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  • Yeast  (58)
  • Springer  (58)
  • Annual Reviews
  • Blackwell Publishing Ltd
  • 2005-2009
  • 1990-1994  (41)
  • 1980-1984  (17)
  • 1991  (41)
  • 1982  (17)
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  • 2005-2009
  • 1990-1994  (41)
  • 1980-1984  (17)
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  • 1
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    Springer
    Journal of industrial microbiology and biotechnology 7 (1991), S. 191-195 
    ISSN: 1476-5535
    Keywords: Yeast ; Trehalose ; Osmotolerance ; Viability
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Summary A total of 12 yeast strains from various genera were examined for their ability to produce ethanol in the presence of high concentrations of glucose. From these studies, the yeastsTorulaspora delbrueckii andZygosaccharomyces rouxii were observed to the most osmotolerant. These osmotolerant yeast strains were also observed to possess high concentrations of intracellular trehalose. Futhermore, these strains were found to be tolerant to long-term storage at −20°C and to storage at 4°C in beer containing 5% (v/v) ethanol. Cells containing high trehalose levels at the time of freezing or cold storage exhibited the highest cell viabilities. Trehalose concentration was observed to increase during growth on glucose, reaching a maximum after 24–48 h. Increasing the incubation temperature from 21 to 40°C also resulted in an increase in intracellular trehalose content. These results suggest that trehalose plays a role in enhancing yeast survival under environmentally stressful conditions.
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  • 2
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    Journal of industrial microbiology and biotechnology 7 (1991), S. 263-268 
    ISSN: 1476-5535
    Keywords: Yeast ; β-Glucanase ; β-Glucosidase catabolite repression ; Sporulation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Summary The activities of three glycosidases, β-glucosidase and β(1,3)- and β(1,6)-glucanases have been monitored during growth and blastospore formation inSaccharomycopsis fibuligera. The assays were carried out on the cell-free culture and in a cell-free extract and a wall autolysate preparation from the growing cells. In complex medium containing 1% glucose an increase in the level of all three enzymes was associated with the transition from mycelium to blastospores. When the level of glucose was increased to 5% blastospore formation was repressed and the level of β-glucanases only increased at the end of the fermentation. The β-glucosidase activity increased during the growth phase. In a defined medium in which slow growth in a wholly yeast-like form was observed, growth was not associated with a high level of β-glucanase activity.
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  • 3
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    Journal of industrial microbiology and biotechnology 7 (1991), S. 35-39 
    ISSN: 1476-5535
    Keywords: 2-Deoxy-d-glucose ; Yeast ; Catabolite repression ; Derepression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Summary The non-metabolizable and toxic glucose analogue 2-deoxy-d-glucose (2-DOG) has been widely employed to screen for regulatory mutants which lack catabolite repression. A number of yeast mutants resistant to 2-DOG have recently been isolated in this laboratory. One such mutant, derived from aSaccharomyces cerevisiae haploid strain, was demonstrated to be derepressed for maltose, galactose and sucrose uptake. Furthermore, kinetic analysis of glucose transport suggested that the high affinity glucose transport system was also derepressed in the mutant strain. In addition, the mutant had an increased intracellular concentration of trehalose relative to the parental strain. These results indicate that the 2-DOG resistant mutant is defective in general glucose repression.
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  • 4
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    Current genetics 19 (1991), S. 329-332 
    ISSN: 1432-0983
    Keywords: Mismatch repair ; Plasmid integration ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A single base pair mismatch (G:T or A:C) in the CYC1 gene of the integrative plasmid pAB218 stimulates up to a five-fold integration into the yeast chromosome. Analysis of chromosomal sites of plasmid integration suggests that the mismatch-stimulated integration is not targeted as would be expected if crossovers, localised in the region of the mismatch, were a necessary step in mismatch repair. Instead, the observed mismatch-stimulated plasmid integration could be due to potentially recombinogenic structures formed during mismatch repair, such as single-stranded gaps or denatured DNA regions extending around the plasmid molecule.
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  • 5
    ISSN: 1432-0983
    Keywords: PET genes ; Yeast ; Mitochondria ; ATP synthase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary This study details the characteristics of two temperature-conditional pet mutants of yeast, strains ts1860 and ts379, which at the non-permissive temperature show deficiencies in the formation of three mitochondrially encoded subunits of the ATP synthase complex. By analysis of mitochondrial translation products, and of mitochondrial transcription in temperature shift experiments from the permissive (22°C) to the non-permissive (36°C) temperature, it was concluded that the nuclear mutations in both mutants primarily inhibit synthesis of ATP synthase subunit 9, and that reductions in subunit 8 and 6 synthesis are secondary pleiotropic effects. Following transfer to 36°C, cells of mutant ts379 display a near complete inhibition of subunit 9 synthesis within 1 h, coincident with a marked reduction in the level of the cognate oli1 mRNA. On the other hand, near complete inhibition of subunit 9 synthesis in strain ts1860 occurs after 3 h at 36°C, at which time there is little change in the level of subunit 9 mRNA. In both mutants the mRNA levels for subunits 6 and 8 are not significantly affected at the time of inhibition of subunit 9 synthesis. Provision of an alternative source of subunit 8, translated extra-mitochondrially for import into the organelle, does not overcome the mutant phenotype of either mutant at 36°C, confirming that subunit 8 is not the sole or primary deficiency in each mutant. The mutants indicate that the products of a least two nuclear genes (designated AEP1 and AEP2) are required for the expression of the mitochondrial oli1 gene and the synthesis of subunit 9. The product of the AEP1 gene (defective in mutant ts1860) is required for translation of oli1 mRNA while the AEP2 product (defective in mutant ts379) is essential either for the stability of oli1 mRNA or for the correct processing of precursor transcripts to the mature message.
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  • 6
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    Current genetics 19 (1991), S. 389-393 
    ISSN: 1432-0983
    Keywords: Yeast ; Pichia inositovora ; Linear plasmids ; Killer toxin
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Pichia inositovora, strain NRRL Y-18709, which contains three linear double-stranded DNA plasmids, pPinl-1, pPinl-2 and pPinl-3, was cured of these plasmids both by growing the strain in the presence of 50 μg/ml bisbenzimide, and by exposure to ultraviolet light. Both cured and uncured strains were tested for growth on a variety of carbon sources. No differences in growth response were detected, indicating no discernible involvement of the linear plasmids in the catabolism of these compounds. Culture supernatants of Pichia inositovora were shown to contain a substance larger than 100 kDa that is toxic to Saccharomyces cerevisiae, strain GS 1688. Toxin activity was optimal in YEPD assay plates containing 50 mM citrate buffer with a pH between 3.4 and 4.2. Culture supernatants from P. inositovora were also weakly active against Cephaloascus albidus, strain NRRL Y-18710, and Citeromyces matritensis, strain NRRL Y-18711. Concentrated supernatants from cured P. inositovora strains did not exhibit these activities, consistent with the hypothesis that this toxic activity is linear plasmid-encoded. Unlike the wellknown Kluyveromyces lactis system or the newly identified P. acaciae system, P. inositovora strains cured of their linear plasmids do not become detectably sensitive to toxin produced by the wild-type strain, suggesting a nonplasmid-encoded immunity function.
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  • 7
    ISSN: 1432-0983
    Keywords: Yeast ; Molecular cloning ; Nitrogen mustard hyper-resistance ; Choline transport
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The recessive hnm1 mutant allele is responsible for hyper-resistance to nitrogen mustard in Saccharomyces cerevisiae. Transformation with a single-copy HNM1 wild-type allele of such hyper-resistant mutants will restore wild-type sensitivity to nitrogen mustard. By contrast the presence of multi-copy vectors containing HNM1, in either a hyper-resistant hnm1 mutant or an HNM1 wild-type, will lead to a novel, mustard-sensitive phenotype unrelated to defects in DNA repair genes. Gene disruption of HNM1 revealed that this gene is nonessential for cells prototrophic for choline (CHO1) but lethal for cells with a cho1 genotype. Sensitivity to nitrogen mustard of wild-type HNM1, but not of hnm1 mutants, depends on the choline content of the growth medium, with cells grown in choline-free medium exhibiting the highest sensitivity. Sequencing of a 300 bp DNA fragment of HNM1 revealed the identity of this gene with the CTR locus, which is responsible for choline transport in Saccharomyces cerevisiae.
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  • 8
    ISSN: 1432-0983
    Keywords: Mutagen hyper-resistance ; Yeast ; Base sequence ; Gene disruption
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A multi-copy plasmid containing the SNQ3 gene confers hyper-resistance to 4-nitroquinoline-N-oxide (4NQO), Trenimon, MNNG, cycloheximide, and to sulfometuron methyl in yeast transformants. Restriction analysis, subcloning, and DNA sequencing revealed an open reading frame of 1950 bp on the SNQ3-containing insert DNA. Gene disruption and transplacement into chromosomal DNA yielded 4NQO-sensitive null mutants which were also more sensitive than the wild-type to Trenimon, cycloheximide, sulfometuron methyl, and MNNG. Hydropathic analysis showed that the SNQ3-encoded protein is most likely not membrane-bound, while the codon bias index points to low expression of the gene.
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  • 9
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    Current genetics 19 (1991), S. 89-94 
    ISSN: 1432-0983
    Keywords: Yeast ; Mitochondria ; Intron ; Mobile
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The mitochondrial and nuclear genomes of 21 yeast species belonging to 12 genera have been tested for the presence of sequences similar to seven S. cerevisiae mitochondrial introns (Sc cox1.1,2,3,4,5c, Sc cob.4 and Sc LSU.1) and one K. lactis mitochondrial intron (Kl cox1.2). Some introns, (Sc cox1.4, Sc cob.4, Sc LSU.1 and Kl cox1.2-all group I type), are widely distributed and are found in species with either basidiomycete or ascomycete affinities. This distribution is suggestive of recent sequence transfer between species. The remaining S. cerevisiae introns cross react with an additional species but with no set pattern. Pulsed field gel electrophoretic studies confirm that none of the tested mitochondrial introns cross react with nuclear DNA. These introns are, therefore, mitochondria-specific. Seven strains of K. lactis exhibit striking variability in intron content. In contrast to all mitochondrial introns tested, two introns of nuclear genes (the K. lactis actin gene and the S. cerevisiae RP29B gene) are not detected beyond their source species.
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  • 10
    ISSN: 1432-0983
    Keywords: Yeast ; Pyruvate decarboxylase ; Gene expression ; Codon usage ; Gene fusion
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Three structural genes encode the pyruvate decarboxylase isoenzymes in the yeast Saccharomyces cerevisiae. PDC1 and PDC5 are active during glucose fermentation where PDC1 is expressed about six times more strongly than PDC5. Expression of PDC6 is weak and seems to be induced in ethanol medium. Consequently, pdc1Δ pdc5Δ double mutants do not ferment glucose and do not grow on glucose medium. Spontaneous mutants, derived from such a pdc1 pdc5 strain, were isolated which could again ferment glucose. They showed pyruvate decarboxylase activity due to a duplication of PDC6. The second copy of PDC6 was expressed under the control of the PDC1 promoter, which was still present in the pdc1 strain. However, the resulting PDC1-PDC6 fusion gene could only partially substitute for PDC1: to achieve normal growth and high pyruvate decarboxylase activity strains carrying PDC1-PDC6 required a functional PDC5 gene which is dispensable in a PDC1 wild-type background. Thus, expression of PDC5 depends on the state of the PDC1 locus: low in the PDC1 wild-type background and high in PDC1-PDC6 fusion strains and, as shown previously, in pdc1 mutants. The activation of PDC5 expression in PDC1-PDC6 strains may be due to particular properties of the PDC1-PDC6 fusion protein or simply to the weaker expression of PDC1-PDC6 in comparison to the wild-type PDC1 gene.
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  • 11
    ISSN: 1432-0983
    Keywords: AEP2 ; Yeast ; Mitochondria ; ATP synthase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The temperature-conditional pet mutant, ts379, of Saccharomyces cerevisiae fails to synthesize mitochondrial ATP synthase subunit 9 at the restrictive temperature due to mutation of a single nuclear locus, AEP2. The inability to synthesize subunit 9 correlates with a lowered accumulation of the cognate oli1 mRNA indicating that the AEP2 product is involved in oli1 transcript maturation or stabilization. The AEP2 gene has been isolated in this study from a wild-type yeast genomic library by genetic complementation of ts379 at the restrictive temperature. A 1740 nucleotide open-reading frame was observed that encodes a basic, hydrophilic protein of 67534 Da which possesses a putative mitochondrial address signal. Disruption of chromosomal DNA within this reading frame produced a non-conditional respiratory mutant unable to synthesize subunit 9, identifying the AEP2 gene. Hybridization analyses indicate that AEP2 is located on chromosome XIII and produces a 2.1 kb poly(A)+ transcript. Two additional open-reading frames were found in close proximity to that of AEP2. The three open-reading frames shared no significant homology with entries in several data bases.
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  • 12
    ISSN: 1432-0983
    Keywords: Pulsed field gel-electrophoresis ; S1 nuclease sensitive sites ; Repair ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Repair under non-growth conditions of DNA double-strand breaks (DSB) and chromatin sites sensitive to S1 endonuclease (SSS) induced by 60Cobalt-gamma rays were monitored in repair-competent and deficient strains of Saccharomyces cerevisiae by pulsed field gelelectrophoresis. In stationary-phase cells of a repair-competent RAD diploid, and an excision-deficient rad3-2 diploid, SSS are repaired as efficiently as DSB, whereas in a repair-competent RAD haploid, and a rad 50-1 diploid, neither SSS nor DSB are repaired. The rad18-2 diploid repairs DSB well but is defective in SSS repair. Obviously, SSS repair in yeast chromatin, like DSB repair, depends on recombination, but unlike DSB repair depends additionally on RAD18 function.
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  • 13
    ISSN: 1432-0983
    Keywords: Yeast ; Mistranslation ; ψ-factor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Chromosomal omnipotent suppressor mutations recovered in ψ+ strains of Saccharomyces cerevisiae were brought into ψ− cytoplasm. SUP46, SUP138 and SUP139 acted as dominant omnipotent suppressors in the ψ− cytoplasm though their suppressor activity was substantially reduced. SUP46 and SUP138 conferred recessive thermosensitivity and antibiotic sensitivity in ψ− cytoplasm as in ψ+ cytoplasm. On the other hand, sup111 through sup115, which acted as recessive omnipotent suppressors in the ψ+ cytoplasm, manifested no, or very low, suppressor activity in the ψ− cytoplasm. They, however, still enhanced the efficiency of the SUP29 tRNA suppressor in ψ− cytoplasm. A multicopy plasmid carrying the wild-type SUP35 gene enhanced the efficiency of sup111 in ψ− cytoplasm.
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  • 14
    ISSN: 1432-0983
    Keywords: Yeast ; Mutant ; p-Fluoro-dl-phenylalanine ; β-Phenethyl-alcohol
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary p-Fluoro-dl-phenylalanine (PFP)-resistant mutants which produce a large amount of β-phenethyl-alcohol, a rose-like flavor component, were isolated from the isogenic strains X2180-1A and X2180-1B of Saccharomyces cerevisiae. Cells of these mutants accumulated phenylalanine and tryptophan more than 3-fold times that of wild-type cells, while they accumulated less than half the tyrosine. The activity of prephenate dehydrogenase (PDG) (EC 1.3.1.12) was markedly decreased while that of 3-deoxy-d-arabino-heptulosonate-7-phosphate synthase (EC 4.1.2.15) was increased. Genetic analysis revealed that the mutation occurred at the TYR1 locus, encoding PDG, and that the mutated TYR1 gene, tyr1-pfp, caused both PFP resistance and β-phenethyl-alcohol overproduction. This was supported by molecular genetic studies with cloned tyr1-pfp DNA.
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  • 15
    ISSN: 1432-0983
    Keywords: Peptides ; Transport ; Regulation ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The transport of small peptides into the yeast Saccharomyces cerevisiae is subject to complex regulatory control. In an effort to determine the number, and to address the function, of the components involved in peptide transport and its regulation, spontaneous mutants resistant to toxic di- and tripeptides were isolated under inducing conditions. Twenty-four mutant strains were characterized in detail and fell into two phenotypic groups; one group deficient in amino acid-inducible peptide uptake, the other with a pleiotropic phenotype including a loss of peptide transport. Complementation analysis of recessive mutations in 12 of these strains defĩned three groups; ptr1 (nine strains), ptr2 (two strains), and ptr3 (one strain). Isolation and screening of 31 additional N-methyl-N-nitro-N-Nitrosoguanidine (MNNG)-induced, peptide transport-deficient mutants produced one ptr3 and 30 ptr2 strains: no additional complementation groups were detected. Uptake of radiolabeled dileucine was negligible in ptr1 and ptr2 strains and was reduced by 65% and 90% in the two ptr3 mutants, indicating that all strains were defective at the transport step. We conclude that the S. cerevisiae amino acid-inducible peptide transport system recognizes a broad spectrum of peptide substrates and involves at least three components. One gene, PTR3, may play an indirect or regulatory role since mutations in this gene cause a pleiotropic phenotype.
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  • 16
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    Journal of molecular evolution 32 (1991), S. 396-404 
    ISSN: 1432-1432
    Keywords: Yeast ; Mitochondrial DNA ; Polymirphism ; Repeated sequences
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A spontaneously arising mitochondrial DNA (mtDNA) variant ofSaccharomyces cerevisiae has been formed by two exta copies of a 14-bp sequence (TTAATTAAATTATC) being added to a tandem repeat of this unit. Similar polymorphisms in tandemly repeated sequences have been found in a comparison between mtDNAs from our strain and others. In 5850 bp of intergenic mtDNA squence, polymorphisms in tandemly repeated sequences of three or more base pairs occur approximately every 400–500 bp whereas differences in 1–2 bp occur approximately every 60 bp. Some polymorphisms are associated wit optional G+C-rich sequences (GC clusters). Two such optional GC clusters and one A+T repeat polymorphism have been discovered in the tRNA synthesis locus. In addition, the variable presence of large open reading frames are documented and mechanisms for generating intergenic sequence diversity inS. cerevisiae mtDNA are discussed.
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  • 17
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    Journal of molecular evolution 32 (1991), S. 439-442 
    ISSN: 1432-1432
    Keywords: Yeast ; Mitochondrial DNA ; ori ; rep ; Polymorphism
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Threeori elements (ori 2,ori 5, andori 7) have been sequenced inSaccharomyces cerevisiae strain Dip 2 and compared to the equivalentori elements of a second strain (B). Bothori 2 andori 5 exhibit 98% base matching between strains Dip 2 and B. In contrast, the thirdori element (ori 7) exhibits extensive sequence rearrangements whereby a segment located downstream in the consensus strain occurs within theori structure in Dip 2. This represents a novel polymorphic form of the yeast mitochondrial genome.
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  • 18
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    Current genetics 20 (1991), S. 471-474 
    ISSN: 1432-0983
    Keywords: Yeast ; DNA replication ; Chemical mutagenesis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Incubation of cdc8 mutants of the yeast Saccharomyces cerevisiae in YPD under permissive conditions, when DNA replication is taking place, prior to transfer to restrictive conditions, strongly stimulates induction of cdc + colonies of ethyl methane sulphonate (EMS)- and methyl methane sulphonate (MMS)-treated yeast strains HB23 (cdc8-1/cdc8-3), HB26 (cdc8-3/cdc8-3) and HB7 (cdc8-1/cdc8-1). After diepoxybutane (DEB) treatment, both the induction of cdc + colonies and their stimulation after incubation in YPD under permissive conditions is low. The results obtained show that stimulation of induction of cdc + colonies under permissive conditions occurs not only after UV-treatment, but also after treatment with such mutagens as EMS and MMS.
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  • 19
    ISSN: 1432-0983
    Keywords: Yeast ; Mitochondrial DNA ; Antibiotic resistance mutations
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A large proportion of the spontaneous erythromycin resistant mutants isolated from a strain carrying a previously-induced chloramphenicol resistance mutation at cap3 do not map at ery1, the locus most often associated with mitochondrial erythromycin resistance. Most of the new mutations are also nonallelic at spil, spi2, and other known antibiotic resistance loci within the 21S rRNA gene; they are allelic with each other and define the new locus, ery2. Induced second-site erythromycin resistant mutants from the cap r3 strain, as well as spontaneous or induced mutants from strains carrying a cap r 1 mutation, all tend to map at eryl. The cap r3 mutation is apparently necessary for the expression of erythromycin resistance resulting from a second mutation at ery2.
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  • 20
    ISSN: 1432-0983
    Keywords: Yeast ; Thermoconditional DNA repair ; Mutagenesis ; Allelism test
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Of two mutant genes (snm1-2 ts and snm2-1 ts) conferring thermoconditional mutagen sensitivity in Saccharomyces cerevisiae one (snm2-1 ts) is shown to be centromere-linked. At the restrictive temperature this allele reduces UV-induced back mutation frequency of the ochre allele hiss-2 but has no influence on forward mutation at the CAN1 locus. Complementation tests and recombination analysis revealed snm2 ts to be allelic with rad5 (rev2).
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  • 21
    ISSN: 1432-0983
    Keywords: Electro-fusion ; Yeast ; Plasmogamy ; Proliferation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Electric field-induced fusion was induced between Saccharomyces cerevisiae protoplasts from the ρ − heterozygous diploid strain 2114 and the respiratory-competent diploid strain 3441, carrying chromosomal markers. Close membrane contact between the cells of the two different strains (ratio 1:1) was achieved by dielectrophoresis in a weak inhomogeneous alternating field (about 1 kV/cm, 2 MHz). Due to dielectrophoresis pearl chains of two or more cells of the two strains are formed between the electrodes. Cell fusion was induced by application of two single square field pulses sufficiently high to induce reversible electrical breakdown in the membrane contact zone between cells within a pearl chain (about 7 to 8 kV/cm field strength and 40 Ms duration). The two subsequent pulses were applied at an interval of about 10 s. Hybrids could be isolated on selection medium in a high yield (compared with conventional fusion techniques). The hybrids were diploid, respiratory-competent and produced prototrophic spores. Thus, the fused hybrids contained only the chromosomal markers of strain 2114 and the cytoplasmic marker for respiratory competence from strain 3441; electro-fusion thus resulted mainly in plasmogamy.
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  • 22
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    Current genetics 6 (1982), S. 93-98 
    ISSN: 1432-0983
    Keywords: Yeast ; Genetic mapping ; Trisomic analysis ; Arginine
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary By use of a set of 8 aneuploid strains of the yeast Saccharomyces cerevisiae, carrying from 1 to 5 identified disomic chromosomes, in crosses to a set of haploid strains collectively bearing 11 unmapped genes, the following chromosome assignments were obtained for these unmapped genes: arg80 on XIII;arg3 on X;car2 on XII; cpa1 and tsm8740 on XV; tsm7269 (=rna6) on II; cpa2 on X or XV; arg82 and tsm4572 on III, IV or XVI; car1 and arg81 on II, IV, VI, VII or XVI. Linkage tests between the unmapped genes and markers located on the chromosomes that had been designated as possible carriers by the previous analysis allowed 8 genes to be localized. The remaining three genes, cpa2, car1 and arg81 (located on fragment F8), could not be positioned on any of the chromosomes indicated by the trisomic analysis, in spite of testing for linkage to markers covering most of the known regions of these chromosomes.
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  • 23
    ISSN: 1432-0983
    Keywords: Petite mutation ; NUC2 nuclease ; Yeast ; RAD52 ; Ethidium bromide
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Defects in the RAD52 gene of the yeast Saccharomyces cerevisiae reduce the levels of the NUC2 endo-exonuclease by approximately 90% compared to the levels in wild-type strains. To examine the potential role of this nuclease in the induction of mitochondrial ‘petite’ mutations, congenic RAD52 and rad52-1 haploids were subjected to treatment with ethidium bromide, a well-known inducer of these mutations. The rad52 strain showed a much higher resistance to ethidium bromide-induced petite formation than the corresponding wild-type strain. Two approaches were taken to confirm that this finding reflected the nuclease deficiency, and not some other effect attributable to the rad52-1 mutation. First, a multicopy plasmid (YEp213-10) carrying NUC2 was transformed into a RAD52 strain. This resulted in an increased fraction of spontaneous petite mutations relative to that seen for the same strain without the plasmid and sensitized the strain carrying the plasmid to peptite induction by ethidium bromide treatment. Second, a strain having a nuc2 allele that encodes a temperaturesensitive nuclease was treated with ethidium bromide at the restrictive and permissive temperatures. Petite induction was reduced under restrictive conditions. Enzyme assays revealed that the RAD52 (YEp213-10) strain had the highest level of antibody-precipitable NUC2 endo-exonuclease whereas the nuc2 and rad52 mutants had the lowest levels. Furthermore, addition of ethidium bromide to the reaction mixture stimulated the activity of the nuclease on double-stranded DNA. Peptite induction by antifolate-mediated thymine nucleotide depletion was also inhibited by inactivation of RAD52 indicating that the effect of reduced NUC2 endo-exonuclease was not restricted to ethidium bromide treatment. Taken collectively, these results indicate that the NUC2 gene product functions in the production of mitochondrial petite mutations.
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  • 24
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    Current genetics 5 (1982), S. 153-155 
    ISSN: 1432-0983
    Keywords: Yeast ; Mutant cell-wall ; Permeability exponentialy
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary When Saccharomyces cerevisiae SY15 rho° mutant cells grown in media stabilized with 10% sorbitol were suspended in 2% sorbitol solutions, 60–70% of the population did not lyse and became permeable to native high molecular weight DNA. Maximal incorporation of DNA to DNase resistant state was measured after 60 min of incubation in presence of 5 μg/ml DNA and 10 mM CaCl2. These results suggest that the fragile mutants might be tested as hosts for transformation of whole yeast cells.
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  • 25
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    Current genetics 20 (1991), S. 1-3 
    ISSN: 1432-0983
    Keywords: Yeast ; Transformation ; Ethanol
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A technique is described in which ethanol is used to improve the genetic transformation of intact yeast (Saccharomyces cerevisiae) cells pretreated with LiAc and PEG. Transformation efficiency was increased with increasing concentrations of ethanol with a peak at 10% concentration. The effect varies with different yeast strains and plasmids and up to a maximum of a 15-fold increase was observed.
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  • 26
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    Current genetics 20 (1991), S. 25-31 
    ISSN: 1432-0983
    Keywords: Yeast ; TSM1 sequence ; Essential gene ; MAT distal cloning
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We have cloned the region from MAT to THR4 on chromosome III of Saccharomyces cerevisiae. Although the region is only 15 kb, the two loci are genetically separated by 22 cM. This is in sharp contrast to the very low level of recombination (2 cM in 22 kb) that is observed in the adjacent CRY1-MAT interval, and suggests that there may be a “hot spot” for recombination in the MAT-THR4 region. The DNA sequence of the first 4.4 kb distal to MAT reveals an open reading frame that we have identified as the essential gene, TSM1. Surprisingly, the TSM1 open reading frame of 1 410 amino acids extends into the MAT locus, such that the 3′-end of the MATα1 transcript ends 15 bp from the 3′-end of the TSM1 open reading frame.
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  • 27
    ISSN: 1432-0983
    Keywords: Yeast ; Enhancer ; Transcriptional elements ; Transcriptional factors ; Regulation
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    Notes: Summary Though highly complex enhancers found in animal cells have not been reported to occur in yeasts they are able to activate the transcription of adjacent genes in yeast cells. Saccharomyces cerevisiae expresses a large number of nuclear proteins that are able to recognize, and specifically bind to, the enhancer sequences of the SV40 animal tumor virus. The complexity of proteins that interact with different elements of the animal enhancers is similar in yeast and animal cell nuclear extracts. Most enhancer motifs, recognized by known trans-acting factors, are protected in footprinting experiments by yeast nuclear proteins.
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  • 28
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Ergosterol ; Squalene synthetase ; Yeast
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    Notes: Summary The ERG9 gene of Saccharomyces cerevisiae has been cloned by complementation of the erg9-1 mutation which affects squalene synthetase. From the 5kkb insert isolated, the functional gene has been localized on a DNA fragment of 2.5 kb. The presence of squalene synthetase activity in E. coli bearing the yeast DNA fragment isolated, indicates that the structural gene encoding squalene synthetase has been cloned. The sequence of the 2.5 kb fragment contains an open reading frame which could encode a protein of 444 amino acids with a deduced relative molecular mass of 51 600. The amino acid sequence reveals one to four potential transmembrane domains with a hydrophobic segment in the C-terminal region. The N-terminus of the deduced protein strongly resembles the signal sequence of yeast invertase suggesting a specific mechanism of integration into the membranes of the endoplasmic reticulum.
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  • 29
    ISSN: 1432-0983
    Keywords: Mitochondria ; Intron ; Telomere ; Yeast
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    Notes: Summary The junctions between X and Y′ subtelomeric repeats in Saccharomyces cerevisiae usually contain a stretch of telomere sequences, (G1–3T)n. Two of three cloned X-Y′ junctions from strain YP1 have a replacement of about 200 bp of X, the internal telomere sequence, and 49 bp of Y′ by a 292 bp sequence. The first 227 bp of this insertion sequence are 100% identical to the fourth intron of cytochrome b. The rest of the insertion has homology to an unknown dispersed nuclear sequence. Recombination among subtelomeric regions can explain the nuclear distribution of this sequence and why telomeres can trap and maintain sequences that would otherwise be lost.
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  • 30
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    Current genetics 6 (1982), S. 99-103 
    ISSN: 1432-0983
    Keywords: Ultraviolet light mutagenesis ; Mitochondrial genome ; Meiosis ; Yeast
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    Notes: Summary Clones derived from ascospores from ultraviolet irradiated diploid cells were examined for the genetic determinants or respiratory properties. Approximately 10% of the cells produced petites of mitochondrial origin at the dose applied. Among 13 asci which produced mitochondrial petites with high frequencies, 6 asci of uniparental type, 0 grandes : 4 petites, were observed. Furthermore, most of the petite spore clones from each individual uniparental ascus showed similar levels of suppressiveness and of mitochondrial gene retention. From these results it is suggested that a single mitochondrial genome participates meiosis in yeast.
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  • 31
    ISSN: 1432-0983
    Keywords: In vitro mutagenesis ; PET-genes ; RNA-leader ; Ribosomal scanning ; Yeast
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    Notes: Summary We report that the major transcription start sites of the yeast PET gene SCO1 are located at positions-149 and -125 relative to the AUG initiation codon of the SCO1 reading frame. The leader sequences of the resulting mRNAs possess a single AUG codon at position-49, which initiates a short open reading frame of three amino acids. The recent finding of a similar situation in the case of the PET gene CBS1 prompted us to address the question as to whether these AUG codons might play some role in the expression of these PET genes. After removal of the upstream AUG codons by site-directed mutagenesis, expression was monitored by use of lacZ fusions and compared to the respective wildtype constructs. Our data show that under all growth conditions tested the leader-contained AUG initiation codons have no significant influence on the expression of both PET genes.
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  • 32
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    Current genetics 6 (1982), S. 179-188 
    ISSN: 1432-0983
    Keywords: Yeast ; Mitochondria ; var1 gene
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    Notes: Summary Several mit mutants mapping within or near the var1 determinant region have been characterized genetically and biochemically. These mutants were isolated using a new enrichment protocol which simplifies the isolation and identification of rare respiration-deficient mutants of yeast. Two of the mutants, PZ200L and PZ206, map in genome segments which flank the known varl gene reading frame; nevertheless, both belong to the same complementation group, apparently that of the varl gene. A third mutant, PZ200R is closely linked to one of the varl allelic determinants now known to be a short insertion within the gene. All three var1 mutants exhibit decreased levels of mitochondrial protein synthesis and negligible activity of the respiratory enzyme complexes. Another cluster of mutants belonging to a separate complementation group from that defined by PZ200L and PZ206 was also mapped and it contains mutants in the nearby serine tRNA gene. The isolation of these mutants in the varl region shows that the varl locus contains information essential for the maintenance of respiration-competent mitochondria. Because these mutants affect mitochondrial protein synthesis, their existence supports the previous hypothesis that the varl protein is an integral component of mitochondrial ribosomes. Furthermore, the mutant sites are present in a DNA sequence that is highly, rich in A+T residues that also contains a gene. Since approximately 50% of the yeast mitochondrial genome is similarly rich in A+T and since most of those regions have not yet been sequenced it is quite possible that other A+T-rich genes may exist.
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  • 33
    ISSN: 1432-0983
    Keywords: Nitrosoguanidine ; Comutation ; Yeast ; Chromosome replication pattern
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    Notes: Summary Contrary to what happens in bacteria, mutations induced by nitrosoguanidine in yeast are not accompanied by an excess of mutations in nearby genes. We have investigated nitrosoguanidine mutagenesis in three regions of the yeast genome: the contiguous DNA segments HIS4A, HIS4B and HIS4C, located on chromosome III; ADE1 and CDC15 separated by about 3 map units on chromosome I; and CAN1, some 50 map units away from the centromere on chromosome V. Revertants at HIS4C never suffered mutations at HIS4A or HIS4B. Reversion at CDC15 did not affect the frequency of mutation at ADE1. No tsm mutations, leading to thermonsensitivity, were found in the immediate vicinity of the locus CAN1 after selecting for canavanine resistant mutants. However, as expected from nitrosoguanidine mutagenesis of replication points and the fixed pattern of chromosome replication, the induced tsm mutations seem not to map randomly over the yeast genome; in fact, two out of the three groups of such tsm mutations studied are located in the same chromosome arm as CAN1, indicating that these two regions are replicated at the same time as CAN1. Replication synchrony is less than perfect, since the tsm mutations of each group affect many different genes.
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  • 34
    ISSN: 1432-0983
    Keywords: Yeast ; Cycloheximide ; Ribosomal mutations
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    Notes: Summary At least four different mutations at the cyh2 locus (rp1X; gene product: YL24) of Saccharomyces cerevisiae confer cycloheximide resistance. The mutant YL24 proteins are either more basic (high-level resistant phenotype), more acidic (low-level resistant phenotype), or unchanged in their electrophoretic mobility (both low-and high-level resistant phenotypes). None of the mutations at other loci seem to induce high-level resistance to cycloheximide.
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  • 35
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    Current genetics 5 (1982), S. 171-180 
    ISSN: 1432-0983
    Keywords: Yeast ; Mitochondrial genes ; Vegetative segregation
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    Notes: Summary A three-factor cross of Saccharomyces cerevisiae involving the cap1, ery1, and oli1 loci was done, with partial pedigree analyses of 117 zygotes. First, second, and third buds were removed and the genotypes of their diploid progeny determined, along with those of the residual zygote mother cell. Results were analyzed in terms of frequencies of individual alleles and of recombinant genotypes in the dividing cells. There is a gradual increase in the frequency of homoplasmic cells and in gene frequency variance during these three generations, as would result from stochastic partitioning of mtDNA molecules between mother and bud, probably coupled with random drift of gene frequencies in interphase cells. These phenomena are more pronounced for buds than for mothers, suggesting that buds receive a smaller sample of molecules. End buds are more likely to be homoplasmic and have a lower frequency of recombinant genotypes than do central buds; an end bud is particularly enriched in alleles contributed by the parent that formed that end of the zygote. Zygotes with first central buds produce clones with a higher recombination frequency than do those with first end buds. These results confirm previous studies and suggest that mixing of parental genotypes occurs first in the center of the zygote. If segregation were strictly random, the number of segregating units would have to be much smaller than the number of mtDNA molecules in the zygote. On the other hand, there is no evidence for a region of the molecule (“attachment point”) which segregates deterministically.
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  • 36
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    Current genetics 6 (1982), S. 29-30 
    ISSN: 1432-0983
    Keywords: Yeast ; Plasmid ; Repair ; Ligase
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    Topics: Biology
    Notes: Summary We show that the DNA ligase encoded or controlled by the cdc9 gene in Saccharomyces cerevisiae is required for replication of plasmid DNA but that excision repair of pyrimidine dimers in plasmid DNA can be completed in its absence.
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  • 37
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    Current genetics 6 (1982), S. 195-201 
    ISSN: 1432-0983
    Keywords: Benomyl resistance ; Yeast ; Mitosis ; Cell cycle mutants
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    Notes: Summary We have isolated 150 benomyl resistant mutants of the fission yeast Schizosaccharomyces pombe. Seven of these mutants were found to be cold sensitive for mitosis. These mutants were the subject of physiological, cytological and genetical characterisation. Growth and division of the seven mutants were similar to the wild type strain at 35 °C. After shift from the permissive (35 °C) to the restrictive temperature (20 °C) the mutants became blocked in mitosis whilst cellular growth continued. Consequently, elongate cells were formed. Six of the seven benomyl resistant mutants became blocked in mitosis at 20 °C with a single aberrant nucleus. In every case the benomyl resistant and cold sensitive phenotype was due to a mutation in a single nuclear gene. These mutants were found to comprise a single genetic linkage group (ben4) and were unlinked to existing TBZ/MBC resistant mutants of S. pombe. Whilst no cross resistance was found in our mutants to TBZ, six of the seven mutants were super sensitive to the spindle poison CIPC. We believe that the phenotype exhibited by these mutants is consistent with a defective tubulin subunit.
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  • 38
    ISSN: 1432-072X
    Keywords: Ions ; Concentration ; Regulation ; Cytoplasm ; Vacuole ; Yeast ; Saccharomyces carlsbergensis
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    Notes: Abstract Intracellular distributions of K+, Mg2+ and orthophosphate under various conditions of cultivation or incubation of the yeast Saccharomyces carlsbergensis were studied by differential extraction of ion pools. The decisive role of vacuolar compartmentation of ions in regulation of K+, Mg2+ and orthophosphate levels in the yeast cytoplasm was shown. The content of intracellular K+ and Mg2+ in yeast increased or decreased primarily depending on the increase or decrease in the vacuolar ion pool. The levels of K+ and Mg2+ in the cytoplasm were practically unchanged. Vacuoles were involved in regulation of Mn2+ concentration in the cytoplasm of the yeast S. carlsbergensis accumulating this ion in the presence of glucose. Alongside the vacuolar compartmentation, the chemical compartmentation, i. e. formation of bound Mg2+, Mn2+ and K+ was, evidently, also involved in the control of ion levels in the cytoplasm. The orthophosphate level in the yeast cytoplasm was regulated by its accumulation in vacuoles and biosynthesis of inorganic polyphosphates in these organelles. The biosynthesis of low-molecular weight polyphosphates occurred parallel to the accumulation of Mg2+ or Mn2+ in vacuoles, thus confirming the availability of the other mechanism for the transport of these ions through the tonoplast differing from the transport mechanism through the plasmalemma.
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  • 39
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    Archives of microbiology 131 (1982), S. 298-301 
    ISSN: 1432-072X
    Keywords: Yeast ; Protoplast ; Compartmentation ; Vacuole ; Trehalose ; Trehalase ; Carbohydrate metabolism ; Saccharomyces cerevisiae
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    Notes: Abstract Protoplasts of Saccharomyces cerevisiae synthesized and degraded trehalose when they were incubated in a medium containing traces of glucose and acetate. Such protoplasts were gently lyzed by the polybase method and a particulate and soluble fraction was prepared. Trehalose was found in the soluble fraction and the trehalase activity mostly in the particulate fraction which also contained the vacuoles besides other cell organelles. Upon purification of the vacuoles, by density gradient centrifugation, the specific activity of trehalase increased parallel to the specific content of vacuolar markers. This indicates that trehalose is located in the cytosol and trehalase in the vacuole. It is suggested that trehalose, in addition to its role as a reserve may also function as a protective agent to maintain the cytosolic structure under conditions of stress.
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  • 40
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    Archives of microbiology 132 (1982), S. 144-148 
    ISSN: 1432-072X
    Keywords: Yeast ; Cell wall ; Mannoproteins ; Envelope turnover ; Concanavalin A
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    Notes: Abstract By pulse and chase labeling experiments, two independent mannoprotein pools have been found associated with the Saccharomyces cerevisiae envelope. One of them probably corresponds to mannoproteins localized in the periplasmic space. These molecules showed a high turnover rate at 28° C. The second pool is formed by intrinsic wall mannoproteins which are apparently stable for long periods of time, after a small initial turnover. These results suggest that at least part of the mannoproteins initially found in the periplasmic space may move into the wall. The time lag between the addition of the radioactive precursors and their incorporation in the cell envelope (20–30 min for amino acids and about 10 min for carbohydrate) indicates that protein formation and carbohydrate incorporation take place in succession. Moreover, bulk glycosylation of mannoproteins seems to occur close in time to the moment of secretion into the periplasmic space.
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  • 41
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    Archives of microbiology 133 (1982), S. 155-161 
    ISSN: 1432-072X
    Keywords: Microtubule ; Nocodazole ; Yeast ; Cell cycle ; Dimorphism ; Fungus ; Wangiella dermatitidis
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    Topics: Biology
    Notes: Abstract The microtubule inhibitor nocodazole {methyl-5-[2-(thienylcarbonyl)-1H-benzimidazol-2-yl]-carbamate} prevented nuclear migration and nuclear division in yeasts and developing multicellular forms of the polymorphic fungus Wangiella dermatitidis. It did not prevent yeast bud formation during at least two or three budding cycles, and caused yeasts to accumulate as premitotic forms with one to three buds. The effects of the drug suggested that at least three control pathways were involved in the yeast cell cycle; that the nocodazole block point was separate from the execution points of two temperature-sensitive mutations which lead to multicellularity; and that microtubules were controlling neither the yeast budding process nor the development of multicellular forms.
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  • 42
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    Archives of microbiology 133 (1982), S. 131-136 
    ISSN: 1432-072X
    Keywords: Fructose-bisphosphatase deficient mutants ; Yeast ; Schizosaccharomyces pombe ; Gluconeogenesis ; Glucose repression
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    Topics: Biology
    Notes: Abstract We showed that in the yeast Schizosaccharomyces pombe, fructose-bisphosphatase is not subject to catabolite inactivation as it was observed in Saccharomyces cerevisiae. However, this enzyme activity is sensitive to catabolite repression in both yeasts. Two mutants lacking completely fructose-bisphosphatase activity were found. They were unable to grow on glycerol medium. They were still respiratory competent and exhibited the ability to derepress partially malate dehydrogenase activity. In glucose exponential phase culture, the parental strain lacks completely the fructosebisphosphatase activity due to catabolite repression. In these conditions, the growth is slowed down only in the mutants eventhough both mutants and their parental strain lack this enzyme activity. Normal sporulation and poor spore germination were observed for one mutant whereas, only in the presence of glucose, normal sporulation and normal spore germination were observed for the second mutant. Mendelian segregation of glycerol growth was found for the well germinating mutant. It is of nuclear heredity. The two mutations appeared to be closely linked.
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  • 43
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    Molecular genetics and genomics 230 (1991), S. 241-250 
    ISSN: 1617-4623
    Keywords: Yeast ; Saccharomyces cerevisiae ; Adenylyl cyclase ; CDC25 ; RAS
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The TFS1 gene of Saccharomyces cerevisiae is a dosage-dependent suppressor of cdc25 mutations. Overexpression of TFS1 does not alleviate defects of temperature-sensitive adenylyl cyclase (cdc35) or ras2 disruption mutations. The ability of TFS1 to suppress cdc25 is allele specific: the temperature-sensitive cdc25-1 mutation is suppressed efficiently but the cdc25-5 mutation and two disruption mutations are only partially suppressed. TFS1 maps to a previously undefined locus on chromosome XII between RDN1 and CDC42. The DNA sequence of TFS1 contains a single long open reading frame encoding a 219 amino acid polypeptide that is similar in sequence to two mammalian brain proteins. Insertion and deletion mutations in TFS1 are haploviable, indicating that TFS1 is not essential for growth.
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  • 44
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    Molecular genetics and genomics 226 (1991), S. 97-106 
    ISSN: 1617-4623
    Keywords: Yeast ; Linear plasmid ; Saccharomyces kluyveri ; Kluyveromyces lactis ; Killer plasmid
    Source: Springer Online Journal Archives 1860-2000
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    Notes: Summary We have determined the complete nucleotide sequence of the linear DNA plasmid, pSKL, isolated from Saccharomyces kluyveri. Sequence analysis showed that pSKL has a high (A+T) content of 71.7%, and that there are 10 open reading frames (ORFs) larger than 250 nucleotides. All 10 ORFs were shown to be transcribed in S. kluyveri cells by S1 nuclease mapping analysis. The localization of ORFs, direction of transcription, and the predicted amino acid sequences of each ORF were quite similar to that of pGKL2, one of the killer plasmids found in Kluyveromyces lactis. The amino acid sequences of the largest two ORFs (ORF2 and ORF6) have homology with several DNA polymerases and RNA polymerases, respectively.
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  • 45
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    Molecular genetics and genomics 226 (1991), S. 145-153 
    ISSN: 1617-4623
    Keywords: Retrotransposons ; Reverse transcription ; Ty elements ; Yeast
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    Notes: Summary Transposition of the yeast transposable element, Ty, has been shown to require a reverse transcription process. By analysing the extrachromosomal Tyspecific nucleic acid molecules associated with overproduced Ty virus-like particles (Ty-VLPs), we identified several reverse transcribed cDNA strands. Most of them resemble the characteristic intermediates of the reverse transcription process described for authentic retroviruses: a (−) strong-stop DNA strand covalently bound to an RNA primer, two elongated (−) strands with one or two long terminal repeat (LTR) sequences and a (+) strong-stop DNA. Surprisingly, complete (+) strands and full-length linear duplex Ty DNA could not be detected. The structural features of two additional (÷) strands may indicate some differences between the mechanisms of (+) strand synthesis in Ty and other retrotransposons or retroviruses.
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  • 46
    ISSN: 1617-4623
    Keywords: Yeast ; Arginine ; Sequence ; Regulation ; Control region
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    Notes: Summary In Saccharomyces cerevisiae, the ARG5,6 gene encodes acetylglutamyl-P reductase and acetylglutamate kinase, two arginine anabolic enzymes which are localized in the mitochondria. The synthesis of both enzymes is co-ordinately controlled by arginine and by three regulatory proteins (ARGRI, ARGRII, and ARGRIII). The ARG5,6 gene was cloned by complementation of an arg5 mutant strain. A subclone containing an EcoRI fragment of about 3.2 kb which complements the arginine requirement was sequenced. This 3163 by sequence contains only one long open reading frame of 2589 nucleotides encoding a protein of 863 amino acids. The size of this protein is in agreement with the length of the unique transcript determined by Northern hybridization. The measurements of ARG5,6 mRNA under various regulatory conditions show no correlation with the enzyme levels. As in other arginine biosynthetic and catabolic genes, the regulation by arginine through the three ARGR proteins thus involves a post-transcriptional control mechanism. By in vitro mutagenesis we created point mutations and deletions in the 5′ non-coding region of the ARG5,6 gene which allowed us to define the primary target of ARGR control. Specific regulation involves two regions: one located between the putative TATA element and the transcriptional initiation site and the second between this site and the first ATG.
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  • 47
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    Molecular genetics and genomics 229 (1991), S. 413-420 
    ISSN: 1617-4623
    Keywords: SCO1 ; Cytochrome oxidase ; Membrane protein ; Mitochondria ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The SCO1 gene of Saccharomyces cerevisiae encodes a 30 kDa protein which is specifically required for a post-translational step in the accumulation of subunits 1 and 2 of cytochrome c oxidase (COXI and COXII). Antibodies directed against a β-Gal::SCO1 fusion protein detect SCO1 in the mitochondrial fraction of yeast cells. The SCO1 protein is an integral membrane protein as shown by its resistance to alkaline extraction and by its solubilization properties upon treatment with detergents. Based on the results obtained by isopycnic sucrose gradient centrifugation and by digitonin treatment of mitochondria, SCO1 is a component of the inner mitochondrial membrane. Membrane localization is mediated by a stretch of 17 hydrophobic amino acids in the amino-terminal region of the protein. A truncated SCO1 derivative lacking this segment, is no longer bound to the membrane and simultaneously loses its biological function. The observation that membrane localization of SCO1 is affected in mitochondria of a rho 0 strain, hints at the possible involvement of mitochondrially coded components in ensuring proper membrane insertion.
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  • 48
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    Molecular genetics and genomics 225 (1991), S. 363-368 
    ISSN: 1617-4623
    Keywords: Yeast ; Metallothionein gene ; Cadmium resistance
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    Notes: Summary A 3.3 kb fragment of yeast genomic DNA was isolated by screening a genomic library constructed in the high copy number 2 micron plasmid YEp351 vector for clones capable of enhancing the degree of resistance of Saccharomyces cerevisiae strain MW3070-8B to cadmium. The insert contained two complete copies of the CUP1 gene open reading frame (183 bp), including the upstream promoter sequences (450 bp) with two conserved metal responsive cis-acting elements. Northern analysis showed that addition of cadmium (0.02 μM) or copper (50 μM) to overnight liquid cultures of yeast induced expression of CUP1 transcripts from both chromosomal and plasmid-borne gene copies. The cloned 3.3 kb DNA in a high copy number plasmid restored copper resistance to the sensitive strain LS70-313Δ, deleted for the CUP1 gene (cup1Δ), but failed to restore cadmium resistance. Thus, CUP1 gene expression in yeast appears to be influenced differently by cadmium and copper ions. Resistance to heavy metal poisoning resulted from enhanced gene product levels attributable to amplification of the CUP1 gene as well as to increased transcriptions. Two distinct gene product levels mediate cadmium and copper resistance; a higher gene product level was required to confer cadmium resistance.
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  • 49
    ISSN: 1617-4623
    Keywords: Meiosis ; Sporulation ; Northern hybridization ; Regulatory circuit ; Yeast
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    Notes: Summary The SGA1 gene encoding glucoamylase is specifically expressed late in meiotic development of the yeast Saccharomyces cerevisiae. We found that accumulation of both enzyme activity and transcripts was regulated negatively by both nutritional signals and a haploid-specific negative regulator gene of meiosis, RME1, and positively by the inducer genes for meiosis, IME1 and IME2. To study the role of sequences upstream of the SGA1 gene in its expression and regulation, we generated internal deletions in the 5′ non-coding region of the gene and chimeric genes with portions of the upstream sequence inserted into a reporter gene. By analyzing the expression of these genes, we have identified both a 19 by upstream activation sequence (UAS) and a 49 by negatively regulating element (NRE). The UAS activated transcription with no requirement for heterozygosity at the mating-type locus, but this activation was still under negative control by nutrients. The NRE showed no UAS-like activity but conferred IME2-dependent (or meiosis-specific) expression on a heterologous promoter. These results suggest that meiosis-specific expression of the SGA1 gene is established by a regulatory hierarchy including positive and negative factors, the actions of which are mediated through the two separate upstream regulatory elements, UAS and NRE, respectively. Also, that two independently acting cascades exist for the regulation of SGA1 expression: one transduces both the mating-type and nutritional signals and includes the IME2 product, which acts to relieve the repression through NRE ; and another transduces only the nutritional signal independently of the above pathway and inhibits positive factors acting on UAS.
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  • 50
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    Molecular genetics and genomics 227 (1991), S. 127-136 
    ISSN: 1617-4623
    Keywords: Yeast ; Killer toxin ; Immunity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A cDNA copy of the M2 dsRNA encoding the K2 killer toxin ofSaccharomyces cerevisiae was expressed in yeast using the yeastADH1 promoter. This construct produced K2-specific killing and immunity functions. Efficient K2-specific killing was dependent on the action of the KEX2 endopeptidase and the KEX1 carboxypeptidase, while K2-specific immunity was independent of these proteases. Comparison of the K2 toxin sequence with that of the K1 toxin sequence shows that although they share a common processing pathway and are both encoded by cytoplasmic dsRNAs of similar basic structure, the two toxins are very different at the primary sequence level. Site-specific mutagenesis of the cDNA gene establishes that one of the two potential KEX2 cleavage sites is critical for toxin action but not for immunity. Immunity was reduced by an insertion of two amino acids in the hydrophobic amino-terminal region which left toxin activity intact, indicating an independence of toxin action and immunity.
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  • 51
    ISSN: 1617-4623
    Keywords: Yeast ; Mitochondria ; Frameshift ; Suppression ; Restriction
    Source: Springer Online Journal Archives 1860-2000
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    Notes: Summary The +1 frameshift mutation, M5631, which is located in the gene (oxi1) for cytochrome c oxidase II (COXII) of the yeast mitochondrial genome, is suppressed spontaneously to a remarkably high extent (20%–30%). The full-length wild-type COXII produced as a result of suppression allows the mutant strain to grow with a “leaky” phenotype on non-fermentable medium. In order to elucidate the factors and interactions involved in this translational suppression, the strain with the frameshift mutation was mutated by MnCl2 treatment and a large number of mutants showing restriction of the suppression were isolated. Of 20 mutants exhibiting a strong, restricted, respiration-deficient (RD) phenotype, 6 were identified as having mutations in the mitochondrial genome. Furthermore, genetic analyses mapped one mutation to the vicinity of the gene for tRNAPro and two others to a region of the tRNA cluster where two-thirds of all mitochondrial tRNA genes are encoded. The degree of restriction of the spontaneous frameshift suppression was characterized at the translational level by in vivo 35S-labeling of the mitochondrial translational products and immunoblotting. These results showed that in some of these mutant strains the frameshift suppression product is synthesized to the same extent as in the leaky parent strain. It is suggested that more than one +1 frame-shifted product is made as a result of suppression in these strains: one is as functional as the wild-type COXII, the other(s) is (are) non-functional and prevent leaky growth on non-fermentable medium. A possible mechanism for this heterogenous frameshift suppression is discussed.
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  • 52
    ISSN: 1617-4623
    Keywords: SNF2 sequence ; Transcriptional regulator ; Gene expression ; Glucoamylase ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We have cloned and sequenced the GAM1 gene which is required for transcription of the STA1 gene encoding an extracellular glucoamylase in Saccharomyces cerevisiae var. diastaticus. Complementation tests indicated that GAM1 is the same gene as SNF2 which is required for derepression of the SUC2 gene encoding invertase. Accumulation of SNF2 RNA was not regulated by the GAM2 and GAM3 genes which are also required for STA1 expression. The SNF2 gene was predicted to encode a 194 kDa highly charged protein with a glutamine-rich tract. A bifunctional SNF2-lacZ fusion protein was shown by immunofluorescence microscopy to be localized to the nucleus, suggesting that the SNF2 protein is located in the nucleus.
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  • 53
    ISSN: 1617-4623
    Keywords: Protein kinase ; Yeast ; CDC28 ; Cell cycle
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A novel protein kinase homologue (KNS1) has been identified in Saccharomyces cerevisiae. KNS1 contains an open reading frame of 720 codons. The carboxy-terminal portion of the predicted protein sequence is similar to that of many other protein kinases, exhibiting 36% identity to the cdc2 gene product of Schizosaccharomyces pombe and 34% identity to the CDC28 gene product of S. cerevisiae. Deletion mutations were constructed in the KNS1 gene. kns1 mutants grow at the same rate as wild-type cells using several different carbon sources. They mate at normal efficiencies, and they sporulate successfully. No defects were found in entry into or exit from stationary phase. Thus, the KNS1 gene is not essential for cell growth and a variety of other cellular processes in yeast.
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  • 54
    Electronic Resource
    Electronic Resource
    Springer
    Molecular genetics and genomics 229 (1991), S. 353-356 
    ISSN: 1617-4623
    Keywords: DNA polymerase ; Gene conversion ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary In Saccharomyces cerevisiae, three different DNA polymerase complexes, POLI, POLII and POLIII, are known to be involved in DNA replication. The catalytic subunit of POLIII is encoded by the essential CDC2 gene. The existence of different thermosensitive non-complementing mutants of CDC2 offers the possibility of using a genetic approach to investigate the involvement of POLIII in induced gene conversion. When cdc2 heteroallelic cells were irradiated and incubated under restrictive conditions, almost no induction of thermoresistant cells could be detected, suggesting an essential role for POLIII in mitotic gene conversion events.
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  • 55
    ISSN: 1617-4623
    Keywords: Translational activation ; Cytochrome b ; Mitochondria ; Yeast ; CRS1 ; CBS2
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The products of the nuclear genes CBS1 and CBS2 are both required for translational activation of mitochondrial apocytochrome b in yeast. We report the intramitochondrial localization of both proteins by use of specific antisera. Based on its solubilization properties the CBS1 protein is presumed to be a component of the mitochondrial membrane; the detergent concentrations needed to release CBS1 from mitochondria are almost the same as for cytochrome c 1. In contrast, CBS2 behaves like a soluble protein, with some characteristics of a membrane-associated protein. A model is presented for translational activation of cytochrome b, which might also be applicable to translational regulation of other mitochondrial genes.
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  • 56
    ISSN: 1617-4623
    Keywords: Yeast ; Transcription ; a- and α-specific genes ; MCM1 ; STE12
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We have examined the relative contributions of MCM1 and STE12 to the transcription of the a-specific STE2 gene by using a 367 by fragment from the STE2 5′-noncoding region to drive expression of a reporter lacZ gene. Mutation of the MCM1 binding site destroyed MCM1 · α2-mediated repression in α cells and dramatically reduced expression in a cells. The residual expression was highly stimulated by exposure of cells to pheromone. Likewise, the loss of STE12 function reduced lacZ expression driven by the wild-type STE2 fragment. In the absence of both MCM1 and STE12 functions, no residual expression was observed. Thus, the STE2 fragment appears to contain two distinct upstream activation sequences (UASs), one that is responsible for the majority of expression in cells not stimulated by pheromone, and one that is responsible for increased expression upon pheromone stimulation. In further support of this idea, a chemically synthesized version of the STE2MCM1 binding site had UAS activity, but the activity was neither stimulated by pheromone nor reduced in ste12 mutants. Although transcription of aspecific genes also requires both MCM1 and STE12, these genes differ from a-specific genes in that they have a single, MCM1-dependent UAS system. The activity of the minimal 26 by UAS from the α-specific STE3 gene was both stimulated by pheromone and reduced in ste12 mutants. These data suggest that at α-specific genes STE12 and MCM1 exert their effects through a single UAS.
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  • 57
    ISSN: 1617-4623
    Keywords: Mitochondria ; Yeast ; Protein targeting ; PET2858 ; Inner membrane protease 1
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The nuclear yeast mutant pet ts2858 is defective in the removal of pre-sequences from the mitochondrially encoded cytochrome oxidase subunit II (COXII) and the processing intermediate of cytochrome b 2 (Cytb 2), a nuclear gene product. In order to identify the genetic lesion in this mutant we have cloned and characterized a DNA region which complements the pet ts2858 mutation. The DNA sequence revealed three open reading frames, one of which is responsible for the complementation. A 570 by reading frame represents the structural gene PET2858, as demonstrated by in vitro mutagenesis, gene expression from a foreign promoter, and allelism tests. PET2858 encodes a 21.4 kDa protein, which is essential for growth on non-fermentable carbon sources and for the proteolytic processing of COXII and the Cytb 2 intermediate. When the N-terminus of the PET2858 protein is fused to a reporter protein, the resulting hybrid molecule is imported into mitochondria. Interestingly, the N-terminal half of the deduced PET2858 protein exhibits 30.7% amino acid identity to the leader peptidase of Escherichia coli. These results suggest that PET2858 codes for a mitochondrial inner membrane protease (IMP1) or at least a subunit of it. This protease is involved in protein processing and export from the mitochondrial matrix.
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  • 58
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    Antonie van Leeuwenhoek 59 (1991), S. 125-127 
    ISSN: 1572-9699
    Keywords: Taxonomy ; Williopsis Salicorniae ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Four strains of an undescribed species of the genus Williopsis were isolated from brackish water. A description of the new species, Williopsis salicorniae (type strain, CBS 8071, NRRL Y-12834), is given.
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