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  • evolution  (474)
  • Saccharomyces cerevisiae  (444)
  • Springer  (918)
  • American Association for the Advancement of Science
  • American Meteorological Society
  • Periodicals Archive Online (PAO)
  • 1995-1999  (473)
  • 1990-1994  (445)
  • 1935-1939
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Year
  • 101
    Electronic Resource
    Electronic Resource
    Parental care in terrestrial gastropods (1994)
    Springer
    Cellular and molecular life sciences 50 (1994), S. 5-14 
    Details
    ISSN: 1420-9071
    Keywords: parental investment ; juvenile survival ; evolution ; gastropods ; molluscs ; ovoviviparity ; viviparity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Parental care in terrestrial gastropods includes the of oviposition sites, production of large, heavily-yolked eggs supplied with calcium carbonate, provisioning of hatchings with eggs in specis with facultative sibling cannibalism, egg retention, and ovoviviparity. Evidence for true viviparity is scarce in terrestrial gastropods, as it is for postlaying care of eggs, though external egg carrying on the shell occurs in a few species. Care of young has not been observed in any terrestrial gastropod species. Provisioning of eggs with nutrients and calcium carbonate might be the most common form of parental investment. Ovoviviparity allows terrestrial gastropods to persist in habitats otherwise unsuitable for oviparous species (e.g. exposed rock walls). An interspecific comparison demonstrates that egg-retaining and ovoviviparous species produce smaller clutches than oviparous species and suggests a cost of parental care.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF01992042
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  • 102
    Electronic Resource
    Electronic Resource
    Evidence for an HLA-C-like locus in the orangutan Pongo pygmaeus (1999)
    Springer
    Immunogenetics 49 (1999), S. 865-871 
    Details
    ISSN: 1432-1211
    Keywords: Key words Orangutan ; MHC class I ; HLA-C ; natural killer cells ; evolution
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  HLA-B and C are related class I genes which are believed to have arisen by duplication of a common ancestor. Previous study showed the presence of orthologues for both HLA-B and C in African apes but only for HLA-B in Asian apes. These observations suggested that the primate C locus evolved subsequent to the divergence of the Pongidae and Hominidae. From an analysis of orangutan Tengku two HLA-C-like alleles (Popy C*0101 and Popy C*0201) were defined as well as three HLA-B-like (Popy-B) alleles. By contrast, no Popy-C alleles were obtained from orangutan Hati, although three Popy-B alleles were defined. Thus an HLA-C-like locus exists in the orangutan (as well as a duplicated B locus), implying that the primate C locus evolved prior to the divergence of the Pongidae and Hominidae and is at least 12–13 million years old. Uncertain is whether all orangutan MHC haplotypes contain a C locus, as the failure to find C alleles in some individuals could be due to a mispairing of HLA-C-specific primers with certain Popy-C alleles. These results raise the possibilities that other primate species have a C locus and that the regulation of natural killer cells by C allotypes evolved earlier in primate evolution than has been thought.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s002510050566
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  • 103
    Electronic Resource
    Electronic Resource
    DNA integration into recipient yeast chromosomes by trans-kingdom conjugation between Escherichia coli and Saccharomyces cerevisiae (1992)
    Springer
    Current genetics 21 (1992), S. 101-108 
    Details
    ISSN: 1432-0983
    Keywords: Trans-kingdom conjugation ; DNA integration ; Saccharomyces cerevisiae ; Escherichia coli
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary IncQ-derived conjugative shuttle vectors, which carried the yeast gene URA3 and/or the yeast autonomously replicating sequence (ARS1), were constructed. Both the ars-plus plasmid pAY205 and the ars-less plasmid pAY201 were successfully transmitted from E. coli to S. cerevisiae by the action of mob and tra. In this trans-kingdom conjugation, plasmid pAY205 could replicate and be retained in transconjugants. Plasmid pAY201 caused the formation of “micro-colonies” of abortive transconjugants due to its transient expression and rapid disappearance. Nevertheless, one per about 103 colonies caused by transmitted pAY201 plasmids were uncurable by integration into the homologous region of a yeast chromosome. Analyses by restriction enzyme mapping and Southern hybridization indicate that this integration is primarily caused by a double crossover during conjugation and not by a single reciprocal recombination.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00318467
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  • 104
    Electronic Resource
    Electronic Resource
    The PAR1 (YAP1/SNQ3) gene of Saccharomyces cerevisiae, ac-jun homologue, is involved in oxygen metabolism (1992)
    Springer
    Current genetics 21 (1992), S. 269-273 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Transcriptional activator ; Oxidative stress ; Glutathione
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The PAR1/SNQ3 gene of S. cerevisiae, which increases resistance to iron chelators in multi-copy transformants, is identical to the YAP1 gene, a yeast activator protein isolated as a functional homologue of the human c-jun oncogene by binding specifically to the AP-1 consensus box. The observed H2O2-sensitivity of par1 mutants has been attributed to an increased sensitivity to reduced oxygen intermediates. Accordingly, par1 mutants did not survive an elevated oxygen pressure and were very sensitive to menadione and methylviologene, two chemicals enhancing the deleterious effects of oxygen. The specific activities of enzymes involved in oxygen detoxification, such as superoxide dismutase, glucose 6-phosphate dehydrogenase and glutathione reductase, were decreased in par1 mutants and increased after PAR1 over-expression. As in the case of oxygen detoxification enzymes, the cellular levels of glutathione were similarly affected. These observations indicate that PAR1/YAP1/SNQ3 is involved in the gene regulation of certain oxygen detoxification enzymes. The finding that H2O2 promotes DNA-binding of human c-jun is consistent with a similar function for PAR1/YAP1/SNQ3 and c-jun in cellular metabolism.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00351681
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  • 105
    Electronic Resource
    Electronic Resource
    An yeast nuclear mutation conferring temperature-sensitivity to the mitochondrial tryptophanyl-tRNA synthetase (1992)
    Springer
    Current genetics 21 (1992), S. 281-283 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Mitochondrial trp-tRNA synthetase ; Nuclear mutation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The conditional respiratory-deficient Saccharomyces cerevisiae mutant pet-ts2281 was complemented by an yeast genomic DNA library. The gene thus isolated was sequenced and proved to be identical to the known MSW1 sequence encoding mitochondrial tryptophanyl-tRNA synthetase (Myers and Tzagoloff 1985). Compared to the wild-type, the ts2281 mutant allele of MSW1 contained a single T→C transition leading to a Leu→Ser replacement at position 294 of the protein sequence. In addition to this mutational alteration, our sequence data for the wild-type gene differ from the originally published MSW1 sequence at five other DNA positions which affect two locally restricted regions of the polypeptide chain. As expected, at the non-permissive temperature ts2281 cells are specifically defective in mitochondrial trp-tRNA formation and, thus, in overall mitochondrial protein synthesis. In addition, the patterns of cytochrome b mRNA maturation intermediates were distinctly different in ts2281 and wild-type yeast cells. The mutational effect of the observed amino-acid substitution in ts2281 is discussed in terms of weakened hydrogen bonding in the C-terminal half of the MSW1-encoded protein.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00351683
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  • 106
    Electronic Resource
    Electronic Resource
    Cloning and expression of Hormoconis resinae glucoamylase P cDNA in Saccharomyces cerevisiae (1993)
    Springer
    Current genetics 24 (1993), S. 38-44 
    Details
    ISSN: 1432-0983
    Keywords: Glucoamylase ; Gene cloning ; Hormoconis resinae ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A cDNA coding for glucoamylase P of Hormoconis resinae was cloned using a synthetic oligonucleotide probe coding for a peptide fragment of the purified enzyme and polyclonal anti-glucoamylase antibodies. Nucleotide-sequence analysis revealed an open reading frame of 1848 base pairs coding for a protein of 616 amino-acid residues. Comparison with other fungal glucoamylase amino-acid sequences showed homologies of 37–48%. The glucoamylase cDNA, when introduced into Saccharomyces cerevisiae under the control of the yeast ADC1 promoter, directed the secretion of active glucoamylase P into the growth medium.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00324663
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  • 107
    Electronic Resource
    Electronic Resource
    Mistranslation of human phosphoglycerate kinase in yeast in the presence of paromomycin (1994)
    Springer
    Current genetics 26 (1994), S. 95-99 
    Details
    ISSN: 1432-0983
    Keywords: Translational fidelity ; Paromomycin ; Stuttering ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Missense errors in the translation of mRNAs in Saccharomyces cerevisiae were screened by looking for charge heterogeneity of proteins on two-dimensional gels resulting from the substitution of charged and neutral amino acids. No such mistranslation was detected in wild-type yeast strains grown in the presence of the translational error-inducing antibiotic paromomycin. However, paromomycin-induced mistranslation of a heterologous mRNA, encoding human phosphoglycerate kinase expressed in yeast, was seen. We suggest that the combination of error-prone translation of a heterologous mRNA, and growth in the presence of paromomycin, leads to an accumulation of mistranslated proteins that can be detected by two-dimensional gel electrophoresis.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00313794
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  • 108
    Electronic Resource
    Electronic Resource
    Normal mitochondrial structure and genome maintenance in yeast requires the dynamin-like product of the MGM1 gene (1993)
    Springer
    Current genetics 24 (1993), S. 141-148 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Dynamin ; Mitochondria ; GTP binding protein
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The isolation and characterization of MGM1, and yeast gene with homology to members of the dynamin gene family, is described. The MGM1 gene is located on the right arm of chromosome XV between STE4 and PTP2. Sequence analysis revealed a single open reading frame of 902 residues capable of encoding a protein with an approximate molecular mass of 101 kDa. Loss of MGM1 resulted in slow growth on rich medium, failure to grow on non-fermentable carbon sources, and loss of mitochondrial DNA. The mitochondria also appeared abnormal when visualized with an antibody to a mitochondrial-matrix marker. MGM1 encodes a dynamin-like protein involved in the propagation of functional mitochondria in yeast.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00324678
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  • 109
    Electronic Resource
    Electronic Resource
    Saccharomyces cerevisiae YDR1, which encodes a member of the ATP-binding cassette (ABC) superfamily, is required for multidrug resistance (1994)
    Springer
    Current genetics 26 (1994), S. 285-294 
    Details
    ISSN: 1432-0983
    Keywords: ABC superfamily ; Multidrug resistance ; Saccharomyces cerevisiae ; YDR1 gene
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A multidrug resistance gene, YDR1, of Saccharomyces cerevisiae, which encodes a 170-kDa protein of a member of the ABC superfamily, was identified. Disruption of YDR1 resulted in hypersensitivity to cycloheximide, cerulenin, compactin, staurosporine and fluphenazine, indicating that YDR1 is an important determinant of cross resistance to apparently-unrelated drugs. The Ydr1 protein bears the highest similarity to the S. cerevisiae Snq2 protein required for resistance to the mutagen 4-NQO. The drug-specificity analysis of YDR1 and SNQ2 by gene disruption, and its phenotypic suppression by the overexpressed genes, revealed overlapping, yet distinct, specificities. YDR1 was responsible for cycloheximide, cerulenin and compactin resistance, whereas, SNQ2 was responsible for 4-NQO resistance. The two genes had overlapping specificities toward staurosporine and fluphenazine. The transcription of YDR1 and SNQ2 was induced by various drugs, both relevant and irrelevant to the resistance caused by the gene, suggesting that drug specificity can be mainly attributed to the functional difference of the putative transporters. The transcription of these genes was also increased by heat shock. The yeast drug-resistance system provides a novel model for mammalian multidrug resistance.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00310491
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  • 110
    Electronic Resource
    Electronic Resource
    Sequence and localization of the gene encoding yeast phosphoglycerate mutase (1991)
    Springer
    Current genetics 20 (1991), S. 167-171 
    Details
    ISSN: 1432-0983
    Keywords: Glycolysis ; Repetitive elements τ/δ ; Promoter ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary In this study we report on the complete nucleotide sequence of the yeast phosphoglycerate mutase gene (GPM1) and its essential 5′ and 3′ non-coding regions. The transcriptional start points were determined by S1-mapping and sequencing of a cDNA clone. Several sequences identified as important for transcriptional regulation in yeast promoters are present upstream of the transcription start point. 3′ to the coding region we sequenced a composite repetitive element which, apparently, originated from a recombination between a delta-and a tau-element. Finally, we mapped the GPM1 gene 13 cM distal to fas1 on chomosome XI.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00312781
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  • 111
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    Molecular cloning of the PEL1 gene of Saccharomyces cerevisiae that is essential for the viability of petite mutants (1993)
    Springer
    Current genetics 24 (1993), S. 307-312 
    Details
    ISSN: 1432-0983
    Keywords: Growth control ; Genetic mapping ; Molecular cloning ; Nucleo-mitochondrial interaction ; Saccharomyces cerevisiae ; Viability of petites
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The PEL1 gene of Saccharomyces cerevisiae is essential for the cell viability of mitochondrial petite mutants, for the ability to utilize glycerol and ethanol on synthetic medium, and for cell growth at higher temperatures. By tetrad analysis the gene was assigned to chromosome III, centromere proximal of LEU2. The PEL1 gene has been isolated and cloned by the complementation of a pel1 mutation. The molecular analysis of the chromosomal insert carrying PEL1 revealed that this gene corresponds to the YCL4W open reading frame on the complete DNA sequence of chromosome III. The putative Pel1 protein is characterized by a low molecular weight of approximately 17 kDa, a low codon adaptation index, and a high leucine content.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00336781
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  • 112
    Electronic Resource
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    Sequence analysis of a Papaver somniferum L. mitochondrial DNA fragment promoting autonomous plasmid replication in Saccharomyces cerevisiae and Kluyveromyces lactis (1993)
    Springer
    Current genetics 24 (1993), S. 366-367 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Papaver somniferum L. ; ARS ; Mitochondrial DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The minimal fragment of mitochondrial DNA from Papaver somniferum L. (poppy) able to promote autonomous plasmid replication in the yeast Saccharomyces cerevisiae was sequenced. Sequence analysis of the 917-bp MK4/8 DNA fragment revealed a high AT content, and the presence of two 12-bp sequences differing from the ARS core consensus of S. cerevisiae only by a T and C insertion, respectively. The mitochondrial insert contains a further six 11-bp sequences with one mismatch to the S. cerevisiae core consensus, more then 20 related sequences with two base pair exchanges, numerous direct and inverted repeats, and many copies of a sequence motif called the ARS box. The original 4.2-kb mitochondrial DNA fragment, as well as the minimal 917-bp subfragment in vector pFL1-E (a variant of YIP5, lacking an origin of replication in yeast), were then tested for their ability to replicate autonomously in another fungus, Kluyveromyces lactis.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00336790
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  • 113
    Electronic Resource
    Electronic Resource
    The ogd1 and kgd1 mutants lacking 2-oxoglutarate dehydrogenase activity in yeast are allelic and can be differentiated by the cloned amber suppressor (1993)
    Springer
    Current genetics 24 (1993), S. 377-381 
    Details
    ISSN: 1432-0983
    Keywords: 2-Oxoglutarate dehydrogenase ; Molecular cloning ; Saccharomyces cerevisiae ; Sequencing ; Suppressor ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The activity of mitochondrial 2-oxoglutarate dehydrogenase in S. cerevisiae can be impaired either by the ogd1 or the kgd1 mutation. The OGD1 gene and two suppressor genes were isolated by complementation of the ogd1 mutant. The complementation of the kdg1 mutant by the OGD1 gene, an allelism test, and meiotic mapping, revealed that the ogd1 and kgd1 mutations are allelic. The two mutations were differentiated by the cloned suppressor gene which was able to partially complement ogd1, but not kgd1. The molecular analysis of the suppressor gene revealed its identity with the natural tRNA CAG Gln gene found in the upstream region of URA10.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00351844
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  • 114
    Electronic Resource
    Electronic Resource
    Starvation for His-tRNAHis in yeast causes translational arrest without a high level of misincorporation of glutamine at histidine codons (1992)
    Springer
    Current genetics 21 (1992), S. 177-182 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Aminoacyl-tRNA synthetase mutant ; PGK overexpression ; In vivo misreading
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The hts1.1 temperature-sensitive histidinyl-tRNA synthetase mutation enables Saccharomyces cerevisiae to be starved for His-tRNAHis by upshift to the non-permissive temperature of 38°C. If yeast behaves similarly to bacterial and mammalian cells, this lack of His-tRNAHis should greatly enhance misreading at histidine codons (CAU/CAC) by Gln-tRNAGln, resulting in substitution of the neutral amino acid glutamine in place of histidine, a basic amino acid. Such misreading causes the isoelectric point (pI) of proteins to shift to lower values, and is readily detectable as “stuttering” on two-dimensional (2D) protein gels. By gel analysis of pulse-labelled proteins of hts1.1 yeast cells that were overexpressing phosphoglycerate kinase (PGK), our study sought to detect this specific translational error in PGK protein. It was not detected by this relatively sensitive technique, indicating that missense errors due to glutamine insertion at histidine codons do not occur in yeast at the readily-detectable level found in bacterial and mammalian cells.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00336838
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  • 115
    Electronic Resource
    Electronic Resource
    Use of reporter genes for the isolation and characterisation of different classes of sporulation mutants in the yeast Saccharomyces cerevisiae (1993)
    Springer
    Current genetics 24 (1993), S. 451-454 
    Details
    ISSN: 1432-0983
    Keywords: Yeast ; Saccharomyces cerevisiae ; Sporulation mutants ; Reporter genes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Reporter genes consisting of sporulation-specific promoters fused to lacZ were used as markers to monitor the sporulation pathway of the yeast Saccharomyces cerevisiae. Strains transformed with these lacZ gene fusions expressed β-galactosidase (assayable on plates using the substrate 5-bromo-4-chloro-3-indolyl-β-D-galactopyranoside, X-gal) in a sporulation-dependent manner. Mutagenesis experiments performed on transformed strains resulted in the recovery of a number of novel sporulation mutants. Three classes of mutants were obtained: those which overexpressed the reporter gene under sporulation conditions, those which did not express the gene under any conditions, and those which expressed the gene in vegetative cells not undergoing sporulation. On the basis of the blue colony-colour produced in the presence of X-gal these have been described as superblue, white, and blue vegetative mutants, respectively. These were further characterised using earlier reporter genes and other marker systems. This study established that the multicopy reporter plasmids chosen do not interfere with sporulation; they are valid tools for monitoring the pathway and they provide a way to isolate mutations not readily selected by other markers.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00351856
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  • 116
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    Physical mapping of the MEL gene family in Saccharomyces cerevisiae (1993)
    Springer
    Current genetics 24 (1993), S. 461-464 
    Details
    ISSN: 1432-0983
    Keywords: Chromosome fragmentation ; MEL gene family ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Nine members, MEL2–MEL10, of the MEL gene family coding for α-galactosidase were physically mapped to the ends of the chromosomes by chromosome fragmentation. Genetic mapping of the genes supported the location of all the MEL genes in the left arm of their resident chromosomes.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00351706
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  • 117
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    Parameters affecting lithium acetate-mediated transformation of Saccharomyces cerevisiae and development of a rapid and simplified procedure (1993)
    Springer
    Current genetics 24 (1993), S. 455-459 
    Details
    ISSN: 1432-0983
    Keywords: Yeast ; Saccharomyces cerevisiae ; Transformation ; Plasmid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have compared a number of procedures for the transformation of whole cells of the yeast Saccharomyces cerevisiae and assessed the effects of dimethylsulphoxide (DMSO) or ethanol, both of which have been reported to enhance transformation efficiency. We find that simplified methods benefit from the addition of one of these compounds, and although differences are observed between strains as to the more beneficial reagent, peak transformation efficiency is, in general obtained with 10% DMSO or 10% EtOH. Increases of between six- and 50-fold are observed, despite a reduction in cell viability, and at this concentration the two compounds are not additive in their effects. The optimum level appears to depend on a balance between improved DNA uptake and reduced cell viability. As a result of this work we present a straightforward and rapid transformation procedure.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00351857
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  • 118
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    Genetic effects of photoactivated psoralens during meiosis in DNA repair mutantpso3-1 ofSaccharomyces cerevisiae (1994)
    Springer
    Current genetics 25 (1994), S. 19-23 
    Details
    ISSN: 1432-0983
    Keywords: Psoralen ; DNA repair mutants ; Gene conversion ; Recombination ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The influence of the DNA repair genePSO3 on photoactivated psoralen-induced meiotic recombination, gene conversion, reverse mutation, and on survival, was assayed in diploid strains ofSaccharomyces cerevisiae homozygous for the wild-type or thepso3-1 mutant allele. Sporulation was normal in thepso3-1 diploid. Wild-type and mutant strains had the same sensitivity to photoactivated monofunctional psoralen (3-CPs+UVA) in meiosis-uncommitted and meiosis-committed stages. The mutant showed higher sensitivity to photoactivated bifunctional psoralen (8-MOP+UVA) during all stages of the meiotic cycle. Mutation induction by 3-CPs+UVA or 8-MOP+UVA in meiosis-committed cells revealed no significant differences between wild-type and thepso3-1 mutant. The status of thePSO3 gene has no influence on the kinetics of induction of gene conversion and crossing-over after 3-CPs+UVA treatment in meiosis-committed cells: gene conversion was blocked while recombination was induced. After treatment with 8-MOP+UVA gene conversion was also blocked in both strains while crossing-over could only be observed in meiosis-committed wild-type cells.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00712961
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  • 119
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    Expression of a yeast gene can be blocked by insertion of short yeast DNA fragments between a UAS and the TATA box (1995)
    Springer
    Current genetics 27 (1995), S. 387-389 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; URS ; FBP1 Transcription
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have constructed a plasmid, pOV10, which facilitates the introduction of putative upstream activating sequences (UAS) or upstream repressing sequences (URS) from yeast genes into plasmids containing CYC1-lacZ fusions. We have observed that the insertion of yeast sequences from 155 to 195 bp between the UAS and the TATA box of a CYC1-lacZ fusion gene can block β-galactosidase expression. It is suggested that this block is related to the formation of nucleosomes on the DNA.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00352109
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  • 120
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    NCA3, a nuclear gene involved in the mitochondrial expression of subunits 6 and 8 of the Fo-F1 ATP synthase of S. cerevisiae (1995)
    Springer
    Current genetics 27 (1995), S. 409-416 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Mitochondrial synthesis ; Nuclear control ; F1Fo-ATPase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Respiratory-competent nuclear mutants have been isolated which presented a cryosensitive phenotype on a non-fermentative carbon source, due to a dysfunctioning of the mitochondrial F1-Fo ATP synthase which results from a relative defect in subunits 6 and 8 of the Fo sector. Both proteins are mtDNA-encoded, but the defect is due to the simultaneous presence of a mutation in two unlinked nuclear genes (NCA2 and NCA3, for Nuclear Control of ATPase) promoting a modification of the expression of the ATP8-ATP6 co-transcript (formerly denoted AAP1-OLI2). This co-transcript matures at a unique site to give two co-transcripts of 5.2 and 4.6 kb in length: in the mutant, the 5.2-kb co-transcript was greatly lowered. NCA3 was isolated from a wild-type yeast genomic library by genetic complementation. The level of the 5.2-kb transcript, like the synthesis of subunits 6 and 8, was partly restored in the transformed strain. A 1011-nucleotide ORF was identified that encodes an hydrophilic protein of 35417 Da. Disruption of chromosomal DNA within the reading frame promoted a dramatic decrease of the 5.2-kb mRNA but did not abolish the respiratory competence of a wild-type strain. NCA3 is located on chromosome IV and produces a single 1780-b transcript.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00311209
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  • 121
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    Failure to detect an antimutator phenotype following disruption of the Saccharomyces cerevisiae DDR48 gene (1995)
    Springer
    Current genetics 27 (1995), S. 496-500 
    Details
    ISSN: 1432-0983
    Keywords: Antimutator ; DDR48 ; Saccharomyces cerevisiae ; Spontaneous mutation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The antimutator phenotype, reportedly conferred by disruption of the Saccharomyces cerevisiae DDR48 gene, was suggested to affect only a specific spontaneous mutational pathway. We attempted to identify the types of mutation that are DDR48-dependent by determining the specificity of the ddr48 antimutator. However, disruption of DDR48 did not decrease the rates of spontaneous forward mutation in a plasmid-borne copy of the yeast SUP4-o gene, the reversion or suppression of the lys2–1 allele, or forward mutation at the CAN1 locus. Interestingly, the latter gene had been reported previously to be subject to the antimutator effect. DNA sequence analysis of spontaneous SUP4-o mutations arising in DDR48 and ddr48 backgrounds provided no evidence for a reduction in the rates of individual mutational classes. Thus, we were unable to verify that disruption of DDR48 causes an antimutator phenotype.
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    Purification and binding properties of the Mal63p activator of Saccharomyces cerevisiae (1995)
    Springer
    Current genetics 27 (1995), S. 509-516 
    Details
    ISSN: 1432-0983
    Keywords: Yeast ; Maltose fermentation ; MAL63 ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Mal63p is a transcriptional activator for maltose fermentation in Saccharomyces cerevisiae. We have purified it to homogeneity from a yeast strain in which the MAL63 gene is under the control of the GAL1–GAL10 promoter. Purification included fractionation of a whole-cell extract by ion-exchange chromatography, chromatography using both non-specific DNA-affinity (calf thymus), and sequence-specific DNA-affinity chromatography. Mal63p activity was assayed by its binding to a fragment of the MAL61–MAL62 promoter, using both filter-binding and electrophoretic-mobility shift assays. DNase-I footprinting identified a new binding site (site 3) between the two previously known sites (sites 1 and 2). Mal63p is a dimer, and methylation-protection experiments identify the recognition motif as: c/a GC N9 c/a GC/g.
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    Sensitivity of sup35 and sup45 suppressor mutants in Saccharomyces cerevisiae to the anti-microtubule drug benomyl (1996)
    Springer
    Current genetics 30 (1996), S. 44-49 
    Details
    ISSN: 1432-0983
    Keywords: Key words Omnipotent suppression ; Microtubules ; Respiratory deficiency ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  SUP35 and SUP45 genes determine the accuracy of translation at the stage of termination. We present indirect evidence indicating that these genes may also control some cellular process mediated by microtubules. A majority of sup35 and sup45 suppressor mutations confer supersensitivity to benomyl, the drug which de-polymerizes microtubules. In addition, data correlating phenotypic manifestations of sup45 suppressor mutations, involving sensitivity to benomyl, respiratory deficiency and a suppressor effect, are also presented.
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    URL: http://dx.doi.org/10.1007/s002940050098
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    Cloning and characterization of the first two genes of the non-oxidative part of the Saccharomyces cerevisiae pentose-phosphate pathway (1996)
    Springer
    Current genetics 30 (1996), S. 404-409 
    Details
    ISSN: 1432-0983
    Keywords: Key words D-ribulose-5-phosphate 3-epimerase ; D-ribose-5-phosphate ketol-isomerase ; Pentose-phosphate pathway ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have cloned and characterized the two remaining unknown genes of the non-oxidative part of the pentose-phosphate pathway of Saccharomyces cerevisiae encoding the enzymes D-ribulose-5-phosphate 3-epimerase (Rpe1p) and D-ribose-5-phosphate ketol-isomerase (Rki1p). Rpe1p has an unexpected high specific activity of 2148 mU × (mg protein)–1 in crude extracts. Deletion mutants of RPE1 show no enzyme activity and are unable to grow on D-xylulose. Unexpectedly, haploid rki1 deletion mutants are not viable. Functional expression of RKI1 was demonstrated following an increase of gene dosage in the haploid rki1 deletion mutant, which restored viability and specific D-ribose-5-phosphate ketol-isomerase activity. Both enzymes show high similarity to the deduced protein sequences of various open reading frames, expressed sequence tags or cDNAs from different organisms.
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    The repair of DNA methylation damage in Saccharomyces cerevisiae (1996)
    Springer
    Current genetics 30 (1996), S. 461-468 
    Details
    ISSN: 1432-0983
    Keywords: Keywords DNA repair ; Methylation damage ; Epistasis analysis ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  The major genotoxicity of methyl methanesulfonate (MMS) is due to the production of a lethal 3-methyladenine (3MeA) lesion. An alkylation-specific base-excision repair pathway in yeast is initiated by a Mag1 3MeA DNA glycosylase that removes the damaged base, followed by an Apn1 apurinic/ apyrimidinic endonuclease that cleaves the DNA strand at the abasic site for subsequent repair. MMS is also regarded as a radiomimetic agent, since a number of DNA radiation-repair mutants are also sensitive to MMS. To understand how these radiation-repair genes are involved in DNA methylation repair, we performed an epistatic analysis by combining yeast mag1 and apn1 mutations with mutations involved in each of the RAD3, RAD6 and RAD52 groups. We found that cells carrying rad6, rad18, rad50 and rad52 single mutations are far more sensitive to killing by MMS than the mag1 mutant, that double mutants were much more sensitive than either of the corresponding single mutants, and that the effects of the double mutants were either additive or synergistic, suggesting that post-replication and recombination-repair pathways recognize either the same lesions as MAG1 and APN1, or else some differ- ent lesions produced by MMS treatment. Lesions handled by recombination and post replication repair are not simply 3MeA, since over-expression of the MAG1 gene does not offset the loss of these pathways. Based on the above analyses, we discuss possible mechanisms for the repair of methylation damage by various pathways.
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    The PSO4 gene of S. cerevisiae is important for sporulation and the meiotic DNA repair of photoactivated psoralen lesions (1995)
    Springer
    Current genetics 27 (1995), S. 207-212 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; pso4-1 mutant Sporulation ; DNA repair ; Meiotic recombination Induced mutagenesis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have evaluated the effect of the Saccharomyces cerevisiae pso4-1 mutation in sporulation and DNA repair during meiosis. We have found that pso4-1 cells were arrested in an early step of meiosis, before premeiotic DNA synthesis, and hence did not produce spores. These results suggest that the PSO4 gene may act at the start point of the cell cycle, as do some SPO and CDC genes. The pso4-1 mutant cells are specifically sensitive to 8-MOP- and 3-CPs-photoinduced lesions, and are found to be severely affected in meiotic recombination as well as impaired in the mutagenic response, as previously described for mitosis. This means that the PSO4 gene is important for the repair 8-MOP-photoinduced lesions, mainly double-strand breaks, and the processing of these lesions into recombinogenic intermediates.
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    Determination of chromosome copy numbers in Saccharomyces cerevisiae strains via integrative probe and blot hybridization techniques (1995)
    Springer
    Current genetics 27 (1995), S. 217-228 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Chromosome copy numbers ; Ploidy probes ; Industrial yeasts
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Methods have been devised for analyzing chromosome copy numbers in S. cerevisiae strains that may be polyploid or aneuploid, as is apparent in the case of many industrial strains. The initial step involved transformation of a strain with an integrative “ploidy probe” transplacement fragment that enable the copy number of the targeted chromosomal locus to be determined via genomic Southern blotting and quantitative probe hybridization. Dual probe co-hybridization to Southern genomic DNA blots was used to extend such locus copy number determinations to other loci within the same chromosome, thereby screening for internal consistency along the length of the chromosome. This approach was also used to extend the analysis to other chromosomes in the genome. The method was established and verified with euploid series laboratory strains and then used to examine chromosome copy numbers in three industrial strains. One brewing strain apparently contained three copies of the chromosomes tested, whilst another brewing and a baking strain showed evidence of aneuploidy.
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    Influence of gene dosage and autoregulation of the regulatory genes INO2 and INO4 on inositol/choline-repressible gene transcription in the yeast Saccharomyces cerevisiae (1997)
    Springer
    Current genetics 31 (1997), S. 462-468 
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    ISSN: 1432-0983
    Keywords: Key words Transcriptional regulation ; Phospholipid biosynthesis ; Saccharomyces cerevisiae ; INO2
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Expression of structural genes of phospholipid biosynthesis in yeast is mediated by the inositol/choline-responsive element (ICRE). ICRE-dependent gene activation, requiring the regulatory genes INO2 and INO4, is repressed in the presence of the phospholipid precursors inositol and choline. INO2 and, to a less extent, INO4 are positively autoregulated by functional ICRE sequences in the respective upstream regions. However, an INO2 allele devoid of its ICRE functionally complemented an ino2 mutation and completely restored inositol/choline regulation of Ino2p-dependent reporter genes. Low-level expression of INO2 and INO4 genes, each under control of the heterologous MET25 promoter, did not alter the regulatory pattern of target genes. Thus, upstream regions of INO2 and INO4 are not crucial for transcriptional control of ICRE-dependent genes by inositol and choline. Interestingly, over-expression of INO2, but not of INO4, counteracted repression by phospholipid precursors. Possibly, a functional antagonism between INO2 and a negative regulator is the key event responsible for repression or de-repression.
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    Secretion of an enzymatically activeTrichoderma harzianum endochitinase bySaccharomyces cerevisiae (1996)
    Springer
    Current genetics 29 (1996), S. 404-409 
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    ISSN: 1432-0983
    Keywords: Biocontrol ; Secretion ; Chitinase ; Expression cloning ; Saccharomyces cerevisiae ; Trichoderma harzianum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A novel endochitinase agar-plate assay has been developed and used to identify 11 full-length cDNAs encoding endochitinase I (ENC I) from aTrichoderma harzianum cDNA library by expression in yeast. The 1473-bpchil cDNA encodes a 424-residue precursor protein including both a signal sequence and a propeptide. The deduced ENC I amino-acid sequence is homologous to other fungal and bacterial chitinases, and the enzyme cross-reacts with a polyclonal antiserum raised against chitinase A1 fromBacillus circulans. TheT. harzianum endochitinase I was secreted into the culture medium by the yeastSaccharomyces cerevisiae in a functionally active form. The purified recombinant enzyme had a molecular mass of 44 kDa, an isoelectric point of 6.3, a pH optimum of 7.0 and a temperature optimum of 20 °C.
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    Cloning and expression of an Aspergillus kawachii endo-1,4-β-xylanase gene in Saccharomyces cerevisiae (1995)
    Springer
    Current genetics 28 (1995), S. 467-473 
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    ISSN: 1432-0983
    Keywords: Aspergillus kawachii ; β-xylanase ; Expression ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract First-strand cDNA was prepared from mRNA isolated from Aspergillus kawachii IFO4308 and the β-xylanase gene (xynC) amplified by using the polymerase chain reaction (PCR) technique. This gene was inserted between the yeast phosphoglycerate kinase (PGK1) gene promoter (PGK1 p) and terminator (PGK1 T) sequences. The PGK1 P-xynC-PGK1 T construct (designated XYN3) was cloned into a multicopy episomal plasmid and the XYN3 gene was expressed in Saccharomyces cerevisiae. Functional β-xylanase (Xyn3) was produced and secreted by the recombinant yeast. Xyn3 was stable between 30 and 50°C, and the optimum temperature and pH were shown to be at 60°C and lower than pH3, respectively. An autoselective fur1::LEU2 XYN3 recombinant strain was developed that allowed β-xylanase production at a level of 300 nkat/ml in a non-selective complex medium.
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    URL: http://dx.doi.org/10.1007/BF00310817
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    Cysteine uptake by Saccharomyces cerevisiae is accomplished by multiple permeases (1999)
    Springer
    Current genetics 35 (1999), S. 609-617 
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    ISSN: 1432-0983
    Keywords: Key words Cysteine uptake ; Amino-acid permeases ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Uptake by Saccharomyces cerevisiae of the sulphur-containing amino acid L-cysteine was found to be non-saturable under various conditions, and uptake kinetics suggested the existence of two or more transport systems in addition to the general amino-acid permease, Gap1p. Overexpression studies identified BAP2, BAP3, AGP1 and GNP1 as genes encoding transporters of cysteine. Uptake studies with disruption mutants confirmed this, and identified two additional genes for transporters of cysteine, TAT1 and TAT2, both very homologous to BAP2, BAP3, AGP1 and GNP1. While Gap1p and Agp1p appear to be the main cysteine transporters on the non-repressing nitrogen source proline, Bap2p, Bap3p, Tat1p, Tat2p, Agp1p and Gnp1p are all important for cysteine uptake on ammonium-based medium. Furthermore, whereas Bap2p, Bap3p, Tat1p and Tat2p seem most important under amino acid-rich conditions, Agp1p contributes significantly when only ammonium is present, and Gnp1p only contributes under the latter condition.
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    Gene-conversion tract directionality is influenced by the chromosome environment (1998)
    Springer
    Current genetics 34 (1998), S. 269-279 
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    ISSN: 1432-0983
    Keywords: Key words Double-strand breaks ; Heteroduplex DNA ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Spontaneous and double-strand break (DSB)-induced gene conversion in Saccharomyces cerevisiae was assayed using non-tandem chromosomal direct repeat crosses and plasmid × chromosome crosses. Each cross involved identical ura3 alleles marked with phenotypically silent restriction fragment length polymorphic (RFLP) mutations at approximately 100-bp intervals. DSBs introduced in vivo at HO sites in one allele stimulated recombination to Ura+ by more than two orders of magnitude. Spontaneous gene-conversion products were isolated from a related strain lacking a functional HO nuclease gene. The multiple markers did not appear to influence the frequency of direct repeat deletions for spontaneous or DSB-induced events. DSB-induced conversion reflected efficient mismatch repair of heteroduplex DNA. Conversion frequencies of equidistant markers on opposites sides of the DSB were similar in the direct repeat cross. In contrast, markers 5′ of the DSB (promoter-proximal) converted more often than 3′ markers in plasmid × chromosome crosses, a possible consequence of crossing-over associated with long conversion tracts. With direct repeats, bidirectional tracts (extending 5′ and 3′ of the DSB) occurred twice as often as in a plasmid × chromosome cross in which DSBs were introduced into the plasmid-borne allele. A key difference between the direct-repeat and plasmid×chromosome crosses is that the ends of a broken plasmid are linked, whereas the ends of a broken chromosome are unlinked. We tested whether linkage of ends influenced tract directionality using a second plasmid × chromosome cross in which DSBs were introduced into the chromosomal allele and found few bidirectional tracts. Thus, chromosome environment, but not linkage of ends, influences tract directionality. The similar tract spectra of the two plasmid × chromosome crosses suggest that similar mechanisms are involved whether recombination is initiated by DSBs in plasmid or chromosomal alleles.
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    Strategy for deletion of complete open reading frames in Saccharomyces cerevisiae (1995)
    Springer
    Current genetics 27 (1995), S. 306-308 
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    ISSN: 1432-0983
    Keywords: Gene deletion ; Open reading frame ; Saccharomyces cerevisiae ; Polymerase chain reaction
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The classical disruption method for yeast genes is by using in vitro deletion of the gene of interest, or of a part of it, with restriction enzymes. We are now routinely using a strategy that takes advantage of polymerase chain reactions (PCRs) which amplify large pieces of DNA. Since this approach results in a complete, precise deletion of the open reading frame, which is replaced by a unique restriction site, the ligated PCR can be used for the insertion of different markers of for two-step gene disruptions without an inserted marker. As we have now used this strategy for the deletion of more than ten genes we have in this report included some hints based on our experience.
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    Molecular cloning and characterization of a novel gene of Candida albicans, CDR1, conferring multiple resistance to drugs and antifungals (1995)
    Springer
    Current genetics 27 (1995), S. 320-329 
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    ISSN: 1432-0983
    Keywords: Multidrug resistance ; Candida albicans ; Saccharomyces cerevisiae ; ABC transporters
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract By functional complementation of a PDR5 null mutant of Saccharomyces cervisiae, we have cloned and sequenced the multidrug-resistance gene CDR1 of Candida albicans. Transformation by CDR1 of a PDR5-disrupted host hypersensitive to cycloheximide and chloramphenicol resulted in resistance to cycloheximide, chloramphenicol and other drugs, such as the antifungal miconazole, with collateral hypersensitivity to oligomycin, nystatin and 2,4 dinitrophenol. Our results also demonstrate the presence of several PDR5 complementing genes in C. albicans, displaying multidrug-resistance patterns different from PDR5 and CDR1. The nucleotide sequence of CDR1 revealed that, like PDR5, it encodes a putative membrane pump belonging to the ABC (ATP-binding cassette) superfamily. CDR1 encodes a 1501-residue protein of 169.9 kDa whose predicted structural organization is characterized by two homologous halves, each comprising a hydrophobic region with a set of six transmembrane stretches, preceded by a hydrophilic nucleotide binding fold.
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    New in-vivo cloning methods by homologous recombination in yeast (1994)
    Springer
    Current genetics 25 (1994), S. 180-183 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; In-vivo cloning ; Non-replicative vectors ; Homologous recombination
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have devised a new strategy to clone DNA sequences from an yeast autonomously-propagating plasmid into a non-autonomous integrative vector by in-vivo recombination. The method consists of a first step in which the replicative plasmid carrying the DNA fragment of interest forms a co-integrate with the non-replicative plasmid by an induced in-vivo reciprocal exchange accompanied by gene conversion. The dimeric plasmid obtained is then purified and cut with an appropriate restriction enzyme and ligated independently to obtain the two intact monomeric plasmids, the original autonomous plasmid plus the new non-autonomous plasmid carrying the subcloned DNA fragment. The dimeric co-integrate can also serve as substrate for a second in-vivo reciprocal exchange that produces new autonomous plasmids carrying the desired DNA fragment. The technique considerably expands the applications of in-vivo cloning in yeast by complementing three important characteristics of previously published methods: (1) it can be used to clone into non-propagating vectors; (2) co-transformation experiments are not required; and (3) the intermediate co-integrate can be used to generate new types of autonomously-propagating plasmids directly. These characteristics are independent of whether the DNA insert is flanked by appropriate restriction sites or whether it does, or does not, express a detectable phenotype in yeast. The method is particularly useful for the cloning of large DNA fragments and can be used for plasmids from organisms other than yeasts.
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    Genetic mapping of 1,3-β-glucanase-encoding genes in Saccharomyces cerevisiae (1992)
    Springer
    Current genetics 22 (1992), S. 283-288 
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    ISSN: 1432-0983
    Keywords: 1,3-β-glucanase genes ; Saccharomyces cerevisiae ; Chromosomal mapping ; Genetic mapping
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The map position of three 1,3-β-glucanase-encoding genes in S. cerevisiae has been determined following conventional meiotic and mitotic mapping combined with recombinant DNA techniques. EXG1, EXG2 and SSG1 were localized to chromosomes XII, IV and XV, respectively, by hybridizing the cloned genes to Southern blots of chromosomes sepaated by pulsed-field gel electrophoresis, in conjunction with the rad52-1-dependent chromosome-loss mapping technique. Meiotic tetrad analyses further localized the EXG1 gene 6.1 centimorgans centromere-proximal to CDC25 on the right arm of chromosome XII. EXG2 was positioned between LYS4 and GCN2 on the right arm of chromosome IV, at distances of 6.2 centimorgans from LYS4 and 4.9 centimorgans from GCN2. Finally, the SSG1 locus mapped on the right arm of chromosome XV, about 8.2 centimorgans to the centromere-proximal side of HIS3.
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    Direct induction of tetraploids or homozygous diploids in the industrial yeast Saccharomyces cerevisiae by hydrostatic pressure (1992)
    Springer
    Current genetics 22 (1992), S. 371-376 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Hydrostatic pressure ; Tetraploidy ; Homozygous diploid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Hydrostatic pressure and a dye plate method were used to investigate the direct induction of tetraploids or homozygous diploids from the industrial diploid or haploid yeast Saccharomyces cerevisiae. Above 200 MPa, hydrostatic pressure greatly inactivated the strains HF399s1 (α haploid), P-540 (a/α diploid), and P-544 (a/α diploid). At the same time, when pressure-treated cells of these strains were spread on a dye plate, some of the visible colonies were stained red/blue or dark blue (variant colonies); the rest stained violet, similar to colonies originating from diploid cells or haploid cells that were not pressure-treated. In addition, above 100 MPa, the formation of variant colonies increased with increasing pressure, and maximized (1x10-1) at 200 and 250 MPa, respectively. The size of almost all variant cells from P-544, P-540, and HF399s1 was visibly increased compared with that of untreated cells and the measured cellular DNA content of P-540 and HF399s1 was double that of untreated cells. Furthermore, based on random spore analysis and mass-matings, induced variants in the diploid strains were found to be tetraploid with an a/a/α/α genotype at the mating-type locus or, in the haploid strains, homozygous diploid with an α/α genotype. From these results we conclude that pressure treatment in combination with a dye plate is a useful method for strain improvement by direct induction of tetraploids or homozygous diploids from industrial strains whether diploid or haploids.
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    Expression of the Saccharomyces cerevisiae CYT2 gene, encoding cytochrome c1 heme lyase (1994)
    Springer
    Current genetics 25 (1994), S. 291-298 
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    ISSN: 1432-0983
    Keywords: Cytochrome c 1 ; Cytochrome c 1 heme lyase ; GRF2p ; Glucose repression ; HAPp ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In this paper we examine the expression of the Saccharomyces cerevisiae CYT2 gene, which encodes cytochrome c 1 heme lyase. This enzyme is required for covalent attachment of heme to apocytochrome c 1, a subunit of the mitochondrial respiratory chain. Transcription of the 1-kb CYT2 mRNA initiates at four prominent sites at a distance of 52–225 bp in front of the AUG start codon. The level of CYT2 mRNA is not influenced by the presence or absence of oxygen or of heme, but it is subject to carbonsource control. The concentration of the CYT2 mRNA is significantly reduced in glucose-grown cells as compared to cells grown under non-repressing conditions. Neither the HAPp activator proteins nor MIG1p, a repressor protein involved in glucose repression, seem to mediate this effect.
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    The Escherichia coli recA gene increases UV-induced mitotic gene conversion in Saccharomyces cerevisiae (1994)
    Springer
    Current genetics 25 (1994), S. 472-474 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; recA gene expression ; UV radiation ; Mitotic gene conversion
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The effect of the Escherichia coli RecA protein on mitotic recombination in the diploid D7 strain of Saccharomyces cerevisiae damaged by UV radiation was investigated. The D7 strain was transformed by two modified versions of the pNF2 plasmid: one, containing the ADH-1 promoter, and the other containing the recA gene tandemly arranged behind the ADH-1 promoter region. Immunological analysis proved the presence of the 38-kDa RecA protein in D7/pNF2ADHrecA transformants. We observed a positive effect of recA gene expression on mitotic gene conversion, mainly at higher doses of UV radiation. The results indicate that a RecA-like activity could participate in steps preceeding mitotic conversion events in yeast.
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    ERV1 is involved in the cell-division cycle and the maintenance of mitochondrial genomes in Saccharomyces cerevisiae (1994)
    Springer
    Current genetics 26 (1994), S. 15-20 
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    ISSN: 1432-0983
    Keywords: Cell-division cycle ; Mitochondrial genome ; Nuclear mutation ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In former studies it was found that the ERV1 gene is essential for cell viability and for the biogenesis of functional mitochondria. A temperature-sensitive nuclear mutant exhibits a severe reduction in all the mitochondrial transcripts. Elimination of the gene leads to growth arrest after a few cell divisions. The putative gene product bears the characteristics of a regulatory factor since it has low expression rate and a high content of charged amino acids. In this study it is further verified that the ERV1 gene alone is responsible for the observed cellular and mitochondrial defects. The 5′ region of the gene is analysed by DNA deletions and complementation studies. Expression of the gene under the control of the GAL1-10 promoter in a disruption strain of ERV1 allows a more detailed specification of its influence on mitochondrial and cellular functions. Immediate and complete loss of mitochondrial genomes is observed after the promoter has been shut off, whereas the yeast cells are still able to grow for a limited time under these conditions. Analysis of the cells by in-vivo DNA flurorescence demonstrates a specific arrest in the cell-division cycle as the terminal phenotype. To further characterize the temperature-sensitive allele of ERV1 the mutated gene has been isolated and sequenced. A single point mutation which leads to the exchange of a single amino acid is found in the reading frame.
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    The yeast nuclear gene MRP-L13 codes for a protein of the large subunit of the mitochondrial ribosome (1994)
    Springer
    Current genetics 26 (1994), S. 8-14 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Nuclear gene ; Mitochondria ; Mitochondrial ribosomal protein
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The nuclear gene MRP-L13 of Saccharomyces cerevisiae, which codes for the mitochondrial ribosomal protein YmL13, has been cloned and characterized. It is a single-copy gene residing on chromosome XI. Its nucleotide sequence was found to be identical to that of the previously reported ORF YK105. A comparison of the predicted protein sequence of the MRP-L13 gene product and the actual N-terminal amino-acid sequence of the isolated YmL13 protein indicated that the mature protein is preceded by a mitochondrial signal peptide of 86 amino-acid residues, which is the longest among all known mitochondrial ribosomal proteins of S. cerevisiae. No sequence similarity was found to any other ribosomal protein in the current databases. The transcription of MRP-L13 was found to be repressed in the presence of glucose. Its protein product is not strictly essential for mitochondrial functions, but disruption of the gene by insertion of LEU2 noticeably affected cellular growth on non-fermentable carbon sources.
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    URL: http://dx.doi.org/10.1007/BF00326298
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    yAP-1- and yAP-2-mediated, heat shock-induced transcriptional activation of the multidrug resistance ABC transporter genes inSaccharomyces cerevisiae (1996)
    Springer
    Current genetics 29 (1996), S. 103-105 
    Details
    ISSN: 1432-0983
    Keywords: Heat-shock response ; Multidrug resistance ; AP-1 homolog ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have examined whether the stress-induced transcriptional activation ofYDR1/PDR5/STS1 is mediated by yAP-1 and yAP-2. Of the stresses examined, heat shock-induced, rapid and transient PDR5 expression became very low in ayap1 yap2 double-gene disruptant, indicating that the yAP proteins mediate the response. Similar results were obtained withSNQ2, a close homologue ofPDR5. A set of 5′-truncation derivatives of thePDR5 gene identified the region from −484 to −434 as being sufficient for the response. A sequence similar to the yAP-1 recognition element recently identified in the stress-responsive yeast genes was found in this region and in the 5′-flanking sequences ofSNQ2.
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    The Saccharomyces cerevisiae HIS3 and LYS2 genes complement the Schizosaccharomyces pombe his5-303 and lys1-131 mutations, respectively: new selectable markers and new multi-purpose multicopy shuttle vectors, pSP3 and pSP4 (1995)
    Springer
    Current genetics 28 (1995), S. 380-383 
    Details
    ISSN: 1432-0983
    Keywords: Autonomously replicating sequence ; Auxotrophy ; Schizosaccharomyces pombe ; Saccharomyces cerevisiae ; Cloning vector ; Selectable marker ; HIS/his ; LYS/lys
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Three new S. pombe plasmids are described. Plasmids pSP3 and pSP4 are two Schizosaccharomyces pombe ars1 multicopy vectors with the Saccharomyces cerevisiae HIS3 or LYS2 genes as selectable markers. They complement the S. pombe his5-303 or lys1-131 mutations, respectively. Plasmid pSPars1 is a vector carrying the S. pombe ars1 and a unique NdeI site which allows the introduction of any selectable marker therefore bringing a unified vector backbone for the construction of new S. pombe/S. cerevisiae/E. coli shuttle vectors. These plasmids permit classical molecular genetic techniques to be performed directly.
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    Secretion of an enzymatically active Trichoderma harzianum endochitinase by Saccharomyces cerevisiae (1996)
    Springer
    Current genetics 29 (1996), S. 404-409 
    Details
    ISSN: 1432-0983
    Keywords: Key words Biocontrol ; Secretion ; Chitinase ; Expression cloning ; Saccharomyces cerevisiae ; Trichoderma harzianum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  A novel endochitinase agar-plate assay has been developed and used to identify 11 full-length cDNAs encoding endochitinase I (ENC I) from a Trichoderma harzianum cDNA library by expression in yeast. The 1473-bp chi1 cDNA encodes a 424-residue precursor protein including both a signal sequence and a propeptide. The deduced ENC I amino-acid sequence is homologous to other fungal and bacterial chitinases, and the enzyme cross-reacts with a polyclonal antiserum raised against chitinase A1 from Bacillus circulans. The T. harzianum endochitinase I was secreted into the culture medium by the yeast Saccharomyces cerevisiae in a functionally active form. The purified recombinant enzyme had a molecular mass of 44 kDa, an isoelectric point of 6.3, a pH optimum of 7.0 and a temperature optimum of 20 °C.
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    Starvation in yeast increases non-adaptive mutation (1999)
    Springer
    Current genetics 35 (1999), S. 77-81 
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    ISSN: 1432-0983
    Keywords: Key words Adaptive mutations ; 6-N-hydroxylaminopurine ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The frequency of reversion in a histidine-requiring mutant of Saccharomyces cerevisiae increases about ten-fold in stationary cells during histidine starvation. Histidine starvation enhances a similar frequency of reversion in a tryptophan-requiring mutant. Starvation, therefore, enhances mutation frequencies in a non-adaptive manner. The base analogue 6-N-hydroxylaminopurine (HAP) added prior to plating on medium with limited histidine strongly increases reversion of the histidine mutant. HAP-induced reversion increases further in stationary starving cells with the same kinetics as that which increases spontaneous reversion. Adding HAP to the stationary starving cells does not produce any effect.
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    Strand interruptions confer strand preference during intracellular correction of a plasmid-borne mismatch in Saccharomyces cerevisiae (1999)
    Springer
    Current genetics 35 (1999), S. 499-505 
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    ISSN: 1432-0983
    Keywords: Key words Heteroduplex repair ; Strand discrimina-tion ; Strand interruptions ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Site-directed mutagenesis was used to construct yeast centromere plasmids in which a strand nick or gap could be placed 5′ or 3′, on either strand, to a reporter gene (SUP4-o) carrying defined base mismatches. The plasmids were then transformed into yeast cells and the direction and efficiency of mismatch repair were assayed by scoring colouring of the transformant colonies. Strands that were nicked were consistently corrected more often than intact strands, but the effect was very small. However, placement of a small gap at the same positions as the nicks resulted in a marked increase in selection for the gapped strand and an enhanced efficiency of mismatch repair. Both the preference for the gapped strand and correction of the mismatch were offset by deletion of the mismatch repair gene PMS1. Together, the results suggest that strand interruptions can direct intracellular mismatch correction of plasmid-borne base mispairs in yeast.
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    Analysis of the genes activated by the FLO8 gene in Saccharomyces cerevisiae (1999)
    Springer
    Current genetics 36 (1999), S. 256-261 
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    ISSN: 1432-0983
    Keywords: Key wordsFLO8 ; Transcriptional regulation ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract It is thought that the FLO8 gene encodes a transcriptional activator of the dominant flocculation gene FLO1 in Saccharomycescerevisiae. To determine other genes which are regulated by FLO8, a detailed comparison of the transcripts from the FLO8 and Δflo8 strains was carried out. In addition to the FLO1 gene, it was found that transcription of the FLO11 and STA1 genes is positively regulated by FLO8. In flo8 strains, not only transcripts of the FLO11, STA1, and FLO1 genes but also invasive growth, extracellular glucoamylase production, and flocculation were undetected. From these results, it is suggested that FLO8 regulates these characteristics via the transcriptional regulation of the FLO11, STA1, and FLO1 genes.
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    Broad antifungal activity of β-isoxazolinonyl-alanine, a non-protein amino acid from roots of pea (Pisum sativum L.) seedlings (1991)
    Springer
    Biology and fertility of soils 11 (1991), S. 203-209 
    Details
    ISSN: 1432-0789
    Keywords: Antifungal activity ; Saccharomyces cerevisiae ; Phytopathogenic fungi ; Heterocyclic non-protein amino acid ; Pisum sativum ; Constitutive plant defence
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Geosciences , Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary β-(Isoxazolin-5-on-2-yl)-alanine (βIA), a heterocyclic non-protein amino acid from root extracts and root exudates of pea seedlings, acts as a potent growth inhibitor of several eukaryotic organisms, including yeasts, phytopathogenic fungi, unicellular green algae, and higher plants. The antibiotic effect on baker's yeast was reversed by l-methionine, l-cysteine, and l-homocysteine. Phytopathogenic fungi such as Botrytis cinerea, Pythium ultimum, and Rhizoctonia solani grown on agar containing βIA were inhibited in the growth of mycelia or in the production of sclerotia. In contrast, no significant inhibition of either Gram-positive or Gram-negative bacteria was observed. Rhizobium leguminosarum, the compatible microsymbiont of Pisum spp., and Rhizobium meliloti were able to tolerate up to 2.9 mM βIA (500 ppm) without any effect on the growth rate. Bradyrhizobium japonicum even gave a positive chemotactic response to βIA. The ecological significance of βIA as a preformed plant protectant during the seedling stage of Pisum spp. and other βIA-containing legumes is discussed.
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    Mechanism of resistance to sulphite in Saccharomyces cerevisiae (1992)
    Springer
    Current genetics 22 (1992), S. 435-440 
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    ISSN: 1432-0983
    Keywords: Sulphite-resistant mutants ; Sulphite uptake ; Acetaldehyde accumulation ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Growth inhibition and cell killing caused by sulphite were reduced in seven Saccharomyces cerevisiae sulphite-resistant independent mutants, compared to their parental strains. Genetic analysis showed that in the seven mutants resistance was inherited as a single-gene dominant mutation and that all the analyzed mutations were allelic, thus identifying a major gene responsible for sulphite resistance in S. cerevisiae. Two of the mutants, MBS20-9 and MBS30, were further characterized. 35S-sulphite uptake experiments showed that the ability to accumulate sulphite was markedly reduced in the two resistant strains. No difference between resistant and sensitive strains with respect to glyceraldehyde-3-phosphate dehydrogenase sensitivity to sulphite, or to intracellular glutathione content, were revealed. In contrast, the extracellular acetaldehyde concentration was higher in the resistant mutants, both in the presence and in the absence of sulphite.
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    Mitochondrial DNA loss by yeast reentry-mutant cells conditionally unable to proliferate from stationary phase (1992)
    Springer
    Current genetics 22 (1992), S. 471-477 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Stationary phase ; mtDNA ; Storage carbohydrate
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Double-mutant cells of the budding yeast Saccharomyces cerevisiae harboring the gcs1-1 and sed1-1 mutations are conditionally defective (cold-sensitive) only for reentry into the mitotic cycle from stationary phase. If already proliferating at the permissive temperature (29°C), these reentry-mutant cells continue to proliferate when transferred to the restrictive temperature of 14°C, but under these conditions reentry-mutant cells lose mitochondrial DNA (mtDNA). In addition, upon exhaustion of the nutrient supply at 14°C, these reentry-mutant cells entered stationary phase at a decreased cell concentration and did not accumulate the reserve carbohydrates trehalose and glycogen. Both of these deficiencies were due to the loss of mtDNA, as shown by the responses of wild-type cells also lacking mtDNA. Mitochondrial status did not affect other aspects of the reentry-mutant phenotype. Although mitochondrial activity and the accumulation of carbohydrate reserves are typical features of cells in stationary phase, the reentry-mutant phenotype reveals that neither entry into nor exit from stationary phase need involve mitochondrial function.
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    Evolutionary conservation of genomic sequences related to the GGP1 gene encoding a yeast GPI-anchored glycoprotein (1993)
    Springer
    Current genetics 23 (1993), S. 19-21 
    Details
    ISSN: 1432-0983
    Keywords: Glycosylphosphatidylinositol anchored-protein ; Southern analysis ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The GGP1 gene encodes the only GPI-anchored glycoprotein (gp115) that has been purified todate in the budding yeast Saccharomyces cerevisiae. It is a single-copy gene whose deduced amino-acid sequence shares no significant homology to any other known protein. In this paper we report a Southern hybridization analysis of genomic DNA from different eukaryotic organisms to identify homologues of the GGP1 gene. We have analyzed DNA prepared from a unicellular green alga (Chlamydomonas eugametos), from two distantly related yeast species (Candida cylindracea and Schizosaccharomyces pombe), and from the common bean Phasoleus vulgaris. The moderate stringency of the experimental conditions and the high specificity of the probes used indicate that a single-copy of GGP1-related sequences exists in all these eukaryotic organisms. The chromosomal localization of the GGP1 gene in S. cerevisiae has also been determined.
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    The STA2 and MEL1 genes of Saccharomyces cerevisiae are idiomorphic (1993)
    Springer
    Current genetics 23 (1993), S. 92-94 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Gene mapping ; Idiomorphism
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The STA2 (glucoamylase) gene of Saccharomyces cerevisiae has been mapped close to the end of the left arm of chromosome II. Meiotic analysis of a cross between a haploid strain containing STA2, and another strain carrying the melibiase gene MEL1 (which is known to be at the end of the left arm of chromosome II) produced parental ditype tetrads only. Since there is no significant DNA sequence similarity between the STA2 and MEL1 genes, or their respective flanking regions, we conclude that these two genes are carried by separate non-hybridizing sequences of chromosomal DNA, either of which can reside at the end of the left arm of chromosome II. By analogy with the mating-type locus of Neurospora crassa, we suggest that the STA2 and MEL1 genes are idiomorphs with respect to one another.
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    Molecular cloning of the yeast OPI3 gene as a high copy number suppressor of the cho2 mutation (1993)
    Springer
    Current genetics 23 (1993), S. 95-101 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Phospholipid synthesis ; Phospholipid-N-methyltransferase ; Mutant ; Over-expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract By functional complementation of the auxotrophic requirements for choline of a cdg1, cho2 double-mutant, by transformation with a genomic DNA library in a high copy number plasmid, two different types of complementing DNA inserts were identified. One type of insert was earlier shown to represent the CHO2 structural gene. In this report we describe the molecular and biochemical characterization of the second type of complementing activity. The transcript encoded by the cloned gene was about 1000-nt in length and was regulated in response to the soluble phospholipid precursors, inositol and choline. A gene disruption resulted in no obvious growth phenotype at 23°C or 30°C, but in a lack of growth at 37°C in the presence of monomethylethanolamine. Null-mutants exhibited an inositol-secretion phenotype, indicative of mutations in the lipid biosynthetic pathway. Complementation analysis, biochemical analysis of the phospholipid methylation pathway in vivo, and comparison of the restriction pattern of the cloned gene to published sequences, unequivocally identified the cloned gene as the OPI3 gene, encoding phospholipid-N-methyltransferase in yeast. When present in multiple copies the OPI3 gene efficiently suppresses the phospholipid methylation defect of a cho2 mutation. As a result of impaired synthesis of phosphatidylcholine, the INO1-deregulation phenotype is abolished in cho2 mutants transformed with the OPI3 gene on a high copy number plasmid. Taken together, these data demonstrate a significantly overlapping specificity of the OPI3 gene product for three sequential phospholipid methylation reactions in the de novo Ptd-Cho biosynthetic pathway.
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    Epitope-tagging vectors designed for yeast (1993)
    Springer
    Current genetics 23 (1993), S. 181-183 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; c-myc epitope ; Fusion
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In order to facilitate the process of epitope-tagging of yeast proteins, we have constructed two Saccharomyces cerevisiae-Escherichia coli shuttle vectors that allow fusion of a sequence encoding an epitope of the human c-myc protein at the 3′ end of any gene. An example of the use of this technique is presented.
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    Genetic and molecular analysis of REC114, an early meiotic recombination gene in yeast (1993)
    Springer
    Current genetics 23 (1993), S. 295-304 
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    ISSN: 1432-0983
    Keywords: Meiosis ; Meiotic recombination ; Saccharomyces cerevisiae ; REC114
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Four new meiotic recombination genes were previously isolated by selecting for mutations that rescue the meiotic lethality of rad52 spo13 strains. One of these genes, REC114, is described here, and the data confirm that REC114 is a meiosis-specific recombination gene with no detectable function in mitosis. REC114 is located on chromosome XIII approximately 4,9 cM from CIN4. The nucleotide sequence reveals an open reading frame of 1262 bp, consensus intron splice sites close to the 3′ end, and indicates that the second exon codes for only seven amino acids. In the promoter region, a URS1 consensus sequence (TGGGCGGCTA), identical to the URS1 found in the promoter of SPO16, is present 93 bp upstream of the translation start site. Northern-blot hybridization demonstrates that REC114 is transcribed only during meiosis and that it is not expressed in the absence of the IME1 gene product, even when IME2 is constitutively expressed.
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    Lack of correlation between trehalase activation and trehalose-6 phosphate synthase deactivation in cAMP-altered mutants of Saccharomyces cerevisiae (1993)
    Springer
    Current genetics 23 (1993), S. 382-387 
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    ISSN: 1432-0983
    Keywords: Trehalase ; Trehalose-6-P synthase ; cAMP mutants ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The rise in cAMP level that follows the addition of glucose or 2,4-dinitrophenol (DNP) to stationaryphase cells of Saccharomyces cerevisiae was accompanied by a marked activation of trehalase (3-fold increase) and a concomitant deactivation of trehalose-6 phosphate synthase (50% of the basal levels). In glucose-grown exponential cells, which are deficient in glucose-induced cAMP signalling, the addition of glucose also prompted a decrease in trehalose-6 phosphate synthase, but had no effect on trehalase activity. Mutants defective in the RAS-adenylate cyclase pathway (ras1 ras2 bcy1 strain), as well as mutants containing greatly reduced protein kinase activity either cAMP-dependent (tpk w1 BCY1 strains) or cAMP-independent (tpk1 w1 bcy1 strains), were unable to show glucose- or DNP-induced trehalase activation but still displayed a clear decrease in trehalose-6 phosphate synthase activity upon addition of these compounds. These data suggest that the activity of trehalose-6 phosphate synthase, as opposed to that of trehalase, is not controlled by the cAMP signalling pathway “in vivo”. Trehalose-6 phosphate synthase was competitively inhibited by glucose (Ki=15 mM) and resulted unaffected by ATP in assays performed “in vitro”.
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    Structure and regulation of the isocitrate lyase gene ICL1 from the yeast Saccharomyces cerevisiae (1993)
    Springer
    Current genetics 23 (1993), S. 375-381 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Isocitrate lyase ; Gene regulation ; Ethanol induction
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The ICL1 gene encoding the isocitrate lyase from Saccharomyces cerevisiae was cloned and sequenced. A reading frame of 557 amino acids showing significant similarity to isocitrate lyases from seven other species could be identified. Construction of icl1 null mutants led to growth defects on C2 carbon sources while utilization of sugars or C3 substrates remained unaffected. Using an ICL1-lacZ fusion integrated at the ICL1 locus, a more than 200-fold induction of β-galactosidase activity was observed after growth on ethanol when compared with glucose-repressed conditions. A preliminary analysis of the ICL1 upstream region identified a 364-bp fragment necessary and sufficient for this regulatory phenotype. Sequence motifs also present in the upstream regions of co-regulated genes were found within this region.
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    Phenotypic identification of amplifications of the ADH4 and CUP1 genes of Saccharomyces cerevisiae (1993)
    Springer
    Current genetics 23 (1993), S. 392-396 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Gene amplification ; ADH4 ; CUP1
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Primary gene amplification, i.e., mutation from one gene copy to multiple gene copies per genome, is important in genomic evolution, as a means of producing anti-cancer drug resistance, and is associated with the progression of tumor malignancy. Primary amplification has not been studied in normal eukaryotic cells because amplifications are extremely rare in these cells. A system has been developed to phenotypically identify co-amplifications of the ADH4 and CUP1 genes of Saccharomyces cerevisiae and 21 independent spontaneous amplifications have been isolated.
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    Donation of information to the unbroken chromosome in double-strand break repair (1993)
    Springer
    Current genetics 23 (1993), S. 414-422 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Donation ; Gene conversion ; Double-strand break repair ; Heteroduplex DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have used transformation of yeast with lincarized plasmids to study the transfer of information to the unbroken chromosome during double-strand break repair. Using a strain which carried the wild-type HIS3 allele, and a linearized plasmid which carried a mutant his3 allele, we have obtained His- transformants. In these, double-strand break repair has resulted in precise transfer of genetic information from the plasmid to the chromosome. Such repair events, we suggest, are gene conversions which entail the formation of heteroduplex DNA on the (unbroken) chromosome. If this suggestion is correct, our results reflect the spatial distribution of such heteroduplex DNA. Transfer of information from the plasmid to the chromosome was obtained at a maximal frequency of 1.5% of the repair events, and showed a dependence with distance. Transformation to His- was also obtained with a 2-kbp insertion and with a deletion of 200 bp. The latter results suggest that gene conversion of large heterologies can occur via repair of a heteroduplex DNA intermediate.
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    MCB elements and the regulation of DNA replication genes in yeast (1993)
    Springer
    Current genetics 24 (1993), S. 185-192 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Cell cycle ; Transcription ; DNA replication
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In eukaryotic organisms, genes involved in DNA replication are often subject to some form of cell cycle control. In the yeast Saccharomyces cerevisiae, most of the DNA replication genes that have been characterized to date are regulated at the transcriptional level during G1 to S phase transition. A cis-acting element termed the MluI cell cycle box (or MCB) conveys this pattern of regulation and is common among more than 20 genes involved in DNA synthesis and repair. Recent findings indicate that the MCB element is well conserved among fungi and may play a role in controlling entry into the cell division cycle. It is evident from studies in higher systems, however, that transcriptional regulation is not the only form of control that governs the cell-cycle-dependent expression of DNA replication genes. Moreover, it is unclear why this general pattern of regulation exists for so many of these genes in various eukaryotic systems. This review summarizes recent studies of the MCB element in yeast and briefly discusses the purpose of regulating DNA replication genes in the eukaryotic cell cycle.
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    Polymorphism within the nuclear and 2μm genomes of Saccharomyces cerevisiae (1991)
    Springer
    Current genetics 20 (1991), S. 189-194 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Bakers' and lager yeast ; Chromosomal and 2 μm DNA polymorphism
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Seven strains of bakers' yeast were obtained as a representative sample of the Spanish baking industry. The nuclear genome was monitored for polymorphism by transverse alternating field electrophoresis (TAFE) and restriction maps of 2 μm DNA were produced. All seven strains were uniquely different when evaluated by their total chromosomal lengths whereas only two 2 μm variants were defined. There was no apparent correlation between chromosomal and plasmid polymorphism. The extensive chromosomal polymorphism within one 2 μm DNA type indicates the rapid and relatively recent evolution of the nuclear genome. The hybrid origin (S. cerevisiae-S.monacensis) of lager yeast was critically evaluated by TAFE analysis of S. cerevisiae and S. carlsbergensis chromosomes. The absence of corresponding S. cerevisiae chromosomes III and XIII in S. carlsbergensis argued against the hybrid origin of lager strains. We discuss limitations of the hybrid origin hypothesis of industrial yeasts and propose that the molecular coevolution observed in 2 μm DNA serves as a useful additional mechanism for rationalization of some of the structural polymorphism of the nuclear genome.
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    URL: http://dx.doi.org/10.1007/BF00326231
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    A novel method for in situ screening of yeast colonies with the β-glucuronidase reporter gene (1991)
    Springer
    Current genetics 20 (1991), S. 437-439 
    Details
    ISSN: 1432-0983
    Keywords: Schizosaccharomyces pombe ; Saccharomyces cerevisiae ; β-glucuronidase ; Colony colour assay ; Fluorometric assay
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Expression of the β-galactosidase gene in yeast has served as a screening marker for many purposes. Here it is shown that in two yeasts, Saccharomyces cerevisiae and Schizosaccharomyces pombe, the β-glucuronidase (GUS) gene can be used as an alternative marker. Since the histochemical substrate can not be taken up by yeast cells, direct colony screening of plates was found to be impossible. However, by a replica plating technique, GUS expression became visibly detectable within 10 min when the GUS gene was strongly expressed. The staining method could still be performed for expression at a 100-fold lower level, but incubation times of several hours were needed. Furthermore, specific GUS expression levels of yeast protein extracts could be quantified by a fluorometric assay which is both very simple to perform and highly sensitive. Since the GUS gene can also tolerate large N-terminal fusions, this method should be particularly attractive for studying such diverse problems as transcriptional and translational regulation or subcellular localization in yeast.
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    URL: http://dx.doi.org/10.1007/BF00317075
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    Pentose-phosphate pathway in Saccharomyces cerevisiae: analysis of deletion mutants for transketolase, transaldolase, and glucose 6-phosphate dehydrogenase (1993)
    Springer
    Current genetics 24 (1993), S. 373-376 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Pentose-phosphate pathway ; Transketolase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Deletion mutants for the yeast transketolase gene TKL1 were constructed by gene replacement. Transketolase activity was below the level of detection in mutant crude extracts. Transketolase protein could be detected as a single protein band of the expected size by Western-blot analysis in wild-type strains but not in the delection mutant. Deletion of TKL1 led to a reduced but distinct growth in synthetic medium without an aromatic amino-acid supplement. We also isolated double and triple mutants for transketolase (tkl1), transaldolase (tal1), and glucose 6-phosphate dehydrogenase (zwf1) by crossing the different mutants. A tal1 tkl1 double mutant grew nearly like wild-type in rich medium. Only the tkl1 zwf1 double and the tal1 tkl1 zwf1 triple mutant grew more slowly than the wild-type in rich medium. This growth defect could be partly alleviated by the addition of xylulose but not ribose. The triple mutant still grew slowly on a synthetic mineral salts medium without a supplement of aromatic amino acids. This suggests the existence of an alternative but limited source of pentose phosphates and erythrose 4-phosphate in the tkl1 zwf1 double mutants. Hybridization with low stringency showed the existence of a sequence with homology to transketolase, possibly a second gene.
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    URL: http://dx.doi.org/10.1007/BF00351843
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    Inactivation of the RAD1 excision-repair gene does not affect correction of mismatches on heteroduplex plasmid DNA in yeast (1992)
    Springer
    Current genetics 21 (1992), S. 261-263 
    Details
    ISSN: 1432-0983
    Keywords: Mismatch correction ; Saccharomyces cerevisiae ; Excision repair ; DNA methylation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The efficiency and direction of mismatch correction in the Saccharomyces cerevisiae SUP4-o gene were not altered by an excision-repair defect (rad1). Although excision-repair functions remove methylated adenine from yeast, adenine methylation at a GATC sequence in SUP4-o did not direct the correction of mismatches via excision repair.
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    URL: http://dx.doi.org/10.1007/BF00336851
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    Calmodulin-dependent protein kinase II and calmodulin are required for induced thermotolerance in Saccharomyces cerevisiae (1995)
    Springer
    Current genetics 27 (1995), S. 190-193 
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    ISSN: 1432-0983
    Keywords: Calmodulin ; Calmodulin-dependent protein kinase II ; Heat shock response ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We show here that yeast mutants lacking calmodulin-dependent protein kinase II fail to fully acquire induced thermotolerance. A similar result was also obtained with mutants depending solely on either the N-terminal half or the C-terminal half of calmodulin. These findings indicate that both calmodulin-dependent protein kinase II and calmodulin are required for induced thermotolerance.
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    URL: http://dx.doi.org/10.1007/BF00313434
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    A genetic analysis of an alpha-amylase super-secretor in yeast; implications for the regulatory pathway (1991)
    Springer
    Current genetics 20 (1991), S. 181-184 
    Details
    ISSN: 1432-0983
    Keywords: Alpha amylase ; Secretion ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Extracellular glucoamylase activity was increased by a gene, which is present in super-secretor, but absent in low-secretor, strains of the yeast Saccharomyces cerevisiae. Genetic data indicated that this super-secretor gene is linked to the STA3 structural gene for glucoamylase. This gene appears to act specifically since it increased the secretion of glucoamylase but not of other secreted enzymes like acid phosphatase and invertase.
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    URL: http://dx.doi.org/10.1007/BF00326229
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    Polymeric genes MEL8, MEL9 and MEL10 — new members of α-galactosidase gene family in Saccharomyces cerevisiae (1991)
    Springer
    Current genetics 20 (1991), S. 269-276 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Melibiose fermentation ; MEL ; Polymeric genes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We used a combination of genetic hybridization analysis and electrokaryotyping with radioactively labelled MEL1 gene probe hybridization to isolate and identify seven polymeric genes for the fermentation of melibiose in strain CBS 5378 of Saccharomyces cerevisiae (syn. norbensis). Four of the MEL genes, i.e. MEL3, MEL4, MEL6 and MEL7, were allelic to those found in S. cerevisiae strain CBS 4411 (syn. S. oleaginosus) whereas three genes, i.e. MEL8, MEL9 and MEL10 occupied new loci. Electrokaryotyping showed that all seven MEL genes in CBS 5378 were located on different chromosomes. The new MEL8, MEL9 and MEL10 genes were found on chromosomes XV, X/XIV and XII, respectively.
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    URL: http://dx.doi.org/10.1007/BF00318514
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    Isolation and primary structure of the ERG9 gene of Saccharomyces cerevisiae encoding squalene synthetase (1991)
    Springer
    Current genetics 20 (1991), S. 365-372 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Ergosterol ; Squalene synthetase ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    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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    URL: http://dx.doi.org/10.1007/BF00317063
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    A mutated ARO4 gene for feedback-resistant DAHP synthase which causes both o-fluoro-dl-phenylalamine resistance and β-phenethyl-alcohol overproduction in Saccharomyces cerevisiae (1991)
    Springer
    Current genetics 20 (1991), S. 453-456 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; β-phenethyl-alcohol ; ARO4 gene ; DAHP synthase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary o-Fluoro-dl-phenylalanine (OFP)-resistant mutants which overproduce β-phenethyl-alcohol were isolated from a laboratory strain of Saccharomyces cerevisiae. Cells of one of the mutants accumulated tyrosine and phenylalanine 1.5–3 fold more than did wild-type cells. Its 3-deoxy-d-arabino-hepturosonate-7-phosphate (DAHP) synthase (EC 4.1.2.15), encoded by ARO4, was free from feedback inhibition by tyrosine. Genetic analysis revealed that the mutation was controlled by a single dominant gene, ARO4-OFP, encoding feedback-resistant DAHP synthase by tyrosine, and that this gene caused both the OFP resistance and β-phenethyl-alcohol overproduction. This was supported by molecular genetic studies using cloned ARO4 both from the wild-type and its mutant strain.
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    Antisense RNA regulation of the ILV2 gene in yeast: a correction (1994)
    Springer
    Current genetics 25 (1994), S. 289-289 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Inducible antisense gene ; Acetolactate synthase ; Bradytrophic phenocopy
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A previous report of the use of antisense RNA to regulate gene expression in yeast is incorrect.
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    URL: http://dx.doi.org/10.1007/BF00357175
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    Isolation and characterization of three mutants with increased sensitivity to photoactivated 3-carbethoxypsoralen in Saccharomyces cerevisiae (1994)
    Springer
    Current genetics 25 (1994), S. 407-411 
    Details
    ISSN: 1432-0983
    Keywords: Psoralen sensitivity ; Saccharomyces cerevisiae ; DNA repair ; Oxidative stress
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The complementation and genetical analysis of yeast mutants sensitive to photoactivated 3-carbethoxy-psoralen define three novel recessive mutant alleles pso-5-1, pso6-1, and pso7-1. Their cross-sensitivity to UV254nm, radiomimetic mutagens, and to chemicals enhancing oxidative stress suggest that these mutants are either impaired in metabolic steps protecting from oxidative stress or in mechanisms of the repair of oxygen-dependent DNA lesions. None of the three novel mutant alleles block the induction of reverse mutation by photoactivated mono- and bi-functional psoralens, nitrogen mustards, or UV254nm.
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    Suppression of a mitochondrial point mutation in a tRNA gene can cast light on the mechanisms of 3′ end-processing (1994)
    Springer
    Current genetics 25 (1994), S. 451-455 
    Details
    ISSN: 1432-0983
    Keywords: tRNA processing ; Saccharomyces cerevisiae ; Mitochondria
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We used a genetic approach to study the nuclear factors involved in the biogenesis of mitochondrial tRNAs. A point mutation in the mitochondrial tRNAAsp gene of Saccharomyces cerevisiae had previously been shown to result in a temperature-sensitive respiratory-deficient phenotype as a result of the absence of 3′ end-processing of the tRNAAsp. Analysis of mitochondrial revertants has shown that all revertants sequenced have a G-A compensatory change at position 53, which restores the hydrogen-bond with the mutated nucleotide. We then searched for nuclear suppressors to identify the nuclear gene(s) involved in mitochondrial tRNA 3′ end-processing. One such suppressor mutation was further characterized: it restores tRNAAsp maturation and growth at 36°C on glycerol medium in heterozygous diploids, but leads to a defective growth phenotype in haploids.
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    Control of glycolytic gene expression in the budding yeast (Saccharomyces cerevisiae) (1995)
    Springer
    Current genetics 29 (1995), S. 1-9 
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    ISSN: 1432-0983
    Keywords: Glycolysis ; Transcriptional activation ; Saccharomyces cerevisiae ; Chromatin structure ; Glucose induction
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
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    URL: http://dx.doi.org/10.1007/BF00313187
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    A mutation in cytochrome oxidase subunit 2 restores respiration of the mutant pet ts1402 (1997)
    Springer
    Current genetics 31 (1997), S. 401-407 
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    ISSN: 1432-0983
    Keywords: Key words Cytochrome oxidase ; Mitochondrial localization ; PET1402/OXA1 ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The yeast PET1402/OXA1 gene encoding a 44.8-kDa protein is required for mitochondrial biogenesis. Substitution of Leu240 to serine in the protein results in an accumulation of the precursor form of the mitochondrially encoded subunit 2 of cytochrome oxidase (Cox2) and temperature-sensitive respiration. This temperature sensitivity can be suppressed by a mutation in the cox2 gene changing Ala189 of the Cox2 protein to proline. In the cox2-ts1402 double mutant respiration is restored without removal of the Cox2 pre-sequence. The suppression suggests an interaction of the Pet1402 protein with the cytochrome oxidase complex. Antibodies raised against the predicted C-terminus and the tagged N-terminus of the Pet1402 protein reacted with a 37-kDa polypeptide. This protein, present in the mitochondrial fraction, is localized within the inner membrane. The difference in size can be explained by the removal of the predicted mitochondrial-targeting sequence from the Pet1402 protein. The mitochondrial localization of the protein points to a direct interaction with the cytochrome oxidase complex.
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    URL: http://dx.doi.org/10.1007/s002940050222
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    A mutant allele of the SUP45 (SAL4) gene of Saccharomyces cerevisiae shows temperature-dependent allosuppressor and omnipotent suppressor phenotypes (1995)
    Springer
    Current genetics 27 (1995), S. 417-426 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Omnipotent suppression ; Nonsense suppression ; SUP45
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Using a plasmid-based termination-read-through assay, the sal4-2 conditional-lethal (temperature-sensitive) allele of the SUP45 (SAL4) gene was shown to enhance the efficiency of the weak ochre suppressor tRNA SUQ5 some 10-fold at 30°C. Additionally, this allele increased the suppressor efficiency of SRM2-2, a weak tRNAGln ochre suppressor, indicating that the allosuppressor phenotype is not SUQ5-specific. A sup + sal4-2 strain also showed a temperature-dependent omnipotent suppressor phenotype, enhancing readthrough of all three termination codons. Combining the sal4-2 allele with an efficient tRNA nonsense suppressor (SUP4) increased the temperature-sensitivity of that strain, indicating that enhanced nonsense suppressor levels contribute to the conditional-lethality conferred by the sal4-2 allele. However, UGA suppression levels in a sup + sal4-2 strain following a shift to the non-permissive temperature reached a maximum significantly below that exhibited by a non-temperature sensitive SUP4 suppressor strain. Enhanced nonsense suppression may not therefore be the primary cause of the conditional-lethality of this allele. These data indicate a role for Sup45p in translation termination, and possibly in an additional, as yet unidentified, cellular process.
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    URL: http://dx.doi.org/10.1007/BF00311210
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    Mutants of Saccharomyces cerevisiae sensitive to oxidative and osmotic stress (1995)
    Springer
    Current genetics 27 (1995), S. 427-434 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Oxidative stress ; Osmotic stress
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Although oxidative stress is involved in many human diseases, little is known of its molecular basis in eukaryotes. In a genetic approach, S. cerevisiae was used to identify elements involved in oxidative stress. By using hydrogen peroxide as an agent for oxidative stress, 34 mutants were identified. All mutants were recessive and fell into 16 complementation groups (pos1 to pos16 for peroxide sensitivity). They corresponded to single mutations as shown by a 2:2 segregation pattern. Enzymes reportedly involved in oxidative stress, such as glucose-6-phosphate dehydrogenase, glutathione reductase, superoxide dismutase, as well as glutathione concentrations, were investigated in wild-type and mutant-cells. One complementation group lacked glucose-6-phosphate dehydrogenase and was shown to be allelic to the glucose-6-phosphate dehydrogenase structural gene ZWF1/MET19. In other mutants all enzymes supposedly involved in oxidative-stress resistance were still present. However, several mutants showed strongly elevated levels of glutathione reductase, gluconate-6-phosphate dehydrogenase and glucose-6-phosphate dehydrogenase. One complementation group, pos9, was highly sensitive to oxidative stress and revealed the same growth phenotype as the previously described yap1/par1 mutant coding for the yeast homologue of mammalian transcriptional activator protein, c-Jun, of the proto-oncogenic AP-1 complex. However, unlike par1 mutants, which showed diminished activities of oxidative-stress enzymes and glutathion level, the pos9 mutants did not reveal any such changes. In contrast to other recombinants between pos mutations and par1, the sensitivity did not further increase in par1 pos9 recombinants, which may indicate that both mutations belong to the same regulating circuit. Interestingly, ten complementation groups were, in parallel, sensitive to osmotic stress, and one mutant allele revealed increased heat sensitivity. Our results indicate that a surprisingly large number of genes seem to be involved in oxidative-stress resistance and a possible overlap exists between osmotic stress and other stress reactions.
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    URL: http://dx.doi.org/10.1007/BF00311211
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    The Saccharomyces cerevisiae gene PSO5/RAD16 is involved in the regulation of DNA damage-inducible genes RNR2 and RNR3 (1998)
    Springer
    Current genetics 34 (1998), S. 124-127 
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    ISSN: 1432-0983
    Keywords: Key wordsPSO5/RAD16 ; Saccharomyces cerevisiae ; Nucleotide excision repair ; Oxidative stress ; Ribonucleotide reductase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The expression of β-galactosidase from DNA damage-inducible RNR2-lacZ and RNR3-lacZ fusion constructs was compared in wild-type (WT) and pso5/rad16 mutant strains after treatment with five mutagens/oxidative stressors. While exposure to the mutagens UVC, 4NQO and H2O2 induced expression of the RNR2-lacZ and RNR3-lacZ fusion constructs in two WT strains, treatment with the two oxidative stressors tBOOH and paraquat did not. In the pso5-1 mutant induction of RNR2-lacZ was largely reduced after UVC and H2O2 while there was no significant induction of β-galactosidase expression after 4NQO treatment for this construct. For RNR3-lacZ there was strongly reduced expression of pso5-1 after UVC and 4NQO while H2O2 failed to induce expression of β-galactosidase. In the WT strains the ranking of the inducing power of the mutagens at 90% survival (as measured in the pso5-1 mutant) was 4NQO〉UVC〉H2O2. Though the WT strains were clearly more resistant that the pso5-1 mutant to the two oxidative stressors paraquat and tBOOH, these substances failed to significantly enhance expression of the RNR2-lacZ and RNR3-lacZ fusion constructs in both the WT and the pso5-1 mutant. Our data suggest that Pso5p/Rad16p has a function in the signal transducing pathway controlling DNA damage-inducible components of nucleotide excision repair.
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    Azf1p is a nuclear-localized zinc-finger protein that is preferentially expressed under non-fermentative growth conditions in Saccharomyces cerevisiae (1998)
    Springer
    Current genetics 34 (1998), S. 287-296 
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    ISSN: 1432-0983
    Keywords: Key words Zinc-finger protein ; Nuclear localization ; Immuno electron microscopy ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In previous studies the AZF1 gene has been identified as a second high-copy number suppressor for a special mutant of the gene for the mitochondrial core enzyme of RNA polymerase. The first high-copy number suppressor of this mutant turned out to be the specificity factor MTF1 for mitochondrial transcription. Up to now, the influence of AZF1 on mitochondrial transcription, its precise localization in the cell and the regulation of its expression has not been determined. The putative protein contains a long stretch of poly-asparagine amino acids and a typical zinc-finger domain for DNA binding. These characteristic structural features were used to create the abbreviation AZF1 (Asparagine-rich Zinc Finger protein). An initial computer analysis of the sequence gave no conclusive results for the presence of a mitochondrial import sequence or a typical nuclear-targeting sequence. A recent more-detailed analysis identified a possible nuclear localization signal in the middle of the protein. Disruption of the gene shows no effect on plates with glucose-rich medium or glycerol. In this report a specific polyclonal antibody against Azf1p was prepared and used in cell-fractionation experiments and in electron-microscopic studies. Both of these clearly demonstrate that the AZF1 protein is localized exclusively in the nucleus of the yeast cell. Northern analysis for the expression of the AZF1 messenger RNA under different growth conditions was therefore performed to obtain new insights into the regulation of this gene. Together with the respective protein-expression analysis these data demonstrate that Azf1p is preferentially synthezised in higher amounts under non-fermentable growth conditions. Over-expression of Azf1p in the yeast cell does not influence the expression level of the mitochondrial transcription factor Mtf1p, indicating that the influence of Azf1p on the suppression of the special mitochondrial RNA polymerase mutant is an indirect one. Subcellular investigation of the deletion mutant by electron microscopy identifies specific ultrastructural cell-division defects in comparison to the wild-type.
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    URL: http://dx.doi.org/10.1007/s002940050398
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    Mutant allele pso7-1, that sensitizes Saccharomyces cerevisiae to photoactivated psoralen, is allelic with COX11, encoding a protein indispensable for a functional cytochrome c oxidase (1999)
    Springer
    Current genetics 36 (1999), S. 124-129 
    Details
    ISSN: 1432-0983
    Keywords: Key words Psoralen sensitivity ; Cytochrome oxidase ; Saccharomyces cerevisiae ; Oxidative stress
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The yeast gene PSO7 was cloned from a genomic library by complementation of the pso7-1 mutant's sensitivity phenotype to 4-nitroquinoline-1-oxide (4NQO). Sequence analysis revealed that PSO7 is allelic to the 1.1-kb ORF of the yeast gene COX11 which is located on chromosome XVI and encodes a protein of 28-kDa localized in the inner mitochondrial membrane. Allelism of PSO7/COX11 was verified by non-complementation of 4NQO-sensitivity in diploids homo- and hetero-allelic for the pso7-1 and cox11::TRP1 mutant alleles. Sensitivity to 4NQO was the same in exponentially growing cells of the pso7-1 mutant and the cox11::TRP1 disruptant. Allelism of COX11 and PSO7 indicates that the pso7 mutant's sensitivity to photoactivated 3-carbethoxypsoralen and to 4NQO is not caused by defective DNA repair, but rather is due to an altered metabolism of the pro-mutagen 4NQO in the absence of cytochrome oxidase (Cox) in pso7-1/cox11::TRP1 mutants/disruptants. Lack of Cox might also lead to a higher reactivity of the active oxygen species produced by photoactivated 3-carbethoxypsoralen. The metabolic state of the cells is important for their sensitivity phenotype since the largest enhancement of sensitivity to 4NQO between wild-type (WT) and the pso7 mutant occurs in exponentially growing cells, while cells in stationary phase or growing cells in phosphate buffer have the same 4NQO resistance, irrespective of their WT/mutant status. Strains containing the pso7-1 or cox11::TRP1 mutant allele were also sensitive to the oxidative stress-generating agents H2O2 and paraquat. Mutant pso7-1, as well as disruptant cox11::TRP1, harboured mitochondria that in comparison to WT contained less than 5% and no detectable Cox activity, respectively.
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    Mitotic recombination and localized DNA double-strand breaks are induced after 8-methoxypsoralen and UVA irradiation in Saccharomyces cerevisiae (1998)
    Springer
    Current genetics 34 (1998), S. 30-42 
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    ISSN: 1432-0983
    Keywords: Key words Mitotic recombination ; DNA double-strand breaks ; Saccharomyces cerevisiae ; 8-Methoxypsoralen plus UVA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Mitotic recombination within the ARG4 gene of Saccharomyces cerevisiae was analysed after treatment of cells with the recombinogenic agent 8-methoxypsoralen (8-MOP) plus UVA. The appearance of DNA double-strand breaks (DSBs) in the ARG4 region during post-treatment incubation was also tested. The results obtained after 8-MOP plus UVA treatment indicate that in mitotic cells: (1) recombination at the ARG4 locus is increased 30 – 500 fold per survivor depending on the strains and the doses employed, (2) the increase of recombination results essentially from gene conversion events which involve the RV site located in the 5′ region of the ARG4 gene twice as often as the Bgl site at the 3′ end, (3) depending on 8-MOP/UVA dose, ectopic gene conversion is associated with reciprocal translocation, (4) DSBs occur preferentially in the ARG 5′ region during post-treatment incubation, as well as in other intergenic regions containing both promoters or/and terminators of transcription, and (5) changes in sequence content in the 5′ region of ARG4, which influences positions and frequencies of DSBs formed during repair, are correlated with a modification of the local chromatin structure.
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    URL: http://dx.doi.org/10.1007/s002940050363
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    Stabilization of methionine-rich protein in Saccharomyces cerevisiae: targeting of BZN protein into the peroxisome (1994)
    Springer
    Current genetics 26 (1994), S. 390-397 
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    ISSN: 1432-0983
    Keywords: Overexpression ; Peroxisomes ; Saccharomyces cerevisiae ; Stabilization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have constructed a gene coding for the 12-kDa intermediate form of the 2s methionine-rich protein from Bertholletia excelsa seeds. This protein, expressed intracellularly in yeast, is characterised by a 20-min balf-life. By adding 11 amino acids corresponding to the peroxisome-targeting sequence (PTSc) of luciferase, we have significantly increased its half-life. This stabilization allowed accumulation of the BZN protein into the peroxisome as judged by cell fractionation. Accumulation of the 12-kDa protein results in a significant increase of the total methionine content in yeast cells (30%) indicating that such a microorganism could represent a practicable protected shuttl for an animal-feed additive.
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    URL: http://dx.doi.org/10.1007/BF00309924
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  • 182
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    Repair of ultraviolet light damage in Saccharomyces cerevisiae as studied with double- and single-stranded incoming DNAs (1992)
    Springer
    Current genetics 21 (1992), S. 93-94 
    Details
    ISSN: 1432-0983
    Keywords: DNA repair ; Incoming DNA ; Saccharomyces cerevisiae ; Ultraviolet light
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Purified double- and single-stranded DNAs of the autonomously replicating vector M13RK9-T were irradiated with ultraviolet light (UV) in vitro and introduced into competent whole cells of Saccharomyces cerevisiae. Incoming double-stranded DNA was more sensitive to UV in excision repair-deficient rad2-1 cells than in proficient repair RAD + cells, while single-stranded DNA exhibited high sensitivity in both host cells. The results indicate that in yeast there is no effective rescue of UV-incoming single-stranded DNA by excision repair or other constitutive dark repair processes.
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    URL: http://dx.doi.org/10.1007/BF00318465
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  • 183
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    The identification of 18 nuclear genes required for the expression of the yeast mitochondrial gene encoding cytochrome c oxidase subunit 1 (1992)
    Springer
    Current genetics 21 (1992), S. 139-146 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Mitochondria ; Cytochrome c oxidase subunit 1 ; RNA processing
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Eighteen nuclear mutants of the yeast Saccharomyces cerevisiae, each disturbed in the biosynthesis of the mitochondrially encoded cytochrome c oxidase subunit 1 (cox 1) and each representing a distinct complementation group, have been examined to identify the level at which COX1 expression is affected. RNA blotting revealed that most have a defect in the processing of COX1 precursor-mRNA; only a few are defective in COX1 transcription and/or pre-mRNA stability. In most RNA-processing mutants, the absence of the COX1 messenger results from a defect in the splicing of one or more COX1 introns. In turn, this defect can be ascribed to a mutation in a nuclear gene which is either directly involved in splicing or else acts indirectly by impairing COX1 translation.
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    URL: http://dx.doi.org/10.1007/BF00318473
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  • 184
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    Sequence analysis of three deficient mutants of cytochrome oxidase subunit I of Saccharomyces cerevisiae and their revertants (1994)
    Springer
    Current genetics 26 (1994), S. 546-552 
    Details
    ISSN: 1432-0983
    Keywords: Cytochrome oxidase ; Revertant ; Mitochondria ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Three respiratory-deficient mutants of cytochrome oxidase subunit I in the yeast mitochondrion have been sequenced. They are located in, or near, transmembrane segment VI, the catalytic core of the enzyme. Respiratory-competent revertants have been selected and studied. The mutant V244M was found to revert at the same site in valine (wild-type), isoleucine or threonine. The revertants of the mutant G251R were of three types: glycine (wild-type), serine and threonine at position 251. A search for second-site mutations was carried out but none were found. Among 60 revertants tested, the mutant K265M was found to revert only to the wild-type allele.
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    URL: http://dx.doi.org/10.1007/BF00309948
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  • 185
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    Cloning and mapping of the CYS4 gene of Saccharomyces cerevisiae (1992)
    Springer
    Current genetics 21 (1992), S. 285-289 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Cysteine biosynthetic ; CYS4 ; Mapping
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A DNA fragment containing the CYS4 gene of Saccharomyces cerevisiae was isolated from a genomic library. The cloned fragment hybridized to the transverse-alternating-field-electrophoresis band corresponding to chromosomes VII and XV. According to the 2 μm DNA chromosome-loss procedure, the cys2 and cys4 mutations, which are linked together and co-operatively confer cysteine dependence, were assigned to chromosome VII. By further mapping involving tetrad analysis, the cys2-cys4 pair was localized between SUP77 (SUP166) and ade3 on the right arm of chromosome VII.
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    URL: http://dx.doi.org/10.1007/BF00351684
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    Genetic analysis of serine biosynthesis and glucose repression in yeast (1992)
    Springer
    Current genetics 21 (1992), S. 295-300 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Serine biosynthesis ; Mutant isolation ; Glucose repression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Serine and glycine biosynthesis in yeast proceed by two pathways; a “glycolytic” pathway, using 3-phosphoglycerate, and a “gluconeogenic” pathway, using glyoxylate. We used a mutation in the cat1 gene to abolish the glucose-repressible “gluconeogenic” pathway and re-isolated two mutants, ser1 and ser2, in the “glycolytic” pathway. The ser1 mutation corresponded to phosphoserine transaminase and ser2 to that of phosphoserine phosphatase. Mutagenesis of a ser1 ser2 cat1 triple mutant facilitated the isolation of a mutation in a new gene, SER10. SER10 appears to be part of a pathway which, under normal growth conditions, is less important in serine biosynthesis. The ser1 ser2 ser10 triple mutants were totally serine auxotrophic on glucose media but serine prototrophic during growth on non-fermentable carbon sources. This phenotype was used to select for possible regulatory mutants that synthesize serine by the gluconeogenic pathway even in the presence of glucose, e.g., with a non-glucose repressible glyoxylate cycle. In an alternative approach to isolate such mutants URA3 and TRP1 expression were placed under the control of the glucose-repressible FBP1 (fructose-1,6-bisphosphatase) promoter. Although both systems resulted in strong selection pressure we could not isolate constitutively derepressed mutants. These results indicate that transcription of glucose-repressible gluconeogenic enzymes is mainly dependent on positive regulatory elements.
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    URL: http://dx.doi.org/10.1007/BF00351686
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  • 187
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    Complementation of the Saccharomyces cerevisiae srb1-1 mutation: an autoselection system for stable plasmid maintenance (1992)
    Springer
    Current genetics 21 (1992), S. 339-344 
    Details
    ISSN: 1432-0983
    Keywords: Yeast ; Saccharomyces cerevisiae ; Lysis mutants ; Plasmid stability
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The autonomously replicating plasmid YEpSS1, containing the S. cerevisiae SOD1 and SRB1 genes, was highly unstable in a wild-type strain. When transformed into a fragile srb1-1 mutant host, the same plasmid displayed different characteristics depending on the growth medium used. Both batch and continuous culture experiments demonstrated that the plasmid was very unstable when the transformed strain SLU15 was grown in the presence of an osmotic stabiliser (10% w/v sorbitol). However, in the absence of the osmoticum, nearly 100% of the cells retained the plasmid and produced the Sod1 protein after 80 generations of growth.
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    URL: http://dx.doi.org/10.1007/BF00351692
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  • 188
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    Analysis of the chromosomal DNA polymorphism of wine strains of Saccharomyces cerevisiae (1992)
    Springer
    Current genetics 22 (1992), S. 1-7 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Wine yeasts ; Chromosome length polymorphism ; TAFE ; Probe hybridization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Wine yeast strains are characterized by a high chromosomal DNA polymorphism. This can be explained partly by a size difference of different variants of specific chromosomes. This difference can reach up to 45% of the size of the chromosome in question. Two strains, SB1 and Eg8, have a very complex chromosomal pattern and show one band hybridizing with probes from two different chromosomes derived from a reference strain. This is an indication of the presence of “hybrid” chromosomes in these wine strains. The most astonishing result concerns chromosome VIII, frequently present in wine strains in two variant forms. The first normal form has a size of about 580 kb while the second is around 1000 kb. These two forms segregate at meiosis and recombine with a normal chromosome VIII from a laboratory strain. Wine yeasts are thus very different from haploid laboratory strains.
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    URL: http://dx.doi.org/10.1007/BF00351734
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  • 189
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    6-Azauracil inhibition of GTP biosynthesis in Saccharomyces cerevisiae (1992)
    Springer
    Current genetics 22 (1992), S. 9-11 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; IMP dehydrogenase ; 6-azauracil ; GTP level
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The addition of 6-azauracil to the growth medium causes a strong reduction of the GTP level in the nucleotide pool of Saccharomyces cerevisiae. In-vitro experiments show a strong inhibition of IMP dehydrogenase activity by 6-azaUMP explaining the preceeding effect. PPR2 mutants, previously characterized by an increased sensitivity to 6-azauracil compared to the wildtype, are specifically susceptible to the lowering of the GTP pool, and are able to grow in presence of 6-azauracil when guanine is added to the medium.
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    URL: http://dx.doi.org/10.1007/BF00351735
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  • 190
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    Construction and growth properties of a yeast strain defective in sterol 14-reductase (1992)
    Springer
    Current genetics 22 (1992), S. 267-272 
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    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Sterol 14-reductase ; Ergosterol ; Fenpropidin
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We have transformed Saccharomyces cerevisiae with a genomic library contained in the replicative vector pFL44. The resulting transformants were screened for resistance to fenpropidin, a specific inhibitor of sterol 14-reductase. A plasmid was isolated that transformed yeast both to resistance to fenpropidin and to an increased specific activity of sterol 14-reductase. Sterol analysis of transformed cells grown in the presence of increasing concentrations of the inhibitor confirmed that resistance was a consequence of over-production of sterol 14-reductase. By chromosomal gene disruption, we have, for the first time, constructed yeast strains defective in sterol 14-reductase. As expected, since yeast in unable to take up sterols in aerobiosis, the disrupted strains do not grow in the presence of oxygen, even if exogenous sterols are supplied. However, disrupted cells grow in anaerobiosis with exogenous oleic acid and ergosterol supplemens. They also grow in aerobiosis if they bear an additional mutation allowing sterol uptake. In this last growth condition the cells require a “sparking” ergosterol supplementation (25nM) and accumulate ignosterol (ergosta-8, 14-dienol) as the end-product of the sterol pathway. These results reveal that ignosterol is not obviously toxic to yeast membranes and strongly suggest that the molecular basis of the antifungal-activity morpholine and piperidine is directly related to the specific inhibition of ergosterol formation.
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    URL: http://dx.doi.org/10.1007/BF00317919
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    Identification of UAS elements and binding proteins necessary for derepression of Saccharomyces cerevisiae fructose-1,6-bisphosphatase (1992)
    Springer
    Current genetics 22 (1992), S. 363-370 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Fructose-1,6-bisphosphatase ; Glucose repression ; Gene activation ; Gluconeogenesis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Fructose-1,6-bisphosphatase is a key enzyme in gluconeogenesis and the FBP1 gene is not transcribed during growth with glucose. Genetic analysis indicated a positive regulation of FBP1 expression after exhaustion of glucose. By linker-deletion analysis, two upstream activation sites (UAS1 and UAS2) were localized and the respective UAS-binding factors (DAP I and DAP II for derepression activating protein) were identified by gel retardation. UAS1 and UAS2 span about 30 bp each, and are separated by approximately 30 bp. Both UAS sites act synergistically. Although UAS1 showed some similarities to the DNA-binding consensus for the general yeast activator Rap1, competition experiments and DEAE-chromatography proved that DAP I and Rap1 correspond to different proteins. Gel retardation by DAP I depended on carbon sources and did not occur in cells growing logarithmically with glucose, whereas a strong retardation signal was obtained with ethanol-grown cells. The present results suggest that DAP I and DAP II are the final regulatory elements for glucose derepression.
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    URL: http://dx.doi.org/10.1007/BF00352437
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    The expression of a specific 2-deoxyglucose-6P phosphatase prevents catabolite repression mediated by 2-deoxyglucose in yeast (1995)
    Springer
    Current genetics 28 (1995), S. 101-107 
    Details
    ISSN: 1432-0983
    Keywords: 2-deoxyglucose ; 2-deoxyglucose-6P phosphatase ; Catabolite repression ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract 2-deoxyglucose (2-DOG), a non-metabolize analogue of glucose, is taken up by yeast using the same transporter(s) as glucose and is phosphorylated by hexokinases producing 2-deoxyglucose-6-P. We found that in DOG R yeasts, 2-DOG was not able to trigger glucose repression, even at concentrations of 0.5%. This result suggests that the specific 2-DOG-6P phosphatase, the enzyme responsible for the DOG R phenotype, may be involved in inhibiting the process of catabolite repression mediated by 2-DOG
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    URL: http://dx.doi.org/10.1007/BF00315774
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    Carbon catabolite regulation of transcription of nuclear genes coding for mitochondrial proteins in the yeast Kluyveromyces lactis (1995)
    Springer
    Current genetics 28 (1995), S. 267-273 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Kluyveromyces lactis ; Transcriptional regulation ; Catabolite repression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Promoter regions of the KlQCR7, KlQCR8 and KlCYC1 genes, coding for subunits of the bc 1-complex and cytochrome c respectively, in the shortterm Crabtree-negative yeast Kluyveromyces lactis differ markedly in sequence from their Saccharomyces cerevisiae counterparts. They have, however, conserved very similar configurations of binding-site motifs for various transcription factors known to be involved in global and carbon-source regulation in S. cerevisiae. To investigate the carbon source-dependent expression of these genes in K. lactis, we have carried out medium-shift experiments and determined transcript levels during the shifts. In sharp contrast to the situation in S. cerevisiae, the level of expression in K. lactis is not affected when glucose is added to a non-fermentable carbon-source medium. However, the genes are not constitutively expressed, but become significantly induced when the cells are shifted from glucose to a nonfermentable carbon source. Finally, induction of transcriptional activation does not occur in media containing both glucose and non-femmentable carbon sources.
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    Analysis of exon and intron mutants in the cytochrome b mitochondrial gene of Saccharomyces cerevisiae (1996)
    Springer
    Current genetics 30 (1996), S. 200-205 
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    ISSN: 1432-0983
    Keywords: Key words Cytochrome b ; Mutants ; Mitochondria ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The nucleotide changes present in a group of five cytochrome b mit– mutants were analyzed at the sequence level. Two single-base changes were found: one (M10-152) generated a nonsense codon in the first exon while the other (M8-181) created a missense substitution in the second exon. The other mutants all have multiple (three) substitutions that either resulted in a missense mutation in a coding region (M17-162) or else changed nucleotides in the last intron of the gene, so blocking its excision (M6-200 and M8-53). The synthesis of mitochondrial polypeptides and the steady state concentration of the complex-III subunits were examined. The Rieske protein and the core-4 and core-5 subunits were much reduced in all mutants. Consequently the overall stability of complex III is very sensitive even to amino-acid substitutions in the cytochrome b protein. Mutant M8-53 provides direct evidence for the proposed role of the P9.1 stem in the core structure of the group-I type last intron of this gene.
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    Expression and secretion of the Candida wickerhamii extracellular β-glucosidase gene, bglB, in Saccharomyces cerevisiae (1996)
    Springer
    Current genetics 30 (1996), S. 417-422 
    Details
    ISSN: 1432-0983
    Keywords: Key wordsβ-glucosidase ; Candida wickerhamii ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The yeast Candida wickerhamii exports a cell-associated β-glucosidase that is active against cellobiose and all soluble cellodextrins. Because of its unique ability to tolerate end-product inhibition by glucose, the bglB gene that encodes this enzyme was previously cloned and sequenced in this laboratory. Using several different promoters and constructs, bglB was expressed in the hosts Escherichia coli, Pichia pastoris, and Saccharomyces cerevisiae. Expression was initially performed in E. coli using either the lacZ or tac promoter. This resulted in intracellular expression of the BglB protein with the protein being rapidly fragmented. Secretion and glycosylation of active β-glucosidase was achieved with several different S. cerevisiae constructs utilizing either the adh1 or the gal1 promoter on 2-µ replicating plasmids. When either the invertase (Suc2) or the BglB secretion signal was used, BglB protein remained associated with the cell wall and appeared to be hyperglycosylated. Expression in P. pastoris was also examined to determine if higher activity and expression could be achieved in a yeast host that usually does not hyperglycosylate. Using the alcohol oxidase promoter in conjunction with either the pho1 or the α-factor secretion signal, the recombinant enzyme was successfully secreted and glycosylated in P. pastoris. However, levels of protein expression from the chromosomally integrated vector were insufficient to detect activity.
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    URL: http://dx.doi.org/10.1007/s002940050151
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    Bleomycin hydrolase (Blh1p), a multi-sited thiol protease in search of a distinct physiological role (1997)
    Springer
    Current genetics 32 (1997), S. 41-51 
    Details
    ISSN: 1432-0983
    Keywords: Key words Bleomycin hydrolase ; Saccharomyces cerevisiae ; Thiol proteases ; Protein amphitropism ; Processing of glycosyl-phosphatidylinositol (GPI) anchor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Bleomycin hydrolase, Blh1p, from yeast was co-purified with Gce1p, a cAMP-binding ectoprotein, anchored to the plasma membrane by a glycosyl-phosphatidylinositol (GPI) anchor. Blh1p is a hydrophilic thiol protease lacking transmembrane domains. We have used polyclonal antibodies to study the topology of the over-expressed protein in yeast and have found that it is amphitropic. Part of Blh1p is associated with plasma membranes, and most of the rest occurs in the cytosol. Both the growth conditions and calcium were found to have minor influences on the topology of Blh1p, in that glucose and the earth-alkali ion slightly enhanced recruitment to the membrane. We have examined the possibility that co-purification of Blh1p with Gce1p has a functional basis, and have observed that over-expression of BLH1 in yeast leads to an acceleration of the glucose-induced amphiphilic to hydrophilic conversion of Gce1p, wherein Blh1p could either directly catalyse the proteolytic removal of the polar headgroup of the GPI anchor subsequent to an initial lipolytic cleavage by a GPI-specific phospholipase C or indirectly modulate the reaction. The data show that a thiol protease is involved, but point to an indirect role of Blh1p in GPI processing. Proteases with similar or overlapping substrate specificity are likely to exist, since deletion of BLH1 neither entails a growth defect on any carbon source tested, nor the loss of proteolytic processing of the GPI anchor of Gce1p. Reduced proteolytic GPI processing is, however, observed in the blh1 mutant and the corresponding acceleration in the respective BLH1 multi-copy transformant.
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    Reciprocal translocation at duplicated RPL2 loci might cause speciation of Saccharomyces bayanus and Saccharomyces cerevisiae (1998)
    Springer
    Current genetics 33 (1998), S. 345-351 
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    ISSN: 1432-0983
    Keywords: Key wordsSaccharomyces bayanus ; Saccharomyces cerevisiae ; Translocation ; Speciation ; Duplicated gene ; RPL2
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract By a genomic comparison of two sibling yeasts, Saccharomyces bayanus and S. cerevisiae, we previously demonstrated that chromosomes II and IV of S. cerevisiae were rearranged into chromosomes 12 and 14 of S. bayanus or vice versa. In the present study we have delimited the translocation break sites in chromosomes II and IV by Southern hybridization using DNA fragments of S. cerevisiae cosmid clones as probes. The results suggest that the reciprocal translocation of chromosomes II and IV had occurred at duplicated RPL2 loci. Furthermore, the translocation sites in S. bayanus were confirmed by the cloning and sequence analysis of the regions flanking RPL2 loci. Several genes in the regions flanking the RPL2 loci were present in the order expected for a translocation at these loci between the two species. These results indicated that the reciprocal translocation between chromosomes II and IV was generated by homologous recombination at duplicated RPL2 loci on the two chromosomes. Therefore, we propose that duplicated genes or duplicated regions play an important role in altering genomic organization during the speciation of S. bayanus and S. cerevisiae.
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    Functional analysis of upstream activating elements in the promoter of the FBP1 gene from Saccharomyces cerevisiae (1998)
    Springer
    Current genetics 33 (1998), S. 406-411 
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    ISSN: 1432-0983
    Keywords: Key words Fructose-1 ; 6-bisphosphatase ; Catabolite repression ; Gluconeogenesis ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have investigated the effect of different carbon sources and of different mutations on the capacity of two elements, UAS1 and UAS2, from the promoter of the FBP1 gene to form specific DNA-protein complexes and to activate expression of a reporter gene. The complexes are observed with nuclear extracts from yeast derepressed on glycerol or ethanol. When hxk2 mutants are grown on glucose the nuclear extracts are able to complex UAS1 but not UAS2, while for wild-type cells grown on galactose only the complex with UAS2 is formed. In contrast, in vivo the operation of both UASs is high in ethanol, moderate to low in glycerol, and negligible in galactose; no expression is observed in glucose even in a hxk2 background. There is no effect of a MIG1 deletion, either in the formation of DNA-protein complexes or on the expression of reporter genes.
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    Positive and negative elements involved in the differential regulation by heme and oxygen of the HEM13 gene (coproporphyrinogen oxidase) in Saccharomyces cerevisiae (1995)
    Springer
    Current genetics 28 (1995), S. 503-511 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; HEM13 regulation ; Heme and oxygen ; CYP1, ROX1, SSN6, TUP1
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The Saccharomyces cerevisiae HEM13 gene codes for coproporphyrinogen oxidase (CPO), an oxygen-requiring enzyme catalysing the sixth step of heme biosynthesis. Its transcription is increased 40–50-fold in response to oxygen- or heme-deficiency. We have analyzed CPO activity and HEM13 mRNA levels in a set of isogenic strains carrying single or double deletions of the CYP1 (HAP1), ROX1, SSN6, or TUPI genes. The cells were grown in the presence or absence of oxygen and under heme-deficiency (hem1Δ background). Both Rox1p and Cyp1p partially repressed HEM13 in aerobic heme-sufficient cells, probably in an independent manner. In the absence of heme, Cyp1p activated HEM13 and strongly repressed ROX1, allowing de-repression of HEM13. Cyp1p had no effect on HEM13 expression in anaerobic cells. Deletions of SSN6 or TUP1 dramatically de-repressed HEM13 in aerobic cells. A series of deletions in the HEM13 promoter identified at least four regulatory regions that are required for HEM13 regulation. Two regions, containing motifs similar to the Rox1p consensus sequences, act as repression sites under aerobic growth. The two other sites act as activation sequences required for full induction under oxygen- or heme-deficiency. Taken together, these results suggest that induction of HEM13 occurs in part through relief of repression exerted by Rox1p and Cyp1p, and in part by activation mediated partly by Cyp1p under heme-deficiency and by unknown factors under oxygen-deficiency.
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    Characterization of a novel α-amylase from Lipomyces kononenkoae and expression of its gene (LKA1) in Saccharomyces cerevisiae (1995)
    Springer
    Current genetics 28 (1995), S. 526-533 
    Details
    ISSN: 1432-0983
    Keywords: α-Amylase ; Lipomyces kononenkoae ; LKA1 ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A highly active α-amylase (76 250 Da) secreted by the raw starch-degrading yeast Lipomyces kononenkoae strain IGC4052B was purified and characterized. Using high performance liquid chromatography (HPLC), end-product analysis indicated that the L. kononenkoae α-amylase acted by endo-hydrolysis on glucose polymers containing α-1,4 and α-1,6 bonds, producing mainly maltose, maltotriose and maltotetraose. The following NH2-terminal amino acids were determined for the purified enzyme: Asp-Cys-Thr-Thr-Val-Thr-Val-Leu-Ser-Ser-Pro-Glu-Ser-Val-Thr-Gly. The L. kononenkoae α-amylase-encoding gene (LKA1), previously cloned as a cDNA fragment, was expressed in Saccharomyces cerevisiae under the control of the PGK1 promoter. The native signal sequence efficiently directed the secretion of the glycosylated protein in S. cerevisiae. De-glycosylation of the enzyme indicated that post-translational glycosylation is different in S. cerevisiae from that in L. kononenkoae. Zymogram analysis indicated that glycosylation of the protein in S. cerevisiae had a negative effect on enzyme activity. Southern-blot analysis revealed that there is only a single LKA1 gene present in the genome of L. kononenkoae.
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    URL: http://dx.doi.org/10.1007/BF00518165
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