was 7.9 X 10(-5) for 4NQO and 10.5 X 10(-5) for 3M4NQO . The base pair substitutions are the main type of induced mutations in ADE2 gene (95 and 89% for 4NQO and 3M4NQO, respectively); among these 40% transversions for 4NQO and 63% for 3M4NQO, GC----AT transitions-32 and 31% for 4NQO and 3M4NQO, respectively, AT----GC transitions-23 and 22% for 4NQO and 3M4NQO, respectively . The results obtained indicate that 4NQO and 3M4NQO induce the same spectrum of mutations in ADE2 gene and that both mutagens are nonspecific in yeast cells. Mol Cell Biol, 1985 Jun, 5(6), 1522 - 4 DNase I-hypersensitive sites in the galactose gene cluster of Saccharomyces cerevisiae; Proffitt JH; Five DNase I-hypersensitive regions were associated with the Saccharomyces cerevisiae galactose gene cluster during both galactose induction and glucose repression of transcription . Four hypersensitive regions were located in areas flanking the GAL cluster genes, and one site occurred within GAL10 . A DNase I-hypersensitive region located between the 5' ends of divergently transcribed GAL10 and GAL1 contained sequences essential for the transcription of both genes. Mol Cell Biol, 1985 Jun, 5(6), 1512 - 21 Saccharomyces cerevisiae coordinates accumulation of yeast ribosomal proteins by modulating mRNA splicing, translational initiation, and protein turnover; Warner JR et al.; The rate of accumulation of each ribosomal protein is carefully regulated by the yeast cell to provide the equimolar ratio necessary for the assembly of the ribosome . The mechanisms responsible for this regulation have been examined by introducing into the yeast cell extra copies of seven individual ribosomal protein genes carried on autonomously replicating plasmids . In each case studied the plasmid-borne gene was transcribed to the same degree as the genomic gene . Nevertheless, the cell maintained a balanced accumulation of ribosomal proteins, using a variety of methods other than transcription . (i) Several ribosomal proteins were synthesized in substantial excess . However, the excess ribosomal protein was rapidly degraded . (ii) The excess mRNA for two of the ribosomal protein genes was translated inefficiently . We provide evidence that this was due to inefficient initiation of translation . (iii) The transcripts derived from two of the ribosomal protein genes were spliced inefficiently, leading to an accumulation of precursor RNA . We present a model which proposes the autogenous regulation of mRNA splicing as a eucaryotic parallel of the autogenous regulation of mRNA translation in procaryotes . Finally, the accumulation of each ribosomal protein was regulated independently . In no instance did the presence of excess copies of the gene for one ribosomal protein affect the synthesis of another ribosomal protein. Proc Natl Acad Sci U S A, 1985 Jun, 82(11), 3785 - 9 RAS2 of Saccharomyces cerevisiae is required for gluconeogenic growth and proper response to nutrient limitation; Tatchell K et al.; Saccharomyces cerevisiae contains two genes with remarkable homology to members of the ras oncogene family . These two genes, RAS1 and RAS2, constitute an essential gene family since spores with disruptions of both genes fail to grow . We report here that strains containing RAS2 disruptions have three distinct phenotypes . First, they fail to grow efficiently on nonfermentable carbon sources . Second, they hyperaccumulate the storage carbohydrates glycogen and trehalose . Third, diploid cells homozygous for the RAS2 disruptions sporulate on rich media . Extragenic suppressors have been isolated that suppress the gluconeogenic defect . These suppressors fall into at least three complementation groups, mutations in two of which bypass the normal requirement of RAS for cell viability, allowing cells containing neither RAS gene to grow . The phenotype of the RAS2 mutant and extragenic suppressors implicate RAS with some function in the normal response to nutrient limitation. Mutat Res, 1985 Jun-Jul, 150(1-2), 217 - 24 Modulation in cytochrome P-420 and P-450 content in Saccharomyces cerevisiae according to physiological conditions and genetic background; von Borstel RC et al.; The diploid strain D5 of Saccharomyces cerevisiae, relative to other strains of yeast, has a large amount of cytochrome P-450 present during the logarithmic phase of growth and a low amount of cytochrome P-420 . As the stationary phase of growth is approached, an increasing intensity of absorbance is observed at 420 nm . If the cells are suspended in buffer during mid-logarithmic growth, the absorbance at 450 nm disappears and absorbance at 420 nm is increased after the cells have been held in buffer for 24 h . At late logarithmic growth, the absorbance at 450 nm is still retained after the cells have been held in buffer for 24 h . Within 44 h of the time of harvest, the absorbance at 450 nm disappears completely and the absorbance at 420 nm is intense . Cytoplasmic petite variants of strain D5 have less of both cytochromes P-450 and P-420 than does the grande D5 strain; the absorbances at 450 and 420 nm are retained up to 96 h when the cells are held in buffer . Haploid spores of strain D5 exhibit absorbances at 450 and 420 nm during the logarithmic phase of growth, and these absorbances are retained after the cells are held in buffer for 24 h . An hypothesis is proposed which states that cytochrome P-450 is the membrane-bound form and cytochrome P-420 is free in the cytosol; the cytochromes interconvert and are active in either state until the associated enzymes disassociate. DNA, 1985 Jun, 4(3), 203 - 10 Expression of rat liver cytochrome P-450MC cDNA in Saccharomyces cerevisiae; Oeda K et al.; Rat liver cytochrome P-450MC cDNA was inserted between the ADH1 promoter and terminator regions of the yeast expression vector pAAH5 . On introduction of the resulting recombinant plasmid pAMC1, Saccharomyces cerevisiae cells synthesized up to 8 X 10(5) molecules per cell of the cytochrome P-450MC protein, most of which was localized in yeast microsomes . Approximately half of the synthesized cytochrome contained heme in the enzyme molecule . These formed a functional electron-transport chain in the microsomes which exhibited aryl hydrocarbon hydroxylase activity toward benzo{a}pyrene. J Gen Microbiol, 1985 Jun, 131 ( Pt 6), 1425 - 32 Regulation by ammonium of glutamate dehydrogenase (NADP+) from Saccharomyces cerevisiae; Bogonez E et al.; The activity of glutamate dehydrogenase (NADP+) (EC 1.4.1.4; NADP-GDH) of Saccharomyces cerevisiae is decreased under conditions in which intracellular ammonia concentrations increases . A high internal ammonia concentration can be obtained (a) by increasing the ammonium sulphate concentration in the culture medium, and (b) by growing the yeast either in acetate + ammonia media, where the pH of the medium rises during growth, or in heavily buffered glucose + ammonia media at pH 7.5 . Under these conditions cellular oxoglutarate concentrations do not vary and changes in NADP-GDH activity appear to provide a constant rate of oxoglutarate utilization . The following results suggest that the decrease in NADP-GDH activity in ammonia-accumulating yeast cells is brought about by repression of synthesis: (i) after a shift to high ammonium sulphate concentrations, the number of units of activity per cell decreased as the inverse of cell doubling; and (ii) the rate of degradation of labelled NADP-GDH was essentially the same in ammonia-accumulating yeast cells and in controls, whereas the synthesis constant was much lower in the ammonia-accumulating cells than in the controls. J Cell Biol, 1985 Jun, 100(6), 1854 - 62 Outer plaque assembly and spore encapsulation are defective during sporulation of adenylate cyclase-deficient mutants of Saccharomyces cerevisiae; Uno I et al.; Sporulation in diploid cells homozygous for the cyr1-2 mutation of the yeast Saccharomyces cerevisiae was examined . This mutation causes a defect in adenylate cyclase and temperature-sensitive arrest in the G1 phase of the mitotic cell cycle . The cyr1-2/cyr1-2 diploid cells were able to initiate meiotic divisions, but produced predominantly two-spored asci at the restrictive temperature . Temperature-sensitive period for production of two-spored asci was approximately 12 h after the transfer of cells to the sporulation medium . The levels of cAMP increased during this period in the wild type and cyr1-2/cyr1-2 diploid cells incubated at the permissive temperature, but remained at an extremely low level in the cyr1-2/cyr1-2 diploid cells incubated at the restrictive temperature . Dyad analysis of the cyr1-2 strain indicated that meiotic products were randomly included into ascospores . Fluorescent microscopy of the cyr1-2/cyr1-2 diploid cells incubated at the restrictive temperature revealed that individual haploid nuclei were enclosed in each of the two spores after meiosis . About half of the cyr1-2/cyr1-2 diploid cells entered normal meiosis 1 producing two normal spindle pole bodies with inner and outer plaques, and the other half entered abnormal meiosis 1 producing one normal spindle pole body and one defective spindle pole body without out plaque . At meiosis II, some cells contained a pair of normal spindle pole bodies and other cells contained pairs of normal and abnormal spindle pole bodies. Biochim Biophys Acta, 1985 May 30, 845(2), 151 - 7 The effect of chemical modification of Saccharomyces cerevisiae on electrophoretic mobility, cell-wall structure and amphotericin B uptake; Ellul H et al.; Saccharomyces cerevisiae NCYC 239 suspended in solutions of NaCl showed two distinct plateaus in plots of electrophoretic mobility vs . pH, corresponding to pKa values of approx . 2 and 5 . This is in contrast to cells suspended in buffer where only a single pKa (4) can be determined . Modification of cells with KI/I2 or nitrous acid led to altered electrophoretic mobility, indicating the presence of sulphydryl and amino groups, respectively, in the yeast cell surface, whereas uranyl nitrate modification had little effect, suggesting phosphate groups to be absent . Electron micrographs showed visible effects of KI/I2 and nitrous acid modification on cell membrane structure, and in these modified cells amphotericin B uptake was rapid . It is suggested that diffusion through the cell wall is the rate-limiting step for amphotericin B uptake . An activation energy of 20 kJ X mol-1 was determined for uptake of amphotericin B by unmodified cells. J Biol Chem, 1985 May 25, 260(10), 5871 - 4 Nuclear mutants of Saccharomyces cerevisiae with altered subunits 4, 5, and 6 of cytochrome oxidase; Koerner TJ et al.; A collection of pet mutants of Saccharomyces cerevisiae has been screened for lesions in cytochrome oxidase . Three different complementation groups have been identified to consist of strains with altered forms of subunits 4, 5, or 6 that are known to be encoded by nuclear genes . The mutant proteins cross-react with antiserum to the holoenzyme or to the individual subunits but exhibit either an increase or decrease in size . In each instance the mutation imparts a respiratory deficient phenotype which is due to reduced levels of cytochrome oxidase activity in the mitochondria . These results indicate that each of the three proteins is required either for the catalytic activity or for the assembly of functional cytochrome oxidase. Arch Biochem Biophys, 1985 May 15, 239(1), 184 - 90 Partial purification and properties of two histone acetyltransferases from the yeast, Saccharomyces cerevisiae; Lopez-Rodas G et al.; Two histone acetyltransferases, A and B, have been extracted and partially purified from yeast cells . The purification scheme included ammonium sulfate precipitation, and chromatography on DEAE-Sepharose and Sephadex G-200 . The basic properties of both enzymes closely correspond to those of acetyltransferase A and B found in higher eucaryotes . Yeast enzyme A elutes from DEAE-Sepharose prior to acetyltransferase B, and it is activated by low concentrations of DNA and strongly inhibited by p-chloromercuribenzoate (PCMB) . Enzyme B is inhibited by DNA over the entire range of concentrations tested and it is less sensitive to PCMB than enzyme A . When assayed with yeast whole histones, enzyme B shows a marked specificity toward histone H4, although H3 and H2B are also accepted as substrates . Enzyme A preferentially catalyzes the acetylation of yeast H2B and H3, with the other two core histones being acetylated to a much lesser extent. J Biol Chem, 1985 May 10, 260(9), 5596 - 602 Attempted expression of a human initiator tRNA gene in Saccharomyces cerevisiae; Drabkin HJ et al.; In attempts to overproduce the wild type and, eventually, mutant human initiator methionine tRNAs for use in structure-function relationship studies, we have investigated the expression of the wild type human initiator tRNA gene in the yeast Saccharomyces cerevisiae, both in vitro and in vivo . We find that the yeast extract, while capable of accurately transcribing several yeast tRNA genes, does not transcribe the human initiator tRNA gene . In addition, when the human initiator tRNA gene is introduced into yeast as part of a 2 mu vector, no expression of the human tRNA gene was detected . A yeast alanine tRNA gene similarly introduced into yeast is expressed efficiently . The block in expression of the human tRNA gene is at the level of transcription and not processing . The yeast cell-free extract can accurately process precursors of the same human initiator tRNA made in a HeLa cell-free extract . Surprisingly, although the human tRNA gene has essentially the same intragenic control elements as the yeast initiator tRNA gene, the human tRNA gene competes extremely poorly for transcription factors in yeast extracts . In the course of screening a yeast DNA bank for initiator tRNA clones we have isolated and sequenced three yeast tRNA genes corresponding to glycine, alanine, and aspartic acid tRNAs . The sequence of glycine tRNA gene differs from the published tRNA sequence in having an additional nucleotide in the variable loop . The alanine tRNA gene codes for a new tRNA . All three genes are transcriptionally active in yeast extracts. J Mol Biol, 1985 May 5, 183(1), 31 - 42 Partial suppression of an ochre mutation in Saccharomyces cerevisiae by multicopy plasmids containing a normal yeast tRNAGln gene; Pure GA et al.; We screened a yeast genomic library for recombinant DNA plasmids that complemented the ultraviolet (u.v.) sensitivity of a strain of Saccharomyces cerevisiae designated rad4-3 that is defective in excision repair of DNA . A multicopy plasmid (pNF4000) with a 9.4 X 10(3) base-pair yeast DNA insert partially complemented the u.v . sensitivity of rad4-3, but not of two other rad4 allelic mutants (rad4-2 and rad4-4), or of other u.v.-sensitive rad mutants . The yeast insert was analyzed by restriction mapping, DNA-DNA hybridization, DNA-tRNA hybridization and DNA sequencing . This analysis revealed the presence of a normal tRNAGln gene, a yeast sigma element situated 5' to the transfer RNA gene, a Ty element and a solo delta element . Deletion analysis of pNF4000 showed that the tRNAGln gene is required for partial complementation of the u.v . sensitivity of rad4-3 . Furthermore, a multicopy plasmid containing a tRNAGln gene derived from a different region of the yeast genome also partially complemented the u.v . sensitivity of rad4-3 . The rad4-3 mutation is suppressed following transformation with a plasmid containing the known ochre suppressor SUP11-o, indicating that it is an ochre mutation . We therefore conclude that when expressed in sufficient quantity, normal tRNAGln (which usually decodes the sense codon CAA) can weakly suppress the nonsense ochre codon UAA, and suggest that this represents an example of wobble occurring at the first rather than at the third position of the codon. J Bacteriol, 1985 May, 162(2), 579 - 83 alpha-Aminoadipate as a primary nitrogen source for Saccharomyces cerevisiae mutants; Zaret KS et al.; In contrast to wild-type strains of the yeast Saccharomyces cerevisiae, lys2 and lys5 mutants are able to utilize alpha-aminoadipate as a primary source of nitrogen . Chattoo et al . (B . B . Chattoo, F . Sherman, D . A . Azubalis, T . A . Fjellstedt, D . Mehnert, and M . Ogur, Genetics 93:51-65, 1979) relied on this difference in the effective utilization of alpha-aminoadipate to develop a procedure for directly selecting lys2 and lys5 mutants . In this study we used a range of mutant strains and various media to determine why normal strains are unable to utilize alpha-aminoadipate as a nitrogen source . Our results demonstrate that the anabolism of high levels of alpha-aminoadipate through the biosynthetic pathway of lysine results in the accumulation of a toxic intermediate and, furthermore, that lys2 and lys5 mutants contain blocks leading to the formation of this intermediate. Radiat Res, 1985 May, 102(2), 254 - 63 An attempt to stimulate mitosis in Saccharomyces cerevisiae with the ultraviolet luminescence from exponential phase cultures of this yeast; Quickenden TI et al.; Neither cell division nor growth of Saccharomyces cerevisiae were stimulated by the ultraviolet luminescence produced by adjacent exponential phase cultures of the yeast . The study included experiments in which the inocula (density = 5 X 10(7) cells cm-3) were irradiated and in which lag phase cultures (densities = 1 X 10(6) or 5 X 10(6) cells cm-3) were irradiated for 30 min with the yeast luminescence . These results do not support the claims of earlier workers that dividing cells can stimulate mitosis in optically coupled cultures by the so-called "mitogenetic effect." Mol Cell Biol, 1985 May, 5(5), 907 - 15 Characterization of an essential Saccharomyces cerevisiae gene related to RNA processing: cloning of RNA1 and generation of a new allele with a novel phenotype; Atkinson NS et al.; The RNA1 gene product is believed to be involved in RNA metabolism due to the phenotype of a single conditionally lethal, temperature-sensitive allele, rna1-1 . We cloned the RNA1 gene and determined that it produces a 1,400-nucleotide polyadenylated transcript . On a multicopy plasmid, the mutant rna1-1 allele partially complements the rna1-1 temperature-sensitive growth defect . This suggests that the temperature-sensitive nature of the rna1-1 allele results from the synthesis of a product with lowered activity or stability at elevated temperatures or from a decrease in synthesis of the rna1-1 product at the restrictive temperature . A chromosomal disruption of RNA1 behaves as a recessive lethal mutation . Haploids bearing the disruption were isolated by sporulating a diploid heterozygous for the disrupted allele and the rna1-1 allele and possessing an episomal copy of the RNA1 gene . Analysis of the rescued haploids bearing the chromosomal disruption indicated that the recessive lethal phenotype of the RNA1 disruption is not merely due to a block in spore germination . Unexpectedly, diploids heterozygous for the disruption and the rna1-1 alleles become aneuploid for chromosome XIII at a frequency of 2 to 5% . It appears that the disrupted RNA1 allele on a multicopy plasmid also promotes aneuploidy for chromosome XIII . Promotion of aneuploidy seems to be a phenotype of this particular allele of RNA1. Mutat Res, 1985 May, 149(3), 359 - 64 Mutagenic effect and mutation spectrum induced by 3H decay in the 8th position of DNA purine bases in Saccharomyces cerevisiae; Korolev VG et al.; Lethal and mutagenic effects and the mutation spectrum induced by 3H decay in the 8th position of adenine and guanine in yeast DNA have been studied . For haploid cells labelled with {8-3H}deoxyadenosinemonophosphate (8-3H-A) and {8-3H}deoxyguanosinemonophosphate (8-3H-G), the lethal efficiencies were determined as (3.0 +/- 0.8) X 10(-3) decay-1 and (3.8 +/- 0.6) X 10(-3) decay-1, respectively, and the mutagenic efficiencies as (5.7 +/- 1.1) X 10(-8) decay-1 and (8.7 +/- 1.4) X 10(-8) decay-1, respectively . The lethal effect of {8-3H}purines may be explained as being due to internal beta-irradiation . In contrast, the local effect of 3H-transmutation was twice as mutagenic as beta-irradiation when the induction of forward gene mutations was examined . Within the spectrum of mutations induced by 8-3H-G, a preference for GC----AT transitions was observed. Mutat Res, 1985 May, 149(3), 339 - 51 Acetone, methyl ethyl ketone, ethyl acetate, acetonitrile and other polar aprotic solvents are strong inducers of aneuploidy in Saccharomyces cerevisiae; Zimmermann FK et al.; A diploid yeast strain D61.M was used to study induction of mitotic chromosomal malsegregation, mitotic recombination and point mutation . Several ketones (including acetone and methyl ethyl ketone) and some organic acid esters (including the methyl, ethyl and 2-methoxyethyl esters of acetic acid) and acetonitrile strongly induced aneuploidy but not recombination or point mutation . Only diethyl ketone induced low levels of recombination and point mutation in addition to aneuploidy . Related compounds were weak inducers of aneuploidy: methyl n-propyl ketone, the methyl esters of propionic and butyric acid, acetic acid esters of n- and iso-propanol and ethyl propionate . No mutagenicity was found with n-butyl and isoamyl acetate, ethyl formate, acetyl acetone (2,5-dipentanone) and dioxane . Methyl isopropyl ketone induced only some recombination and point mutation but no aneuploidy . Efficient induction was only observed with a treatment protocol in which growing cells were exposed to the chemicals during a growth period of 4 h at 28 degrees C followed by incubation in ice for more than 90 min, usually overnight for 16-17 h . Aneuploid cells could be detected in such cultures during a subsequent incubation at growth temperature if the chemical was still present . Detailed analysis showed that there was a high incidence of multiple events of chromosomal malsegregation . It is proposed that the mutagenic agents act directly on tubulin during growth so that labile microtubules are formed which dissociate in the cold . When cells are brought back to temperatures above the level critical for reassembly of tubulin and allowed to grow, faulty microtubules are formed. J Bacteriol, 1985 May, 162(2), 763 - 7 Formation of septum-like structures at locations remote from the budding sites in cytokinesis-defective mutants of Saccharomyces cerevisiae; Slater ML et al.; Cell wall structures that partition membrane-bound portions of cytoplasm were formed at sites along the peripheral wall when a cytokinesis-defective cell division cycle mutant (cdc3) of Saccharomyces cerevisiae was grown at a restrictive temperature . The appearance of these structures, as observed in electron micrographs, was similar to that of normal septa . Aberrant septa were also detected in cytokinesis mutants harboring mutations cdc10, cdc11, and cdc12, after growth at 37 degrees C . Formation of the abnormal septa was abolished by the introduction, in a cdc3-containing strain, of additional cell cycle mutations that precluded events leading to cytokinesis and cell division . These results showed that septum formation can occur in the absence of cytokinesis . Formation of the abnormal structures was controlled by the same sequences of cell cycle events as formation of normal septa but was not subject to the spatial controls that ensure association of the septum with the budding site. Biochim Biophys Acta, 1985 Apr 25, 834(2), 249 - 55 Purification of a phospholipase B inhibitor from Saccharomyces cerevisiae; Witt W et al.; The phospholipase B activity of plasma membrane vesicles from Saccharomyces cerevisiae is inhibited by the 100 000 X g supernatant of mechanically disrupted yeast cells . A 1850-fold purification of the inhibitor activity was achieved by gel filtration, ion exchange chromatography with DEAE-cellulose and hydrophobic interaction chromatography with Octyl-Sepharose . SDS-polyacrylamide gel electrophoresis of the purified inhibitor revealed two main bands with an apparent Mr of 60 000 and 26 500 . The phospholipase B activity was strongly reduced but not completely blocked by this preparation, while the lysophospholipase and transacylase reactions, which are catalyzed by the same membrane-bound enzymes (Witt, W . et al . (1984) Biochim . Biophys . Acta 795, 108-116), were not affected. Eur J Biochem, 1985 Apr 15, 148(2), 405 - 6 Abnormal amino acid metabolism in mutants of Saccharomyces cerevisiae affected in the initiation of sporulation; Dickinson JR et al.; Amino acid metabolism was examined during sporulation in a wild-type strain of Saccharomyces cerevisiae and in a mutant homozygous for the spd1 mutation (derepression of sporulation) . In the wild-type initiation of sporulation involves a rise in intracellular glutamate . In the derepressed mutant there was no further increase of intracellular glutamate: glutamate was already present at a greater concentration than in the wild-type . These elevated glutamate levels are apparently due to reduction in the activity of 2-oxoglutarate dehydrogenase . One consequence of the elevated glutamate levels in diploids homozygous for the spd1 mutation is the production of considerable amounts of glycyl-L-proline, glutathione and saccharopine. Nucleic Acids Res, 1985 Apr 11, 13(7), 2357 - 72 The nucleotide sequence of the RAD3 gene of Saccharomyces cerevisiae: a potential adenine nucleotide binding amino acid sequence and a nonessential acidic carboxyl terminal region; Reynolds P et al.; The RAD3 gene of Saccharomyces cerevisiae is required for excision of pyrimidine dimers and is essential for viability . We present the nucleotide sequence of the RAD3 protein coding region and its flanking regions, and the deduced primary structure of the RAD3 protein . In addition, we have mapped the 5' end of RAD3 mRNA . The predicted RAD3 protein contains 778 amino acids with a calculated molecular weight of 89,779 . A segment of the RAD3 protein shares homology with several adenine nucleotide binding proteins, suggesting that RAD3 protein may react with ATP . The twenty carboxyl terminal amino acids of RAD3 protein are predominantly acidic; however, deletion of this acidic region has no obvious effect on viability or DNA repair. J Biol Chem, 1985 Apr 10, 260(7), 4531 - 40 Characterization of factors and DNA sequences required for accurate transcription of the Saccharomyces cerevisiae 5 S RNA gene; Taylor MJ et al.; Cell-free extracts prepared from yeast cells have previously been shown to direct selective and accurate in vitro transcription of tRNA and 5 S RNA genes . We have further analyzed the transcription factors and DNA sequences required for in vitro transcription of the yeast 5 S RNA gene . Fractionation of a yeast extract has identified a 5 S RNA gene-specific factor required, in addition to the two previously described tRNA factors (Klekamp, M . S., and Weil, P . A . (1982) J . Biol . Chem . 257, 8432-8441), for accurate transcription of the 5 S RNA gene by RNA polymerase III . Transcription of variant 5 S RNA genes has indicated that a region of the gene extending from residue +57 to residue +99 is essential for directing specific initiation of transcription . Although the 5' flanking and initial coding sequence is not absolutely required for transcription of the gene, some of the variant genes which have substitutions in this region are less actively transcribed than the wild-type gene . Transcription initiates on some of the variant genes at a position equivalent to +1 in the substituted sequence, while on other variant genes transcription initiates further upstream. Appl Environ Microbiol, 1985 Apr, 49(4), 863 - 6 Image analysis method for the rapid counting of Saccharomyces cerevisiae cells; Costello PJ et al.; An image analysis system which incorporates a microscope, video camera, monitor, and Apple computer and which uses image area to count Saccharomyces cerevisiae cells is described and evaluated . Yeast cell suspensions of densities of up to 100 X 10(6) cells per ml can be counted when viewed in the counting chamber of a hemacytometer . The yeast image area measured depends upon the light intensity used to illuminate the yeast cells, the sharpness of the image focused on the monitor, and the grey level selected when scanning the digitized image on the monitor of the Apple computer, all of which can be controlled . The image area also depends upon the yeast strain and medium in which the culture is grown, but it is not affected by the concentration of sugar or ethanol in which the yeast cells are suspended . Yeast growth measured by image analysis can be calibrated to give results similar to those obtained with hemacytometer counting . Yeast cells can be counted in the presence of high cell densities of bacteria by adjusting the grey level at which the digitized image is scanned. Mol Cell Biol, 1985 Apr, 5(4), 816 - 22 Isolation of the SUP45 omnipotent suppressor gene of Saccharomyces cerevisiae and characterization of its gene product; Himmelfarb HJ et al.; The Saccharomyces cerevisiae SUP45+ gene has been isolated from a genomic clone library by genetic complementation of paromomycin sensitivity, which is a property of a mutant strain carrying the sup45-2 allele . This plasmid complements all phenotypes associated with the sup45-2 mutation, including nonsense suppression, temperature sensitivity, osmotic sensitivity, and paromomycin sensitivity . Genetic mapping with a URA3+-marked derivative of the complementing plasmid that was integrated into the chromosome by homologous recombination demonstrated that the complementing fragment contained the SUP45+ gene and not an unlinked suppressor . The SUP45+ gene is present as a single copy in the haploid genome and is essential for viability . In vitro translation of the hybrid-selected SUP45+ transcript yielded a protein of Mr = 54,000, which is larger than any known ribosomal protein . RNA blot hybridization analysis showed that the steady-state level of the SUP45+ transcript is less than 10% of that for ribosomal protein L3 or rp59 transcripts . When yeast cells are subjected to a mild heat shock, the synthesis rate of the SUP45+ transcript was transiently reduced, approximately in parallel with ribosomal protein transcripts . Our data suggest that the SUP45+ gene does not encode a ribosomal protein . We speculate that it codes for a translation-related function whose precise nature is not yet known. Mol Cell Biol, 1985 Apr, 5(4), 787 - 96 Alpha-factor structural gene mutations in Saccharomyces cerevisiae: effects on alpha-factor production and mating; Kurjan J; The role of alpha-factor structural genes MF alpha 1 and MF alpha 2 in alpha-factor production and mating has been investigated by the construction of mf alpha 1 and mf alpha 2 mutations that totally eliminate gene function . An mf alpha 1 mutant in which the entire coding region is deleted shows a considerable decrease in alpha-factor production and a 75% decrease in mating . Mutations in mf alpha 2 have little or no effect on alpha-factor production or mating . The mf alpha 1 mf alpha 2 double mutants are completely defective in mating and alpha-factor production . These results indicate that at least one alpha-factor structural gene product is required for mating in MAT alpha cells, that MF alpha 1 is responsible for the majority of alpha-factor production, and that MF alpha 1 and MF alpha 2 are the only active alpha-factor genes. Mol Cell Biol, 1985 Apr, 5(4), 610 - 8 Expression of the Escherichia coli dam methylase in Saccharomyces cerevisiae: effect of in vivo adenine methylation on genetic recombination and mutation; Hoekstra MF et al.; The Escherichia coli DNA adenine methylase (dam) gene has been introduced into Saccharomyces cerevisiae on a yeast-E . coli shuttle vector . Sau3AI, MboI, and DpnI restriction enzyme digests and Southern hybridization analysis indicated that the dam gene is expressed in yeast cells and methylates GATC sequences . Analysis of digests of total genomic DNA indicated that some GATC sites are not sensitive to methylation . The failure to methylate may reflect an inaccessibility to the methylase due to chromosome structure . The effects of this in vivo methylation on the processes of recombination and mutation in mitotic cells were determined . A small but definite general increase was found in the frequency of mitotic recombination . A similar increase was observed for reversion of some auxotrophic markers; other markers demonstrated a small decrease in mutation frequency . The effects on mutation appear to be locus (or allele) specific . Recombination in meiotic cells was measured and was not detectably altered by the presence of 6-methyladenine in GATC sequences. Proc Natl Acad Sci U S A, 1985 Apr, 82(7), 2106 - 10 Construction of telocentric chromosomes in Saccharomyces cerevisiae; Surosky RT et al.; We describe a simple method for the construction of large chromosomal deletions in yeast . Diploid yeast cells were transformed with DNA fragments that replace large regions of the chromosomes by homologous recombination . Using this method, we have constructed a telocentric chromosome III in which approximately equal to 100 kilobases (kb) of DNA has been removed from the left arm of the chromosome, so that the centromere is 12 kb from the left telomere . This telocentric chromosome is mitotically stable . Its rate of loss in a diploid strain is 2.5-7.4 X 10(-4) per cell division compared to a rate of loss of 0.36-1.8 X 10(-4) per cell division for a normal chromosome III . It also segregates 2+:2- with fidelity during meiosis . The construction of systematic deletions in a chromosome should be useful in determining the essential features for proper chromosomal segregation and replication. Proc Natl Acad Sci U S A, 1985 Apr, 82(7), 2019 - 23 Influenza viral (A/WSN/33) hemagglutinin is expressed and glycosylated in the yeast Saccharomyces cerevisiae; Jabbar MA et al.; Recombinant plasmids were constructed in which genes coding for either the entire or the signal-minus (amino acid residues 2-17 deleted) hemagglutinin (HA) of WSN influenza virus were placed under the control of the alcohol dehydrogenase I gene promoter of Saccharomyces cerevisiae . Both recombinant plasmids were shown to direct the synthesis of HA-specific polypeptides that were detected by immunoprecipitation with antiviral antibodies . The complete HA produced in yeast had an approximate Mr of 70,000 and was glycosylated, as determined by the endoglycosidase H sensitivity, and was bound to membrane . Therefore, the complete HA polypeptide possessing the signal sequence probably traversed the yeast secretory pathways . Signal-minus HA, on the other hand, had a lower molecular weight and was nonglycosylated . The specific binding of yeast HA with antiviral antibodies could be competitively inhibited by influenza viral HA, demonstrating that the HA produced in yeast contained antigenic determinants of the native viral HA. J Biol Chem, 1985 Mar 25, 260(6), 3542 - 7 Phosphorolytic cleavage of diadenosine 5',5'''-P1,P4-tetraphosphate . Properties of homogeneous diadenosine 5',5'''-P1,P4-tetraphosphate alpha, beta-phosphorylase from Saccharomyces cerevisiae; Guranowski A et al.; Novel enzymatic activity which splits diadenosine 5',5'''-P1,P4-tetraphosphate (Ap4A) phosphorolytically has been found in extracts from Saccharomyces cerevisiae . One of the two alpha,beta-anhydride bonds between Ap4A phosphate residues undergoes phosphorolysis, and ATP (pppA) plus ADP (ppA) are the products of the reaction according to the equation: AppppA + P*i----pppA + p*pA The reaction is dependent on the presence of divalent metal ions; Mn2+ or Mg2+ sustain the greatest rates of reaction . Among analogues of the Ap4A substrate, Ap5A and Gp4G, but not p4A and Ap3A, are substrates, and corresponding products are p4A plus ADP, and GTP plus GDP, the phosphate being incorporated into the nucleoside 5'-diphosphates . In the reactions, phosphate can be substituted with arsenate . Arsenolysis of Ap4A, Ap5A, or Gp4G leads to ATP plus AMP, p4A plus AMP, and GTP plus GMP, respectively . The name diadenosine tetraphosphate alpha,beta-phosphorylase (ADP-forming) is proposed for the new enzyme . The phosphorylase has been purified to apparent homogeneity and behaves as a single polypeptide chain of Mr = 40,000 . Optimum activity of the enzyme is at pH 8.0 and the sulfhydryl groups are essential for catalysis . At saturating Ap4A, the rate constant for the reaction is 36 s-1 and the Km value for Ap4A is 60 microM (37 degrees C, 50 mM Hepes/KOH (pH 8.2), 500 microM MnCl2, 10 mM K2HPO4, 1 mM 2-mercaptoethanol, and 2% glycerol) . The Km values for phosphate and arsenate are 1 and 3 mM, respectively. J Biol Chem, 1985 Mar 10, 260(5), 3108 - 15 Transfer RNA splicing in Saccharomyces cerevisiae . Secondary and tertiary structures of the substrates; Lee MC et al.; Secondary and tertiary structures of four yeast tRNA precursors that contain introns have been elucidated using limited digestion with a variety of single-strand- and double-strand-specific nucleases . The pre-tRNAs, representing the variety of intron sizes and potential structures, were: pre-tRNALeuCAA, pre-tRNALeuUAG, pre-tRNAIleUAU, and pre-tRNAPro-4UGG . Conventional tRNA cloverleaf structure is maintained in these precursors except that the anticodon loop is interrupted by the intron . The intron contains a sequence which is complementary to a portion of the anticodon loop and allows the formation of a double helix often extending the anticodon stem . The 5' and 3' splicing cleavage sites are located at either end of this helix and are single-stranded . The intron is the most sensitive region to nuclease cleavage, suggesting that it is on the surface of the molecule and available for interaction with the splicing endonuclease . Absence of Mg2+ or spermidine renders the dihydrouridine and T psi C loops of these precursors highly sensitive to nuclease digestion . These ionic effects mimic those observed for tRNAPhe and suggest that the tRNA portion of these precursors has native tRNA structure . We propose consensus secondary and tertiary structures which may be of significance to eventual understanding of the mechanism of yeast tRNA splicing. J Biol Chem, 1985 Mar 10, 260(5), 3084 - 9 Elongation factor 1 alpha from Saccharomyces cerevisiae . Rapid large-scale purification and molecular characterization; Thiele D et al.; Cytoplasmic elongation factor 1 alpha (EF-1 alpha) was purified to homogeneity from the yeast Saccharomyces cerevisiae using a large-scale procedure . The three steps of purification used were batch adsorption on phosphocellulose, phosphocellulose chromatography and, as the last step, GDP-Sepharose or Biorex column chromatography . The protein is very basic (pI = 9.2) and has an apparent molecular mass of 49 kDa, as determined by polyacrylamide gel electrophoresis using denaturing conditions . It is one of the most abundant proteins in yeast (about 5% of total soluble protein), as shown by two-dimensional gel electrophoresis and by immunological titration . A strong immunological and structural homology was found between yeast EF-1 alpha and elongation factors from other sources . Common immunological features were found between yeast and wheat germ EF-1 alpha . Tryptic hydrolysis of yeast EF-1 alpha in the presence of 25% glycerol generated a large trypsin-resistant polypeptide (Mr = 43,000) which had the same NH2-terminal sequence as the proteolyzed product from rabbit reticulocyte, Artemia salina EF-1 alpha and Escherichia coli EF-Tu . Completed DNA sequence determination of one structural gene for yeast EF-1 alpha confirmed a remarkable conservation of several protein sequence domains in yeast and animal EF-1 alpha (Cottrelle, P., Thiele, D., Price, V., Memet, S., Micouin, J.Y., Marck, C., Buhler, J.M . Sentenac, A., and Fromageot, P . (1985) J . Biol . Chem . 260, 3090-3096). Genetics, 1985 Mar, 109(3), 481 - 92 Mutations leading to expression of the cryptic HMRa locus in the yeast Saccharomyces cerevisiae; Kassir Y et al.; Mutations leading to expression of the silent HMRa information in Saccharomyces cerevisiae result in sporulation proficiency in mata1/MAT alpha diploids . An example of such a mutation is sir5-2, a recessive mutation in the gene SIR5 . As expected, haploids carrying the sir5-2 mutation are nonmaters due to the simultaneous expression of HMRa and HML alpha, resulting in the nonmating phenotype of an a/alpha diploid . However, sir5-2/sir5-2 mata1/MAT alpha diploids mate as alpha yet are capable of sporulation . The sir5-2 mutation is unlinked to sir1-1, yet the two mutations do not complement each other: mata1/MAT alpha sir5-2/SIR5 SIR1/sir1-1 diploids are capable of sporulation . In this case, recessive mutations in two unlinked genes form a mutant phenotype, in spite of the presence of the normal wild-type alleles . The PAS1-1 mutation, Provider of a Sporulation function, is a dominant mutation tightly linked to HMRa . PAS1-1 does not affect the mating ability of a strain, yet it allows diploids lacking a functional MATa locus to sporulate . It is proposed that PAS1-1 leads to partial expression of the otherwise cryptic a1 information at HMRa. Arch Biochem Biophys, 1985 Mar, 237(2), 292 - 9 Extraction of proteins from Saccharomyces cerevisiae ribosomes under nondenaturing conditions; Lee JC et al.; The differential sensitivity of ribosomal proteins to removal by salts has been studied . Proteins were extracted from the large and small subunits of cytoplasmic ribosomes from Saccharomyces cerevisiae by washing the individual subunits with a series of solutions containing increasing concentrations of NH4Cl (0.74-3.56 M) for a defined time (20 min) at 0 degrees C . The molar ratio of magnesium to ammonium ions of 1:40 was maintained to protect the ribosomal subparticles from complete disassembly . Proteins extracted under each salt condition were analyzed for composition by two-dimensional polyacrylamide gel electrophoresis . The relative quantity of each protein was determined . Most proteins were not removed from the ribosomal particle completely by any one condition, but were preferentially enriched in a single fraction . Whereas most proteins could be solubilized, several proteins remained predominantly or exclusively with the final core particle . The kinetics of protein release from both subunits at a single NH4Cl concentration (0.74 M) were also studied . Release of protein was time dependent, i.e., longer extraction generally removed more of the same proteins . However, prolonged treatment (240 min) of subunits, even at the same salt concentration, resulted in removal of additional species of proteins in varying amounts . Among the ribosomal RNA species, only the 5 S RNA species was released from the ribosomal particles upon treatment. Eur J Biochem, 1985 Mar 1, 147(2), 273 - 9 Intracellular maturation and secretion of acid phosphatase of Saccharomyces cerevisiae; Schonholzer F et al.; To elucidate intracellular maturation and secretion of acid phosphatase of Saccharomyces cerevisiae we prepared a monoclonal antibody that recognizes specifically the protein moiety of this cell surface glycoprotein . With this antibody membranes and soluble fractions of wild-type cells, grown in low-phosphate medium in the presence and absence of tunicamycin, were examined by the immunoblot technique . Similarly, secretory mutants, blocked at distinct steps in the secretory pathway at the restrictive temperature as well as a strain harboring several copies of the structural gene PHO5 for repressible acid phosphatase, were analyzed . The data suggest the following sequence of events in acid phosphatase maturation and secretion: three unglycosylated precursors with molecular masses of 60 kDa, 58 kDa and 56 kDa are synthesized into membranes of the endoplasmic reticulum, where these are core glycosylated in a membrane-bound form . They appear on sodium dodecyl sulfate gels as bands with molecular masses of 76 kDa and 80 kDa . Owing to a rate-limiting maturation step, occurring after core glycosylation, they can accumulate in a membrane-bound form . At the Golgi apparatus outer carbohydrate chains are attached to the core and the enzyme appears in a soluble form, indicating a release of acid phosphatase from the membrane between the endoplasmic reticulum and the Golgi . Pulse-chase experiments suggest that the time for acid phosphatase synthesis and its transport to the Golgi is about 5 min. J Bacteriol, 1985 Mar, 161(3), 831 - 5 Rapid method for isolation and screening of cytochrome c oxidase-deficient mutants of Saccharomyces cerevisiae; McEwen JE et al.; We describe here a new method for the specific isolation of cytochrome c oxidase-deficient mutants of Saccharomyces cerevisiae . One unique feature of the method is the use of tetramethyl-p-phenylenediamine as a cytochrome c oxidase activity stain for yeast colonies . The staining of yeast colonies by tetramethyl-p-phenylenediamine is dependent upon a functional cytochrome c oxidase and is unaffected by other lesions in respiration . Since the tetramethyl-p-phenylenediamine colony staining reaction is rapid and simple, it greatly facilitates both the identification and characterization of cytochrome c oxidase-deficient mutants . Another feature of the method, which is made possible by the tetramethyl-p-phenylenediamine colony stain, is the use of an op1 parent strain for the isolation of nuclear pet or mitochondrial mit mutants in specific protein-coding genes . A parent strain that carries this marker selects against rho0 or rho- classes of pleiotropic respiratory-deficient mutants, since these are lethal in op1 strains . We have used this method to isolate 123 independently derived cytochrome c oxidase-deficient pet mutants and 300 independently derived mit mutants. J Biol Chem, 1985 Feb 25, 260(4), 2253 - 7 Characterization of a specific alpha-mannosidase involved in oligosaccharide processing in Saccharomyces cerevisiae; Jelinek-Kelly S et al.; Fractionation of a crude extract from Saccharomyces cerevisiae X-2180 on Sepharose 6B in the presence of 0.5% Triton X-100 resolves two enzyme fractions containing alpha-mannosidase activity . Fraction I which is excluded from the gel contains alpha-mannosidase activity toward both p-nitrophenyl-alpha-D-mannopyranoside and Man9GlcNAc oligosaccharide as substrates, whereas Fraction II which is included in the gel contains only oligosaccharide alpha-mannosidase activity . The latter enzyme is very specific and removes a single mannose residue from Man9GlcNAc, whereas the alpha-mannosidase activity of Fraction I removes several mannose residues from Man9GlcNAc oligosaccharide . High resolution 1H NMR analysis of the Man8GlcNAc formed from Man9GlcNAc in the presence of the alpha-mannosidase of Fraction II showed only a single isomer with the following structure: (see formula; see text) This specific enzyme is most probably involved in processing of oligosaccharide during biosynthesis of mannoproteins . The mannose analog of 1-deoxynojirimycin (50-500 microM), dideoxy-1,5-imino-D-mannitol, inhibits the oligosaccharide alpha-mannosidase activities of Fractions I and II to about the same extent, but has no effect on the nonspecific alpha-mannosidase which acts on p-nitrophenyl-alpha-D-mannopyranoside. Nucleic Acids Res, 1985 Feb 25, 13(4), 1327 - 39 Biogenesis of mitochondria: DNA sequence analysis of mit- mutations in the mitochondrial oli1 gene coding for mitochondrial ATPase subunit 9 in Saccharomyces cerevisiae; Ooi BG et al.; The nucleotide sequence of the yeast mitochondrial olil gene has been obtained in a series of mit- mutants with mutations in this gene, which codes for subunit 9 of of the mitochondrial ATPase complex . Subunit 9 is the proteolipid, 76 amino acids in length, necessary for the proton translocation function of the membrane Fo-sector . These mutants were classified on the basis of their rescue by a petite strain shown here to retain the entire wild-type olil gene . The mutation in one mit- strain removes a positively charged residue (Arg39----Met) which is likely to be located in a segment of subunit 9 that protrudes from the inner mitochondrial membrane . In a second mit- mutant, a negatively charged residue replaces a conserved glycine residue (Gly18----Asp) in a glycine-rich segment of the protein that is most likely embedded within the membrane . Other mit- mutations result in frameshifts with predicted products 7, 65 and 68 amino acid residues long . In each mit- mutant, there is the loss of one or more of the amino acid residues that are highly conserved among diverse species . The location and nature of specific changes pinpoint amino acid residues in subunit 9 essential to the activity of the mitochondrial ATPase complex. Eur J Biochem, 1985 Feb 15, 147(1), 191 - 6 Mitochondrial transcription and processing of transcripts during release from glucose repression in 'resting cells' of Saccharomyces cerevisiae; Zennaro E et al.; Mitochondrial transcription and processing of transcripts have been investigated at different stages of release from glucose repression in resting cells of Saccharomyces cerevisiae . Transcripts were identified by hybridization with nick-translated or terminally labelled gene-specific probes . This allowed the determination of the steady-state levels of individual transcripts in the mitochondrial RNA population . Results showed different gene-specific patterns of response to respiratory induction: no increase in the level of transcripts (oxi2); a rapid increase in the steady-state levels of all transcripts (cob); a very strong increase in the processing of the high-molecular-mass precursors (oxi3 and oli2); an increase in the level of stable circular transcripts (oxi3) . As a whole the results indicate specific and differentiated effects of release from glucose repression on the expression of the different mitochondrial genes and demonstrate the importance of processing events in mitochondrial regulation. Mol Gen Mikrobiol Virusol, 1985 Feb, (2), 33 - 5 {Gene transfer using fusion of isolated cell nuclei with protoplasts of the yeast Saccharomyces cerevisiae}; Becher D et al.; Hybrid clones of Saccharomyces cerevisiae with different genotypes have been obtained by polyethyleneglycol induced fusion of isolated cellular nuclei with protoplasts . The genetic instability of complete nuclei after fusion results in formation of different genotypes. Int J Pept Protein Res, 1985 Feb, 25(2), 187 - 96 Synthesis and biological activity of N epsilon-acyl derivatives of a Saccharomyces cerevisiae mating pheromone; Shenbagamurthi P et al.; We report a general method for acylation of the N epsilon-amino group of the lysyl residue in peptides . The procedure involves acylation using p-nitrophenyl esters and 1-hydroxybenzotriazole in organic solvents to yield a series of fatty acyl mating pheromones of Saccharomyces cerevisiae . The fatty acyl group does not influence coupling of peptide fragments . Biological activities of the synthesized alpha-factor mating pheromones derivatized with acetyl, butyryl, caprylyl and lauryl groups are nearly equivalent to the activity of unacylated alpha-factor . The N epsilon-stearyl-alpha-factor is biologically inactive . The procedures reported in this communication can be used to increase hydrophobicity of lysine-containing peptides when the lysyl group is not essential for activity. Can J Microbiol, 1985 Feb, 31(2), 109 - 18 Ultrastructure of Saccharomyces cerevisiae strain AG1-7 and its responses to changes in environment; Willison JH et al.; Asynchronous populations of the budding yeast Saccharomyces cerevisiae strain AG1-7 were examined by freeze-fracture electron microscopy for ultrastructural changes occurring in response to changes in the environment, specifically the following: temperature (23 or 37 degrees C); cell density (exponential, early stationary, and stationary phases); various periods of nitrogen starvation at low cell density, and return of nitrogen-starved cells to nitrogen-replete medium . This information has been gathered in preparation for ultrastructural examination of comparable responses of temperature-sensitive cell-cycle mutants . The plasma membrane was found to be particularly responsive to changes in environment . A high proportion (75%) of cells in exponential phase populations at 37 degrees C displayed paracrystalline arrays of plasma membrane particles, whereas this proportion was much lower (20%) at 23 degrees C in the same medium; plasma membrane grooves were longer at 37 than at 23 degrees C . In budded cells, the mother cell displayed paracrystalline arrays more frequently than the bud . Entry of cells into stationary phase, either through permitting population growth or by limiting nitrogen supply, resulted in increases in numbers of paracrystalline arrays and grooves . Groove depth also increased . The paracrystalline-array and groove-density responses were independent, both during entry into stationary phase and during the subsequent lag phase . Unusual groove forms appeared during stationary phase in high cell density populations, but not in low cell density nitrogen-starved populations . "Aggregate" and "geometric" tonoplast forms, previously described in strain A364A when grown under some of the conditions used here, were not found in AG1-7 under any of the conditions used here . It was demonstrated that particle-free patches can arise rapidly on the tonoplast of AG1-7 in response to temperature change from 37 to 23 degrees C . During stationary phase, spherosomes (lipid droplets) increased in size, particularly in response to nitrogen depletion . After 72 h of nitrogen starvation, about 10% of cell volume consisted of spherosomes . Changes in vacuolar content and mitochondrial form were also noted during entry into stationary phase. Genetics, 1985 Feb, 109(2), 303 - 32 Meiotic recombination between duplicated genetic elements in Saccharomyces cerevisiae; Jackson JA et al.; We have studied the meiotic recombination behavior of strains carrying two types of duplications of an 18.6-kilobase HIS4 Bam HI fragment . The first type is a direct duplication of the HIS4 Bam HI fragment in which the repeated sequences are separated by Escherichia coli plasmid sequences . The second type, a tandem duplication, has no sequences intervening between the repeated yeast DNA . The HIS4 genes in each region were marked genetically so that recombination events between the duplicated segments could be identified . Meiotic progeny of the strains carrying the duplication were analyzed genetically and biochemically to determine the types of recombination events that had occurred . Analysis of the direct vs . tandem duplication suggests that the E . coli plasmid sequences are recombinogenic in yeast when homozygous . In both types of duplications recombination between the duplicated HIS4 regions occurs at high frequency and involves predominantly interchromosomal reciprocal exchanges (equal and unequal crossovers) . The striking observation is that intrachromosomal reciprocal recombination is very rare in comparison with interchromosomal reciprocal recombination . However, intrachromosomal gene conversion occurs at about the same frequency as interchromosomal gene conversion . Reciprocal recombination events between regions on the same chromatid are the most infrequent exchanges . These data suggest that intrachromosomal reciprocal exchanges are suppressed. J Bacteriol, 1985 Feb, 161(2), 778 - 80 Interaction of UAG suppressors and omnipotent suppressors in Saccharomyces cerevisiae; Song JM et al.; Haploids bearing the dominant UAG suppressor, SUP7-a, and various alleles of the omnipotent suppressor sup35 were examined . The presence of the UAG suppressor reduced the efficiency of some alleles of sup35, and caused other sup35 alleles to be lethal . A nonclassical interaction of the dominant suppressor tRNA and the ribosome is proposed to explain these observations. J Bacteriol, 1985 Feb, 161(2), 769 - 71 Expression of a Saccharomyces cerevisiae photolyase gene in Escherichia coli; Sancar GB; A 3.3-kilobase PvuII fragment carrying the PHR1 gene of Saccharomyces cerevisiae has been cloned into an Escherichia coli expression vector and introduced into E . coli strains deficient in DNA photolyase . Complementation of the E . coli phr-1 mutation was observed, strongly suggesting that the yeast PHR1 gene encodes a DNA photolyase. J Bacteriol, 1985 Feb, 161(2), 596 - 601 Alpha mating type-specific expression of mutations leading to constitutive agglutinability in Saccharomyces cerevisiae; Doi S et al.; Two mutants of Saccharomyces cerevisiae have been isolated and characterized . The mutants were constitutively agglutinable at 36 degrees C, the temperature at which wild-type cells agglutinate only after induction by mating pheromone . The mutant cells had other properties specific for the normal alpha cell type, i.e., conjugation with a cells, response to a mating pheromone, and production of alpha mating pheromone . The two mutations, cag1 and cag2, were recessive and expressed only in alpha cells . cag1 is linked very closely to the MAT locus, but cag2 is unlinked to the MAT locus . These cag mutations complemented ste3-1 . These results indicate that CAG genes are novel alpha-specific genes involved in the regulation of sex agglutinin synthesis. Biochemistry, 1985 Jan 29, 24(3), 753 - 9 Saccharomyces cerevisiae structural cell wall mannoprotein; Frevert J et al.; A novel mannoprotein fraction with an average molecular weight of 180 000 has been isolated from Saccharomyces cerevisiae mnn9 mutant cell wall that was solubilized by beta-glucanase digestion . The same material could be extracted from purified wall fragments with 1% sodium dodecyl sulfate . The protein component, 12% by weight, is rich in proline, whereas the carbohydrate, mainly mannose, is about evenly distributed between asparagine and hydroxyamino acids . Endoglucosaminidase H digestion of the isolated mannoprotein reduced its average molecular weight to 150 000, but the mannoprotein, while still embedded in the cell wall, was inaccessible to the enzyme . Biosynthesis and translocation of the mannoprotein were investigated by following incorporation of {3H}proline into this fraction . In the presence of tunicamycin, both mnn9 and wild-type X2180 cells made a mannoprotein fraction with an average molecular weight of 140 000, whereas in the absence of the glycosylation inhibitor, the mnn9 mutant made material with a molecular weight of 180 000 and the mannoprotein made by wild-type cells was too large to penetrate the polyacrylamide gel . Although the cell wall mannoprotein was resistant to heat and proteolytic enzymes, attempts to isolate the carbohydrate-free component failed to yield any characteristic peptide material. J Biol Chem, 1985 Jan 25, 260(2), 1271 - 9 Nucleolytic processing of a tRNAArg-tRNAAsp dimeric precursor by a homologous component from Saccharomyces cerevisiae; Engelke DR et al.; A subcellular extract from Saccharomyces cerevisiae has been used to transcribe cloned yeast tRNA genes in vitro and to process the primary transcripts at the 5' and 3' termini . Chromatographic fractionation of the extract has separated the transcription components from two distinct nucleolytic activities: an endonuclease that cleaves the precursors to produce mature 5' termini; and a 3'-5' exonuclease . These fractions have been used to elaborate a processing pathway for the dimeric primary transcript of the yeast tRNAArg-tRNAAsp gene pair . Under optimal conditions in vitro this gene is expressed at a rate of 200 transcripts/gene/hour, initiating at position -10 with respect to the mature 5' terminus of tRNAArg and terminating near position +160 . The primary transcripts are cleaved by an endonuclease to give tRNAAsp with a mature 5' terminus, and a pre-tRNAArg monomer with a 5' leader and 3' trailer sequences . A second endonuclease cleavage of pre-tRNAArg generates the mature 5' terminus of tRNAArg . The endonuclease cleavages are not ordered . Exonuclease activity(ies) remove the spacer sequences from the 5' mature tRNAArg, and trim the 3' trailer portion from tRNAAsp . Exonucleolytic removal of the 3' trailer does not require prior endonuclease action, but removal of the spacer sequences from pre-tRNAArg is incomplete without prior removal of the 5' leader sequences. FEBS Lett, 1985 Jan 7, 179(2), 307 - 10 An antisuppressor mutant of Saccharomyces cerevisiae deficient in isopentenylated tRNA has reduced delta 2-isopentenylpyrophosphate: tRNA-delta 2-isopentenyl transferase activity; Laten HM et al.; We have previously reported the isolation and initial characterization of a mutation in Saccharomyces cerevisiae, designated mod5-1, that reduces the capacity of altered tyrosine tRNAs to suppress ochre nonsense mutations . The mutation results in the virtual elimination of the modified tRNA nucleoside, N6-delta 2-(isopentenyl) adenosine, normally found adjacent to the anticodons of certain tRNA species . We demonstrate here that MOD5 codes for delta 2-isopentenylpyrophosphate: tRNA-delta 2-isopentenyl transferase, or a protein that regulates its synthesis. Eur J Biochem, 1985 Jan 2, 146(1), 95 - 100 Purification and characterization of the indole-3-glycerolphosphate synthase/anthranilate synthase complex of Saccharomyces cerevisiae; Prantl F et al.; The indole-3-glycerolphosphate synthase/anthranilate synthase complex from Saccharomyces cerevisiae was purified to apparent homogeneity . The native complex with Mr approximately equal to 130 000 consists of two different subunits, the TRP2 gene product with Mr = 64 000 and the TRP3 gene product with Mr = 58 000 . The larger polypeptide was identified as anthranilate synthase and is active in vitro with ammonia as cosubstrate without need of complex formation . The smaller polypeptide carries both glutamine amidotransferase activity and indole-3-glycerolphosphate synthase activity . Various steady-state kinetic parameters as well as the amino acid composition of the two polypeptides were determined. Curr Genet, 1985, 10(1), 35 - 7 Suppression of temperature sensitive mutations in oncogene-related CDC genes in Saccharomyces cerevisiae by catabolite repression resistance and cytoplasmic petite mutations; Egilsson V et al.; The "start" cell division control genes CDC36 and CDC28 have been reported to contain a certain sequence homology to tissue oncogenes (ets and some protein kinase encoding oncogenes respectively) . Here we report that temperature sensitive mutations in these genes are suppressed in cytoplasmic "petite" mutants and catabolite repression resistant mutants. Curr Genet, 1985, 10(1), 21 - 7 Maintenance and copy number control of ARS1 plasmids in Saccharomyces cerevisiae . Evidence of a mating type effect; Gullov K et al.; Following mating of a and alpha isogenic haploids we observe that the frequency of plasmid bearing cells, during selective growth, increases three fold . By examining the mitotic stability, the frequency of plasmid bearing cells during the cell cycle and the copy number of ARS1 plasmids in isogenic haploid and diploid cells, we show that the apparent stability of circular ARS1 plasmids in a/alpha cells is largely due to a diminished copy number in these cells . This observation is fully comprehensible with the model for plasmid segregation as presented by Murray and Szostak (1983) . In order to account for the differences in copy numbers, alpha and alpha/alpha isogenic strains were compared . Likewise a number of mating type nonspecific sterile mutants were compared with the parental Ste+ strain . It seems that a diminished copy number is established when the MATa1/MAT alpha 2 regulatory system (Klar et al . 1981) is switched on, since the effect is observed in Sir- strains only. Curr Genet, 1985, 10(1), 15 - 20 A mutant of Saccharomyces cerevisiae with impaired maintenance of centromeric plasmids; Larionov VL et al.; A mutant with unstable maintenance of hybrid plasmids containing either one of the centromeric loci CEN3, CEN6, CEN11 and ars1 or the replicator of the 2 mu plasmid has been obtained . The frequency of loss of hybrid plasmids in the mutant was up to 3.10(-1) per one generation versus 10(-2) in the original strain . The unstable maintenance of minichromosomes in the mutant is controlled by a recessive nuclear gene, named SMC for stability of minichromosomes . Loss of some minichromosomes is connected with impairment of their segregation in cell division . In diploids homozygous for smc mitotic chromosomal segregation is not affected but sporulation is impaired . The question of adequacy of usage of minichromosomes for selection of mutants with impaired function of centromeric loci is discussed. Microbiol Sci, 1985 Jan, 2(1), 10 - 3 Saccharomyces cerevisiae as a model eukaryote for studies on mitochondriogenesis; Trivedi A et al.; A great deal of progress has been made in elucidating the underlying mechanisms which control the interplay between the nuclear and mitochondrial genomes during biogenesis of mitochondria . The advantage of using the yeast Saccharomyces cerevisiae in these studies over other eukaryotic cells will be discussed. Curr Genet, 1985, 9(7), 529 - 32 UV-inducible proteins in Saccharomyces cerevisiae; Rolfe M; Two UV-inducible proteins have been detected in the yeast, Saccharomyces cerevisiae . The proteins have molecular weights of 78,000 Daltons and 23,000 Daltons . This induction is specific for UV-irradiation as exposure to X-rays, mitomycin C and heat shock does not result in the synthesis of the proteins . The larger (78 kD) protein is induced in various rad strains and in a rho degree cir degree strain . Attempts are being made to isolate the genes coding for these inducible proteins. Curr Genet, 1985, 9(5), 341 - 4 General and specific controls of lysine biosynthesis in Saccharomyces cerevisiae; Urrestarazu LA et al.; Six of the eight enzymes of the alpha-aminoadipate pathway for the biosynthesis of lysine in Saccharomyces cerevisiae were examined for repressibility to lysine and for susceptibility to the general control of amino acid biosynthesis . All of the enzymes exhibited a 2 to 4 fold lower level of specific activity in the wildtype strain X2180 when grown in lysine supplemented medium as compared to minimal medium . However, levels of only three of the enzymes, alpha-aminoadipate reductase, saccharopine reductase, and saccharopine dehydrogenase, were derepressed in the leaky lysine mutant 7305d and leaky arginine mutant 7853-6c when grown in minimal medium . These observations are characteristic of enzymes under general control of amino acid biosynthesis . The remaining three enzymes, homocitrate synthetase, homoaconitase and homoisocitrate dehydrogenase were repressed in 7305d cells grown in minimal or lysine supplemented medium. Mol Gen Genet, 1985, 200(2), 291 - 4 Mutation affecting the specific regulatory control of lysine biosynthetic enzymes in Saccharomyces cerevisiae; Ramos F et al.; A Saccharomyces cerevisiae mutant which exhibits a considerably increased cellular lysine pool has been isolated and characterized . Assay of enzymes of the lysine and arginine pathways shows that the mutation harboured by this mutant alters the specific repression of lysine but does not influence the general control of amino acid biosynthesis . Because it is recessive to the wild-type allele and acts pleiotropically on the synthesis of several lysine pathway enzymes, this regulatory mutation has been denominated lys80-1 (or lysR--1) . It is believed to affect the synthesis or the structure of a factor which plays a negative role in the control of LYS gene expression. Mol Gen Genet, 1985, 199(1), 59 - 63 Expression of the RAD1 and RAD3 genes of Saccharomyces cerevisiae is not affected by DNA damage or during the cell division cycle; Nagpal ML et al.; The RAD1 and RAD3 genes of Saccharomyces cerevisiae are required for excision repair of UV damaged DNA . In addition, the RAD3 gene is essential since rad3 deletions are recessive lethals . We have examined the induction of the RAD1 and RAD3 genes by DNA damage and during the cell division cycle . We have made fusions of the RAD1 and RAD3 genes with the Escherichia coli lacZ gene encoding beta-galactosidase . Beta-galactosidase activity was measured in a Rad+ yeast strain containing the RAD1-lacZ or the RAD3-lacZ fusion, either in a multicopy replicating plasmid or as a single copy integrant resulting from transformation with an integrating plasmid which transforms yeast by homologous recombination in the yeast genome . No induction of beta-galactosidase activity occurred after ultraviolet light (UV) or 4-nitroquinoline-1-oxide (NQO) treatment . Haploid cells of mating type a were synchronized by treatment with alpha factor and beta-galactosidase activity was determined during different cell cycle stages . No change in beta-galactosidase activity was observed in the strain containing the RAD1-lacZ or the RAD3-lacZ fusion integrated in the yeast genome. Curr Genet, 1985, 10(3), 187 - 95 Saccharomyces cerevisiae strains sensitive to inorganic mercury . II . Effect of glucose; Sakamoto E et al.; Saccharomyces cerevisiae strains sensitive to inorganic mercury (Ono and Sakamoto 1985) did not grow well on the medium rich in glucose and poor in peptone . This growth inhibition, like growth inhibition caused by inorganic mercury, was relieved by exogenous tyrosine . Sugars such as fructose and mannose were as inhibitory as glucose, but glycerol was not at all . Galactose was inhibitory but not so much as glucose . A gal2 mutation (defective in galactose uptake) partly relieved growth inhibition caused by excess galactose . Moreover, it was found that some of revertants which gained ability to grow well in the presence of excess glucose were defective in the glucose uptake . From these observations, we conclude that growth inhibition of the inorganic mercury sensitive strains by excess sugar is a consequence of the catabolite regulation . In other words, the inorganic mercury sensitive strains are hyper-sensitive to the catabolite regulation due to the presence of the HGS2-1 allele. Curr Genet, 1985, 9(7), 533 - 8 UV-inducible transcripts in Saccharomyces cerevisiae; Rolfe M; Differential colony hybridisation has been used to identify DNA sequences in Saccharomyces cerevisiae corresponding to RNA transcripts whose levels increase 5-10 fold following UV-irradiation . Four sequences have been identified, three of which share sequence homology and hybridize to the same set of genomic DNA fragments . The fourth sequence appears to be distinct, however each DNA sequence hybridizes to a similar sized RNA transcript which is approximately 4.0 kb long . The relationships between these DNA sequences and their potential protein products is discussed. Curr Genet, 1985, 9(4), 253 - 7 Cloning and mapping of CDC40, a Saccharomyces cerevisiae gene with a role in DNA repair; Kassir Y et al.; The cdc40 mutation has been previously shown to be a heat-sensitive cell-division-cycle mutation . At the restrictive temperature, cdc40 cells arrest at the end of DNA replication, but retain sensitivity to hydroxyurea (Kassir and Simchen 1978) . The mutation has also been shown to affect commitment to meiotic recombination and its realization . Here we show that mutant cells are extremely sensitive to Methyl-Methane Sulfonate (MMS) when the treatment is carried out at restrictive temperature . Incubation at 37 degrees C prior to, or after MMS treatment at 23 degrees C, does not result in lower survival . It is concluded that the CDC40 gene product has a role in DNA repair, possibly holding together or protecting the DNA during the early stages of repair . The CDC40 gene was cloned on a 2.65 kb DNA fragment . A 2 mu plasmid carrying the gene was integrated and mapped to chromosome IV, between trp4 and ade8, by the method of marker loss . Conventional tetrad analysis has shown cdc40 to map 1.7 cM from trp4. Basic Life Sci, 1985, 36, 231 - 42 Characterization of a tightly centromere-linked gene essential for meiosis in the yeast Saccharomyces cerevisiae; Yeh E et al.; The centromere region in the yeast Saccharomyces cerevisiae is characterized by short DNA fragments, less than 1,000 bp in length, that are capable of stabilizing entire chromosomes throughout mitotic and meiotic cell divisions . The CEN fragments are organized in a unique chromatin structure and are surrounded by ordered arrays of nucleosomal subunits . RNA transcripts are found 200-300 bp from the centromere, and lie within this ordered chromatin array . No transcripts have been detected through the centromere itself . We have examined the expression and cellular function of a tightly centromere-linked transcript on chromosome 11, (CEN11)L . The (CEN11)L transcript is present at constitutive levels throughout the mitotic and meiotic cell cycles . Disruption of the coding sequences in vivo has no effect on cell viability or mitotic growth, but the cells are unable to sporulate . Genetic complementation with known mutants in sporulation (spo10, spo13) has defined (CEN11)L as a new locus that appears to be required during both meiotic segregation divisions. Folia Microbiol (Praha), 1985, 30(6), 501 - 5 Effect of clomiphene on the content of sterols and fatty acids in Saccharomyces cerevisiae; Rezanka T et al.; During cultivation of Saccharomyces cerevisiae clomiphene regulates both quantitative and qualitative production of sterols and fatty acids as identified by gas chromatography and mass spectrometry . The content of sterols decreases to 75%, the production of fatty acids is comparable with that in the control . The occurrence of sterols increases; sterols with methyl group in position 4, without double bond in position 22 and with double bond in position 24(25) or 24(28) predominate . Among fatty acids shorter saturated and monoene acids are primarily produced, 2-hydroxy acids practically disappeared. Gene, 1985, 39(1), 95 - 101 Complete nucleotide sequence of the hexokinase PI gene (HXK1) of Saccharomyces cerevisiae; Kopetzki E et al.; The nucleotide sequence of the yeast glycolytic hexokinase isoenzyme PI-gene, HXK1, has been determined by sequencing the yeast DNA insert of the previously isolated plasmid HXK1 clone {Entian et al., Mol . Gen . Genet . 198 (1984) 50-54} . The structural gene sequence included 1452 bp coding for 484 amino acid (aa) residues corresponding to the Mr of 153 605 for the HXK1 monomer . Several initiation regions and termination points were located using nuclease S1 mapping . The HXK1 sequence was 76% homologous with that of HXK2, which is responsible for triggering glucose repression in yeasts . Since HXK1 is not involved in this regulatory system, the regulatory function of HXK2 must correspond to one or more of the differences between both isoenzymes . Most changes in the amino acid sequence were statistically distributed; however, four clustered regions with more than five altered aa residues were identified. Gene, 1985, 38(1-3), 205 - 14 The histidine permease gene (HIP1) of Saccharomyces cerevisiae; Tanaka J et al.; The histidine-specific permease gene (HIP1) of Saccharomyces cerevisiae has been mapped, cloned, and sequenced . The HIP1 gene maps to the right arm of chromosome VII, approx . 11 cM distal to the ADE3 gene . The gene was isolated as an 8.6-kb BamHI-Sau3A fragment by complementation of the histidine-specific permease deficiency in recipient yeast cells . We sequenced a 2.4-kb subfragment of this BamHI-Sau3A fragment containing the HIP1 gene and identified a 1596-bp open reading frame (ORF) . We confirmed the assignment of the 1596-bp ORF as the HIP1 coding sequence by sequencing a hip1 nonsense mutation . Analysis of the amino acid (aa) sequence of the HIP1 gene reveals several hydrophobic stretches, but shows no obvious N-terminal signal peptide . We have constructed a deletion of the HIP1 gene in vitro and replaced the wild-type copy of the gene with this deletion . The hip1 deletion mutant can grow when it is supplemented with 30 mM histidine, 50 times the amount required for the growth of HIP1 cells . Revertants of this deletion mutant able to grow on a normal level of histidine arise by mutation in unlinked genes . Both these observations suggest that there are additional, low-affinity pathways for histidine uptake. Mol Gen Genet, 1985, 201(1), 99 - 106 The use of plasmid DNA to probe DNA repair functions in the yeast Saccharomyces cerevisiae; White CI et al.; The survival of plasmid YRp12 treated in vitro with ultraviolet- or gamma-radiation, or with restriction endonucleases, has been used to investigate in vivo RAD gene activity in Saccharomyces cerevisiae . Yields of pyrimidine dimers or single and double strand breaks in plasmid DNA were assayed by physical methods . The biological effects of these damages were assayed by transformation of wild-type cells and rad mutants from each of the major groups of radiosensitive mutants . After UV-irradiation plasmid survival depended qualitatively on the same host functions that are needed for cellular survival . After gamma-irradiation no such correspondence was found . Apart from a RAD52-dependent stimulation of transformation efficiency at low doses, other host repair functions had little effect . Stimulation of transformation corresponded with the production of double- but not single-strand breaks in plasmid sequences homologous with the yeast genome and may be linked with a transient increase in mitotic stability . More generally these data also show that transformation events using the LiCl protocol may entail the uptake of a very low number of plasmid molecules per cell over a 10-fold range of DNA concentrations. Mol Gen Genet, 1985, 201(1), 1 - 6 Partial deprivation of GTP initiates meiosis and sporulation in Saccharomyces cerevisiae; Varma A et al.; We have investigated the physiological conditions under which meiosis and the ensuing sporulation of Saccharomyces cerevisiae are initiated . Initiation of sporulation occurs in response to carbon, nitrogen, phosphorus, or sulfur deprivation, and also, when met auxotrophs are partially starved for methionine, but not after starvation of other amino acid auxotrophs . It also occurs after partial starvation of pur or gua auxotrophs for guanine but not after starvation of ura auxotrophs for uracil . Under all these sporulation conditions the concentrations of both guanine nucleotides (GTP) and S-adenosylmethionine (SAM) decrease whereas those of other nucleotides show no trend . We show that the decrease of guanine nucleotides is essential for the initiation of meiosis and sporulation: when a gua auxotroph, also lacking one of the two SAM synthetases, is starved for guanine but supplemented with 0.1 mM methionine, GTP decreases while SAM slightly increases and yet the cells sporulate. Mol Gen Genet, 1985, 200(3), 458 - 62 An improved assay for DNA ligase reveals temperature-sensitive activity in cdc9 mutants of Saccharomyces cerevisiae; Barker DG et al.; A sensitive and quantitative assay for DNA ligase has been developed which is suitable for the analysis of crude cell extracts of yeast . The assay is sufficiently sensitive to detect the low levels of DNA ligase activity remaining in cdc9 mutants of Saccharomyces cerevisiae . Indeed, we have been able to show that this residual activity is temperature-sensitive, thus establishing finally that CDC9 is the structural gene for DNA ligase. Mol Gen Genet, 1985, 200(1), 74 - 9 Isolation of the catalase A gene of Saccharomyces cerevisiae by complementation of the cta1 mutation; Cohen G et al.; As a first step in an analysis of the DNA regions involved in the control of the catalase A gene of Saccharomyces cerevisiae by glucose, heme, and oxygen this gene has been cloned . Catalase A-deficient mutants were obtained by UV mutagenesis of a ctt1 mutant strain specifically lacking catalase T . All the catalase A-deficient mutants obtained fall into one complementation group . The single recessive mutation causing specific lack of catalase A was designated cta1 . Several overlapping DNA fragments complementing the cta1 mutation were obtained by transforming ctt1 cta1 double mutants with a yeast gene library in vector YEp13 . Hybrid selection of RNA with the help of one of the cloned DNAs followed by in vitro translation of this RNA and identification of the protein synthesized with catalase A-specific antibodies showed that the catalase A structural gene has been cloned . A single copy of this gene is present in the yeast genome . Transcription of the catalase A gene cloned into vector YEp13 is repressed by glucose . The DNA isolated hybridizes to a 1.6 kb polyA+-RNA virtually absent from heme-deficient cells, presumably catalase A mRNA. J Basic Microbiol, 1985, 25(3), 161 - 74 Electron microscopic study of purified polysaccharide components glucans and mannan of the cell walls in the yeast Saccharomyces cerevisiae; Kopecka M; In this study electron-microscopic characteristics of platinum shadowed or negatively stained preparations of purified beta-(1----3)-D-glucan, beta-(1----6)-D-glucan, and mannan were examined . These polysaccharides were isolated from cell walls of the yeast Saccharomyces cerevisiae . While purified samples of beta-(1----6)-D-glucan and mannan proved to be amorphous in structure and homogenous in appearance, the purified beta-(1----3)-D-glucan, isolated and presented to us as alkali-insoluble yeast glucan A2, was not homogenous . It consisted of (i) fibrillar component, (ii) amorphous matrix, and (iii) chitin bud scars . The ultrastructure of beta-(1----3)-D-glucan present in the glucan A2 sample did not change after treatment with 0.5 M acetic acid at 75 degrees C for 2 hours . After treatment with 1 M NaOH for 3 days at 4 degrees C scar material was removed by centrifugation and after a subsequent acidification of supernatant with acetic acid both the microfibrillar and the amorphous components were still present . It was concluded that beta-(1----3)-D-glucan component consists of molecules probably differing in their physico-chemical properties such as D . P., the degree of branching, conformation, and that cannot be separated by the methods currently used for their isolation. Gene, 1985, 34(1), 55 - 61 Molecular cloning of the RAD10 gene of Saccharomyces cerevisiae; Prakash L et al.; We have cloned the RAD10 gene of Saccharomyces cerevisiae and physically mapped it to a 1.0-kb DNA fragment . Strains containing disruptions of the RAD10 gene were found to show enhanced UV sensitivity compared with the previously characterized rad10-1 or rad10-2 mutants . The UV sensitivity of the disruption mutant is comparable to the highly UV sensitive rad1-19, rad2-delta, and rad3-2 mutants. Antonie Van Leeuwenhoek, 1985, 51(1), 1 - 10 Parental age and the life-span of zygotes of Saccharomyces cerevisiae; Muller I; Isolated cells of Saccharomyces cerevisiae were mated by micromanipulation and the reproductive capacity of the resulting zygotes was determined . The mating frequency was dependent on the age of the parents: conjugations between young cells and cells which had completed more than two thirds of their life-span were very rare events . The life-span of a zygote was very similar to the life-span of its shorter-lived parent . If one of the parent cells had budded several times prior to fusion, the life-span of the zygote was reduced correspondingly, i.e . there was no 'rescue by hybridization.' In four crosses the distribution of buds on both of the parent cells was recorded . In three of these four crosses the buds were evenly distributed, and in one the alpha-parent had three times as many buds as the a-parent. Microbios, 1985, 42(167), 37 - 44 Growth and adaptation of Saccharomyces cerevisiae at different cadmium concentrations; Minney SF et al.; The effect of 0, 5, 10 and 25 mg l-1 cadmium on the growth of Saccharomyces cerevisiae in defined medium has been investigated . It was found that the length of the lag phase increased with cadmium concentration and that metal uptake during the lag phase occurred only at a cadmium concentration of 25 mg l-1 . However, metal uptake occurred at all cadmium concentrations during the exponential phase . The yeast was gradually adapted to cadmium by a series of subcultures which resulted in a decrease in the length of the lag phase . Adaptation also caused a reduction in the cadmium uptake during the lag phase at 25 mg l-1 cadmium but did not affect uptake during the exponential phase at any concentration . A single passage through cadmium-free medium partially reversed the adaptation process . Sulphide production was enhanced significantly when the yeast was grown in the presence of increasing cadmium concentrations . However, at 5 mg l-1 cadmium, adapted cells produced less sulphide than unadapted cells, whilst at 10 and 25 mg l-1 cadmium the production of sulphide was similar for adapted and unadapted cells. Microbios, 1985, 42(167), 17 - 25 Biochemical and genetic analysis of an acatalasic rho+ mutant of Saccharomyces cerevisiae; Maqueda M et al.; The previously described acatalasic rho+ strain R-6 has been studied in order to determine the type of mutation responsible for its inability to produce catalase . Induction conditions for catalase activity and temperature influence on the behaviour of this strain were assayed . Furthermore, haploid progeny arising from crosses between R-6 and other strains (rho+, rho- and rho0) was examined for catalase levels . From the data obtained a hypothetical interaction between nuclear and mitochondrial genomes in relation to catalase biosynthesis is discussed. Mol Gen Genet, 1985, 199(1), 21 - 5 Cloning and mapping of the sporulation gene, spoT7, in Saccharomyces cerevisiae; Tanaka H et al.; In order to isolate a DNA fragment able to complement a sporulation-deficient mutation in Saccharomyces cerevisiae, a simple screening procedure was devised which was based on the difference in osmotic sensitivity between protoplasts and spores . A plasmid (pHT7) containing a 13 kb DNA insert that complemented the spoT7 mutation was isolated from a yeast genomic library prepared in the vector YEp13 . Gene spoT7 was linked to rna1 at 1.2 cM and to mak27 at 7.2 cM on the right arm of chromosome XIII . Mapping of the cloned gene following integration into the chromosome showed that the cloned gene was allelic to spoT7 and that a part of the RNA1 gene was also cloned into the same fragment . Gene spoT7 was localized on a 5 kb DNA fragment by further subcloning. Mol Gen Genet, 1985, 199(1), 14 - 20 A mutant plasmid with increased stability of holding and polymerization in Saccharomyces cerevisiae; Harashima S et al.; A yeast mutant plasmid, pX, showing increased stability of holding in mitotic and meiotic cell division, was isolated from an unstable plasmid, YRp7, which consists of pBR322 and a 1.4 kilobase (kb) fragment of Saccharomyces cerevisiae carrying the TRP1 gene and the autonomously replicating sequence, ARS1 . The pX plasmid exists as circular molecules in a series of polymeric forms from the monomer to the 20-mer or more, consisting, except for the monomer, of even numbers of unit molecules in tandem arrangement . The pX monomer consists solely of a yeast DNA fragment of 849 base pairs (bp) containing the TRP1 and ARS1 sequences derived from the 1.4 kb yeast fragment of YRp7 . The copy number of pX was calculated to be 20 as monomer units per genome . The ARS1 region was delimited to 80 bp by this mutation and the region essential for the ARS function was discussed. Gene, 1985, 33(2), 215 - 26 Factors affecting heterologous gene expression in Saccharomyces cerevisiae; Mellor J et al.; The 'promoter' fragment from the yeast phosphoglycerate kinase (PGK) gene has been used to direct the expression of human interferon-alpha-2 (IFN alpha 2) on a high-copy-number plasmid in yeast . The yields of IFN alpha 2 are only 1-3% of yeast total protein, whereas the maximum yield of PGK produced by the PGK gene on a high-copy-number plasmid is at least 50% . IFN alpha 2 is turned over more rapidly than PGK but in addition a major reason for the relatively low level of IFN alpha 2 is that IFN-specific RNA levels are much lower . This does not reflect differences in plasmid copy number or integrity, or differences in the 5' and 3' untranslated regions of the transcripts or DNA flanking regions . It appears that the presence of heterologous coding sequences, or the absence of specific yeast sequences causes a reduction in heterologous RNA levels in yeast. Gene, 1985, 33(2), 159 - 68 Transcription of the human adenovirus E1a gene in Saccharomyces cerevisiae; Handa H et al.; The early region 1a (E1a) and its flanking sequences of human adenovirus type 5 (Ad5) have been cloned in the yeast-Escherichia coli shuttle vector YEp13 and transferred into the yeast Saccharomyces cerevisiae . The E1a-specific RNAs were produced in the transformed yeast cells . The 5' ends of these transcripts were capped but were lacking 10 to 45 nucleotides from the 5' end of the proper E1a mRNA . These transcripts terminated approx . 1000 nucleotides downstream from the proper 3' end . No splicing of the E1a-specific RNA could be detected in the yeast cells. Basic Life Sci, 1985, 31, 425 - 34 Mutation induction by excess deoxyribonucleotides in Saccharomyces cerevisiae; Brendel M; Excess dTMP is toxic and mutagenic with exponentially growing dTMP efficient uptaking yeast strains 831 rho+ and 833 rho . The respiratory deficient strain 833 exhibits a tenfold sensitivity to the genotoxicity of excess dTMP . Mutant yield in the forward mutation system CAN1----can1 after dTMP excess is comparable to that found after irradiation with UV254nm . Excess dTMP is a poor mutagen in stationary phase cells of both strains . Mutagenicity of excess dTMP is not found in an ochre mutant allele (ade2-1) . Exposure of exponentially growing cells to other deoxyribonucleotides (dCMP, dAMP, and dGMP) reveal these nucleotides to have mutagenic potential as well. Zentralbl Mikrobiol, 1985, 140(1), 3 - 11 {Effect of cadmium, zinc, lead and mercury on enzyme activity in Saccharomyces cerevisiae}; Grafl HJ et al.; The in vivo influence of cadmium, zinc, lead, and mercury on the activities of intracellular enzymes in Saccharomyces cerevisiae has been studied . Cadmium and lead cause a significant increase of the enzyme activities . Zinc and mercury do not affect the enzyme activities but they intensify considerably the effects of cadmium and lead . A reduction of enzyme activities is found only rarely . Interactions between the heavy metals tested can lead to synergism or to antagonism or rather to an addition of the effects . The findings suggest that under in vivo conditions heavy metals show only an indirect influence on intracellular enzymes. Chromosoma, 1985, 91(2), 113 - 20 Synaptonemal complexes of normal and mutant yeast chromosomes (Saccharomyces cerevisiae); Moens PB et al.; Synaptonemal complex (SC) analysis of six laboratory yeast strains showed the SC karyotypes to be repeatable within strains . Chromosomal differences were found between strains . In five of the strains, two SCs insert into the nucleolus . This represents a single bivalent with a nucleolar organizer in a medial position as is suggested by genetic data or two bivalents each with a terminal nucleolar organizer . In the first interpretation, n = 16; in the second, n = 17 . Strain Tris has a single nucleolar SC and n = 17 . In strains DCx374, DCx416 and x8366a the genetically determined rearrangements of linkage group III could not be identified . Presumably the short SC (0.33 micron) associated with linkage group III cannot accommodate an inversion loop or a translocation configuration . The strains however were found to harbour a reciprocal translocation involving the nucleolar chromosome . Trisomy for one of the longer chromosomes was observed in Tris and spo10 . It is concluded that rearrangements of the medium and long but not short yeast chromosomes can be detected cytologically . Measurements of nuclear volumes show SC length to vary with artifactually induced swelling of the nucleus . Linear regression of SC length over nuclear radius indicates that actual SC length is only about one-half the observed length . As a result the DNA packing per SC unit length is higher than previously estimated. Anal Biochem, 1985 Jan, 144(1), 179 - 85 Purification of phosphoenolpyruvate carboxykinase from Saccharomyces cerevisiae and its use for bicarbonate assay; Tortora P et al.; Electrophoretically homogeneous phosphoenolpyruvate carboxykinase (EC 4.1.1.49) from Saccharomyces cerevisiae was obtained in high yields by means of a two-step purification procedure consisting of ion-exchange chromatography and affinity chromatography on adenosine 5'-monophosphate-Sepharose 4B . In the latter step the binding of the enzyme to the resin specifically required the presence of Mn2+ . The enzyme was eluted when Mn2+ was removed by addition of ethylenediaminetetraacetate to the elution buffer . Homogeneity, molecular weight, and subunit composition of phosphoenolpyruvate carboxykinase were checked by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration . A factor which caused an underestimation of the enzyme activity in crude extracts was identified as adenylate kinase . Finally, a method is proposed for the enzymatic assay of bicarbonate using a purified phosphoenolpyruvate carboxykinase preparation. Mol Cell Biol, 1985 Jan, 5(1), 226 - 35 Regulation of CDC9, the Saccharomyces cerevisiae gene that encodes DNA ligase; Peterson TA et al.; We have cloned CDC9, the structural gene for Saccharomyces cerevisiae DNA ligase, and investigated its transcriptional regulation both as a function of cell cycle stage and after UV irradiation . The steady-state level of DNA ligase mRNA increases at least fourfold in late G1, after the completion of start but before S phase . This high level of CDC9 mRNA then decays with an apparent half-life of ca . 20 min and remains at a low basal level throughout the rest of the cell cycle . The accumulation of CDC9 mRNA in late G1 is dependent upon the completion of start but not the CDC7 and CDC8 functions . Exposure of cells to UV light elicits an eightfold increase in DNA ligase mRNA levels. Mol Cell Biol, 1985 Jan, 5(1), 17 - 26 RAD3 gene of Saccharomyces cerevisiae: nucleotide sequence of wild-type and mutant alleles, transcript mapping, and aspects of gene regulation; Naumovski L et al.; We determined the complete nucleotide sequence of the RAD3 gene of Saccharomyces cerevisiae . The coding region of the gene contained 2,334 base pairs that could encode a protein with a calculated molecular weight of 89,796 . Analysis of RAD3 mRNA by Northern blots and by S1 nuclease mapping indicated that the transcript was approximately 2.5 kilobases and did not contain intervening sequences . Fusions between the RAD3 gene and the lac'Z gene of Escherichia coli were constructed and used to demonstrate that the RAD3 gene was not inducible by DNA damage caused by UV radiation or 4-nitroquinoline-1-oxide . Two UV-sensitive chromosomal mutant alleles of RAD3, rad3-1 and rad3-2, were rescued by gap repair of a centromeric plasmid, and their sequences were determined . The rad3-1 mutation changed a glutamic acid to lysine, and the rad3-2 mutation changed a glycine to arginine . Previous studies have shown that disruption of the RAD3 gene results in loss of an essential function and is associated with inviability of haploid cells . In the present experiments, plasmids carrying the rad3-1 and rad3-2 mutations were introduced into haploid cells containing a disrupted RAD3 gene . These plasmids expressed the essential function of RAD3 but not its DNA repair function . A 74-base-pair deletion at the 3' end of the RAD3 coding region or a fusion of this deletion to the E . coli lac'Z gene did not affect either function of RAD3. Farmaco {Sci}, 1985 Jan, 40(1), 3 - 13 Photobiological properties of furothiocoumarins in Saccharomyces cerevisiae; Juttermann R et al.; Nine furothiocoumarins corresponding to psoralen, 8-methylpsoralen, 3-carbethoxy-8-methylpsoralen, 8-methoxypsoralen, pseudopsoralen, angelicin, isopseudopsoralen, allopsoralen and pseudoisopsoralen were synthesized by treating furocoumarins with phosphorus pentasulfide . Photobiological studies on haploid yeast cells (Saccharomyces cerevisiae) revealed that the furothiocoumarins exert some photoactivity on cell survival and the induction of mitochondrial damage . In most cases, the furothiocoumarins were less active than their furocoumarinic counterparts and exhibited a preference for a monofunctional type of action . A certain photochemotherapeutic activity can be suggested. Radiat Environ Biophys, 1985, 24(1), 1 - 7 Influence of different inhibitors on the activity of the RAD54 dependent step of DNA repair in Saccharomyces cerevisiae; Siede W et al.; The recombinagenic pathway of DNA repair in yeast was characterized by the effect of different inhibitors on the temperature-dependent survival after gamma-irradiation in haploid cells of the thermoconditional mutant rad54-3 . Blocking protein synthesis with cycloheximide in replicating cells caused partial inhibition of the RAD54 dependent function but some repair activity remained detectable . This indicates that gamma-rays can induce RAD54 activity above some constitutive level of function . Inhibition of DNA replication by hydroxyurea efficiently blocked the RAD54 dependent function in stationary-phase cells but not in logarithmic-phase cells . In logarithmic-phase cells, we found a strong inhibitory effect of caffeine on the RAD54 mediated repair process. Proc Natl Acad Sci U S A, 1985 Jan, 82(1), 168 - 72 RAD6 gene of Saccharomyces cerevisiae encodes a protein containing a tract of 13 consecutive aspartates; Reynolds P et al.; The RAD6 gene of Saccharomyces cerevisiae is required for postreplication repair of UV-damaged DNA, for induced mutagenesis, and for sporulation . We have mapped the transcripts and determi