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  • Immunocytochemistry  (352)
  • Yeast  (235)
  • Springer  (587)
  • American Chemical Society
  • 1985-1989  (365)
  • 1980-1984  (222)
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Year
  • 101
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    Springer
    Current genetics 6 (1982), S. 93-98 
    ISSN: 1432-0983
    Keywords: Yeast ; Genetic mapping ; Trisomic analysis ; Arginine
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary By use of a set of 8 aneuploid strains of the yeast Saccharomyces cerevisiae, carrying from 1 to 5 identified disomic chromosomes, in crosses to a set of haploid strains collectively bearing 11 unmapped genes, the following chromosome assignments were obtained for these unmapped genes: arg80 on XIII;arg3 on X;car2 on XII; cpa1 and tsm8740 on XV; tsm7269 (=rna6) on II; cpa2 on X or XV; arg82 and tsm4572 on III, IV or XVI; car1 and arg81 on II, IV, VI, VII or XVI. Linkage tests between the unmapped genes and markers located on the chromosomes that had been designated as possible carriers by the previous analysis allowed 8 genes to be localized. The remaining three genes, cpa2, car1 and arg81 (located on fragment F8), could not be positioned on any of the chromosomes indicated by the trisomic analysis, in spite of testing for linkage to markers covering most of the known regions of these chromosomes.
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  • 102
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    Current genetics 8 (1984), S. 49-55 
    ISSN: 1432-0983
    Keywords: Protoplast fusion ; Yeast ; Yarrowia lipolytica ; Kluyveromyces lactis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Prototrophic hybrids have been obtained by the fusion of auxotrophic haploid strains of the two yeasts Yarrowia (Saccharomycopsis) lipolytica and Kluyveromyces lactis. The hybrid fusants had a colonial morphology intermediate between that of the two parent strains, were uninucleate, and contained an approximately diploid amount of DNA per cell. The growth rates of all the fusants on a minimal glucose medium were slower than those of the two parents. Two of the fusants studied could utilise a novel range of carbon sources. All of these data suggested that the hybrids contained a diploid nucleus formed by the fusion of the two haploid parental nuclei. However, analytical CsCl density gradient centrifugation demonstrated that the nuclear DNA of the fusants was derived almost entirely from the Y. lipolytica parent. Moreover, an examination of the protein constitution of the fusants by two-dimensional gel electrophoresis showed that their protein patterns were indistinguishable from that of Y. lipolytica. Two possible mechanisms for the formation of a diploid nucleus containing DNA derived almost entirely from one of the haploid parents are discussed.
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  • 103
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    Current genetics 9 (1985), S. 279-284 
    ISSN: 1432-0983
    Keywords: Virus-like particles ; Double-stranded RNA ; Yeast ; Yarrowia lipolytica
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Four out of the 24 strains of the yeast Yarrowia lipolytica we have checked for the presence of virus-like particles (VLPs) proved to contain encapsidated double-stranded RNA (dsRNA) molecules, 4.9 kb long. A major VLP polypeptide of MW 80,000 was observed in all 4 cases, and a second one of MW 77,000 in three cases. dsRNA from the VLPs harboring only the larger polypeptide showed little homology with the 3 others. We have found no homology between VLP dsRNAs and host DNA or dsRNAs from Saccharomyces cerevisiae, and no relationship between the presence of VLPs and a possible killer phenomenon in Y. lipolytica.
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  • 104
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    Current genetics 7 (1983), S. 473-480 
    ISSN: 1432-0983
    Keywords: ars sequences ; Yeast ; Chlamydomonas
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A hybrid plasmid (pLG4) containing pBR325 and the yeast arg4 gene was constructed then used to isolate DNA fragments of Chlamydomonas able to promote high frequency transformation of yeast. Three plasmids containing EcoRI restriction fragments of chloroplast DNA and two plasmids containing Aval fragments of nuclear DNA were shown to support autonomous replication of plasmids in yeast. The three EcoRI fragments correspond to restriction fragments R4, R5 and R11 of native chloroplast DNA. These fragments are clustered in the physical map of chloroplast DNA constructed by Rochaix (1978). All isolated plasmids were shown to transform yeast at high frequency but the yeast transformants were quite unstable mitotically. Potential cloning sites are still available in the new plasmids which could be used as vectors in yeast and possibly in Chlamydomonas itself.
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  • 105
    ISSN: 1432-0983
    Keywords: D-xylose fermentation ; Yeast ; Protoplast fusion ; Ploidy
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Protoplast fusion technique was employed in the preparation of presumptive diploid, triploid and tetraploid strains of the D-xylose fermenting yeast C. shehatae CBS2779. Prototrophic selection technique was employed for the recovery of presumed fusant strains. The hybrid nature of the presumptive diploid, triploid and tetraploid strains was confirmed by analysing I) the nuclear condition; II) the cell size and the cell volume of the parental and fusant strain; III) the cellular DNA content and IV) the induced and spontanenous mitotic segregation of properties in these strains. The increased level of ploidy was found to have an effect on the rate of ethanol production from D-xylose.
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  • 106
    ISSN: 1432-0983
    Keywords: Yeast ; Mitochondrial DNA ; Antibiotic resistance mutations ; Suppressor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Strains that are genotypically sensitive to chloramphenicol and also contain one of the nuclear suppressors of mitochondrial chloramphenicol resistance (Waxman et al. 1979) were constructed. A manganese mutagenesis on such a strain produced chloramphenicol resistant mutants, most of which resulted from mutations in nuclear genes. These mutants may be either dominant or recessive, and they probably do not code for membrane proteins. The few mitochondrial mutants fall into several classes, but all result from mutations in the 21S rRNA gene. The suppressor allele effectively prevents the appearance of the most common group of mitochondrial mutants (those that map at cap1), and thereby enhances the selection of novel mutants in the region.
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  • 107
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    Current genetics 9 (1985), S. 533-538 
    ISSN: 1432-0983
    Keywords: cDNA hybridisation ; UV inducible RNA ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Differential colony hybridisation has been used to identify DNA sequences in Saccharomyces cerevisiae corresponding to RNA transcripts whose levels increase 5–10 fold following UV-irradiation. Four sequences have been identified, three of which share sequence homology and hybridise to the same set of genomic DNA fragments. The fourth sequence appears to be distinct, however each DNA sequence hybridises to a similar sized RNA transcript which is approximately 4.0 kb long. The relationships between these DNA sequences and their potential protein products is discussed.
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  • 108
    ISSN: 1432-0983
    Keywords: Yeast ; TEF genes ; Gene disruption
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Two genes,TEF1 andTEF2, encode the protein elongation factor EF-1α in the yeastSaccharomyces cerevisiae. We have generated yeast haploid strains containing eitherTEF1 orTEF2 interrupted by insertion of a large piece of foreign DNA. Cells which contain either one functional copy of the EF-1α genes are viable. In contrast, attempts to isolate a yeast haploid strain with bothTEF1 andTEF2 inactivated have failed suggesting that the double gene disruption is a lethal event.
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  • 109
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    Current genetics 3 (1981), S. 213-220 
    ISSN: 1432-0983
    Keywords: Mitochondrial genetics ; Yeast ; Suppressiveness ; Triploid analysis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A genetically defined highly suppressive petite yeast strain (ρ −cob+AsEoCoOoPo) was crossed with a grande strain carrying a multiply marked mitochondrial genome (ρ +ArErCrO rpr). Petite diploid progeny, isolated from individual zygotic clones consisting either of wholly petite or mixtures of grande and petite cells, were characterised genetically by crossing to grande haploids. The diploid petites were found to closely resemble the petite parent and in general not to carry mitochondrial markers from the grande parent. In the petites from the mixed clones recombination was detected, but only within the region of homology between the genomes. These observations are inconsistent with models of suppressiveness based on destructive recombination and suggest that the petite genome eliminates the grande genome from zygotic progeny through being preferentially replicated. The most plausible model to explain the observed pattern of zygotic clones postulates a limited number of mDNA replication sites in zygotes, competition for sites between input mDNA molecules and an advantage in this competition for suppressive ρ − mDNA.
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  • 110
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    Current genetics 3 (1981), S. 229-233 
    ISSN: 1432-0983
    Keywords: Yeast ; Nascent DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Two species of newly synthesised DNA larger than average replicons have been found in yeast. Their molecular weights are 60 million and 90 million daltons respectively. The exact nature of these molecules is not certain. They may represent entirely novel species of cellular DNA or they could be concatameric replication intermediates of some particular fraction of DNA, such as mitochondrial DNA or rDNA. Alternatively they could result from the fusion of adjacent completed replicons in a small cluster.
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  • 111
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    Current genetics 5 (1982), S. 153-155 
    ISSN: 1432-0983
    Keywords: Yeast ; Mutant cell-wall ; Permeability exponentialy
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary When Saccharomyces cerevisiae SY15 rho° mutant cells grown in media stabilized with 10% sorbitol were suspended in 2% sorbitol solutions, 60–70% of the population did not lyse and became permeable to native high molecular weight DNA. Maximal incorporation of DNA to DNase resistant state was measured after 60 min of incubation in presence of 5 μg/ml DNA and 10 mM CaCl2. These results suggest that the fragile mutants might be tested as hosts for transformation of whole yeast cells.
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  • 112
    ISSN: 1432-0983
    Keywords: Yeast ; Mitochondria ; Intragenic recombination ; Mutant polypeptides
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Genetic and biochemical studies were performed with mutants allocated to the mitochondrial oxi2 gene. Recombinational analysis of 19 oxi2 mutants was performed using α and a mutant strains derived from the same genetic background. The frequencies of wild-type recombinants in oxi2 − × oxi2 − crosses varied from 0.002 to 17%. The map of oxi2 mutations constructed on the basis of these frequencies shows many internal inconsistencies. In the course of rho − deletion mapping five classes of oxi2 mutations were distinguished. The results of deletion analysis are in agreement with those of recombinational mapping. The analysis of mitochondrial translation products by SDS-polyacrylamide electrophoresis of 20 oxi2 mutants shows that 17 of them are connected with conspicuous changes of 22 kd polypeptide band corresponding to subunit III of cytochrome oxidase. At least four of them carried instead of subunit III clearly visible significantly shorter polypeptides (12.8 to 20.1 kd). These were, most likely, shorter fragments of subunit III resulting from chain termination mutations. Colinearity was observed between the lenght of new polypeptides and the positions of the respective mutations on the recombinational map. These data confirm hat oxi2 encodes subunit III of cytochrome oxidase and suggest that translation of the oxi2 gene is in the direction from V303 to V273.
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  • 113
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    Current genetics 7 (1983), S. 285-288 
    ISSN: 1432-0983
    Keywords: Arginyl-tRNA-Protein transferase ; Yeast ; Post-translational modification
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A mutant of Saccharomyces cerevisiae deficient in arginyl-tRNA-protein transferase has been isolated. The responsible mutation designated ate1, was localized near the centromere of chromosome VII. It probably involves the structural gene for the transferase since residual enzyme activity in the mutant is temperature-sensitive.
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  • 114
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    Current genetics 7 (1983), S. 489-492 
    ISSN: 1432-0983
    Keywords: Mitochondrial genes ; Yeast ; Vegetative segregation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Genes in mitochondria and chloroplasts segregate rapidly during vegetative reproduction. Models to explain this vegetative segregation invoke either random segregation of organelle DNA molecules, or nonrandom segregation with random recombination events. All such models are basically stochastic. To look at vegetative segregation we took heteroplasmic (HET) cells containing mitochondrial mutations at the cap1, eryl and olil loci from several crosses. HETs were repeatedly selected and subcloned. Even after three to five successive subclonings (approximately 60–100 generations) some cells remained heteroplasmic. This confirms and extends previous observations of persistent HETs by Rank and Bech-Hansen (1972) and Forster and Kleese (1975), and by Bolen et al. (1980) for chloroplast genes in Chlamydomonas.
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  • 115
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    Current genetics 8 (1984), S. 29-32 
    ISSN: 1432-0983
    Keywords: Antisuppression ; Suppression ; tRNA ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The effect of a previously isolated antisuppressor mutation from bakers' yeast, that reduced the efficiency of the tyrosine-inserting ochre suppressor, SUP7-o, on other tyrosine-inserting ochre suppressors has been determined. As expected, the antisuppressor mutation, mod5-1, restricted the capacity of all eight tyrosine-inserting ochre suppressors to suppress nonsense mutations. Based on the suppression of five ochre alleles in the presence of mod5, the eight class I suppressors can be grouped into three subclasses. The most efficient subclass had only one member, SUP4-o. Members of the second group included SUP2-o, SUP3-o, SUP7-o, and SUP8-o. The third and least efficient subclass included SUP5-o, SUP6-o, and SUP1 1-o. These differences in efficiencies are a function of the relative expression of the eight genes encoding tRNATYR.
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  • 116
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    Current genetics 8 (1984), S. 85-92 
    ISSN: 1432-0983
    Keywords: Chromosome map ; Yeast ; Schizosaccharomyces pombe ; Gene conversion
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The genetic map of the nuclear genome of the fission yeast Schizosaccharomyces pombe has been extended by mitotic and meiotic mapping data. A total of 158 markers are now assigned to the three linkage groups known in this organism, and 118 of them have been located on the corresponding chromosome map. Chromosome II and III each consist of one linkage group. There is some indication that the two large fragments which define chromosome I are meiotically linked, but the linkage observed is significant at the P = 0.05 level only. The length of the map is at least 1,700 map units, corresponding to an average of about 8 kilobases per map unit. The latter figure is comparable to the one obtained for intragenic recombination in the sup3 gene (Hofer et al. 1979). The basic frequency of gene conversion as measured for 21 genes varies according to a distribution of Poisson (with a modal value of 0.6% conversion per meiosis and per gene), in sharp contrast with Saccharomyces cerevisiae (Fogel et al. 1980) and Ascobolus immersus (Nicolas 1979). This may reflect the rarity of gene or region-specific rec alleles in S. pombe and may be related to the homothallism of this organism.
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  • 117
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    Current genetics 8 (1984), S. 353-358 
    ISSN: 1432-0983
    Keywords: Hygromycin B ; Yeast ; Plasmids
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Saccharomyces cerevisiae is normally sensitive to the drug hygromycin B; a hygromycin B concentration of 200 µg/ml in agar plates is sufficient to completely inhibit growth. We constructed yeast-E. coli bifunctional plasmids which confer hygromycin B resistance to Saccharomyces cerevisiae. Promoters and amino terminal coding regions of a heat shock gene, a heat shock cognate gene, and the phosphoglycerate kinase gene from yeast were fused to a bacterial hygromycin B resistance gene. In all three cases, yeast cells containing plasmids with the hybrid hygromycin B resistance gene were resistant to high levels of the drug. Yeast cells containing these plasmids can also be directly selected after transformation by using hygromycin B. The intact bacterial hygromycin B resistance gene and the kanamycin resistance gene from Tn903 were also tested in yeast for their ability to confer resistance to hygromycin B and G418. The intact bacterial genes were not effective in conferring drug resistance to yeast cells.
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  • 118
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    Current genetics 7 (1983), S. 85-92 
    ISSN: 1432-0983
    Keywords: Yeast ; RAD52 ; Cloning ; S1 and BAL31 Deletions
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The RAD52 gene of Saccharomyces cerevisiae has previously been shown to be involved in both recombination and DNA repair. Here we report on the cloning of this gene. A plasmid containing a 5.9 kb yeast DNA fragment inserted into the BamH1 site of the YEp13 vector has been isolated and shown to complement the X-ray sensitive phenotype of the rad52-1 mutation. The rad52-1 cells containing the plasmid form larger colonies than similar cells having lost the plasmid. This plasmid has been shown not to complement either the U.V. sensitivity or the recombination defect of the E. coli recA mutation. From the insert various fragments have been subcloned into the YRp7 and YIp5 vectors. Integration events of two of the subclones have been genetically mapped to the chromosomal location of RAD52, indicating that the structural gene has been cloned. A 1.97 kb BamH1 fragment subcloned into YRp7 in one orientation complements the rad52-1 mutation, while the same fragment in the opposite orientation fails to complement. Various other subclones indicate that a BglII site, within the BamH1 fragment, is in the RAD52 gene. This BglII site has been deleted by Sl-nuclease digestion and the resulting deletion inactivates the RAD52 gene. BAL31 deletions from one end of a 1.9 kb Sal1-BamH1 fragment have been isolated; up to 0.9 kb can be deleted without loss of RAD52 activity, indicating that the RAD52 gene is approximately 1 kb or less in length.
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  • 119
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    Current genetics 8 (1984), S. 69-76 
    ISSN: 1432-0983
    Keywords: Yeast ; Ethidium bromide ; Meiosis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Ethidium bromide was found to inhibit nuclear and mitochondrial DNA synthesis during meiosis which resulted in the inhibition of meiotic gene conversion and sporulation and was also lethal. Protection from the effects of ethidium bromide on meiotic gene conversion and survival was found to coincide with DNA synthesis, but it is possible that protection from sporulation inhibition occurs only later in meiosis.
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  • 120
    ISSN: 1432-0983
    Keywords: Yeast ; cdc8-1 mutation ; Mitotic recombination
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary In a diploid strain homozygous for the cdc8-1 mutation, a block in DNA synthesis caused by restrictive temperature resulted in a significant increase in the frequency of intragenic recombination at the HOM2 locus. Under restrictive conditions, incorporation of radioactivity into DNA was reduced to 2% of the control and alkaline sucrose gradient centrifugation revealed that only short DNA fragments were synthesized. There was no considerable fragmentation of template DNA during incubation of cdc8-1 strains under restrictive conditions.
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  • 121
    ISSN: 1432-0983
    Keywords: Telomeres ; Recombination ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Natural termini from macronuclear DNA of the ciliated protozoans Tetrahymena thermophila and Oxytricha fallax can support telomere formation in yeast. However, plasmids carrying these ciliate termini are modified by the addition of DNA which hybridizes to the synthetic oligonucleotide poly [d(C-A)], a sequence which also hybridizes to terminal restriction fragments from yeast chromosomes but not to Tetrahymena or Oxytricha macronuclear DNAs. Thus, in yeast, the creation of new telomeres on ciliate termini involves the acquisition of yeast-specific terminal sequences presumably by either recombination or non-templated DNA synthesis. The RAD52 gene is required for the majority of yeast mitotic and meiotic recombination events. Moreover, the absence of an active RAD52 gene product results in high rates of chromosome loss. Here we demonstrate that terminal restriction fragments from Tetrahymena macronuclear ribosomal DNA (rDNA) support the formation of modified telomeres in a yeast strain carrying a defect in the RAD52 gene. Moreover, linear plasmids bearing these modified ciliate termini are stably propagated in rad52 − cells.
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  • 122
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    Current genetics 8 (1984), S. 333-340 
    ISSN: 1432-0983
    Keywords: polA+ ; DNA polymerase I ; Cloning ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The E. coli polA + gene has been subcloned from a specialised λ transducing phage onto a low copy number plasmid. Plasmid-encoded DNA polymerase I was synthesised at 2 to 3 times the wild-type E. coli level, and was biochemically indistinguishable from chromosomally-encoded protein. It was able to counteract the radio sensitivity of polA1, polAex1, polAex2 and polA12 mutants, but no complementation of polA107 mutants occurred, even though the plasmid polA+ gene was expressed. S. cerevisiae ars-1 or 2 μ replicative sequences were introduced into the polA+ plasmid. Transformation of yeast with these constructs increased total DNA polymerase levels 2–20 times, depending upon assay conditions. The additional activity was discriminated from yeast DNA polymerases by its ability to use low concentrations of substrate, by its resistance to chemical inhibition, and by co-electrophoresis with pure DNA polymerase I and its proteolytic fragments. The polA+ gene was expressed in yeast without the aid of yeast promotor sequences. However, deletion of cloned DNA more than 99 base pairs in front of the structural gene prevented expression in yeast but not in E. coli, indicating that the two organisms use different sequences for expression of the plasmid polA+ gene.
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  • 123
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    Current genetics 8 (1984), S. 471-475 
    ISSN: 1432-0983
    Keywords: β-Glucanase ; Expression ; Heterologous DNA ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A cloned endo-1,3-1,4-β-glucanase gene from the Gram-positive bacterium B. subtilis has been located by deletion analysis on a 1.4 kb PvuI-ClaI DNA fragment. This gene has been sub-cloned in the yeast LEU2 vector pJDB207 to produce a hybrid plasmid designated pEHB9. pEHB9 has been transformed to S. cerevisiae and shown to direct the synthesis of an endo-1,3-1,4-β-glucanase in yeast. The β-glucanase activity was low and could only be detected in crude cell extracts of yeast harbouring pEHB9.
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  • 124
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    Current genetics 9 (1985), S. 259-262 
    ISSN: 1432-0983
    Keywords: Yeast ; Ty1 ; Trans ; Deletion
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary DEL1 strains of the yeast Saccharomyces cerevisiae exhibit a high rate of deletions of the three linked genes, CYC1, OSM1, and RAD7. Classical genetic methods showed that DELI segregated as a single Mendelian gene closely linked to CYC1. In addition, genetic evidence suggested that DEL1 was both cis- and trans-dominant (Liebman et al. 1979). Molecular analysis of deletions isolated from a haploid DEL1 strain established that deletion formation was mediated by recombination between yeast transposable elements, Ty's (Liebman et al. 1981). We now report the molecular characterization of deletions isolated from diploids in the trans configuration. This analysis reveals that these deletions probably arose in a two-step process involving mitotic recombination followed by Ty-mediated deletion formation in cis.
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  • 125
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    Current genetics 9 (1985), S. 285-291 
    ISSN: 1432-0983
    Keywords: Killer toxin ; Plasmid selection ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Transformants of sensitive yeast strains containing an expressed cDNA copy of the yeast killer toxin-immunity gene could be selected for by exposure to added killer toxin. For strain AH-22 the transformation frequency was approximately 10% that obtained by selection for leucine prototrophy. The procedure required time for expression of immunity prior to selection, and a screening step to remove non-transformed survivors. Under conditions where active toxin was produced, transformants containing the toxin-immunity gene were at a selective advantage, and cells losing the plasmid were killed. This resulted in self selection of transformants, and affords a way of maintaining plasmid stability in protrophic strains.
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  • 126
    ISSN: 1432-0983
    Keywords: Regulation ; Urea ; Catabolism ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Urea amidolyase and the high affinity urea uptake system are induced by allophanate. durM − and durL − recessive mutations, which are easily obtained, totally prevent this induction. They are not linked to each other nor to the concerned structural genes. Despite an intensive hunt, no mutation of repressor or classical operator type has been selected. We conclude that urea amidolyase and urea uptake induction involves at least two positive elements coded for by the durM and durL genes.
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  • 127
    ISSN: 1432-0983
    Keywords: Yeast ; Catalase ; Trehalose ; Glycogen
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Mutations at the GLC1 locus in Saccharomyces cerevisiae result in a major deficiency in synthesis of catalase T, but do not affect catalase A. Three independent glc1 mutations were shown to have the same pleiotropic phenotype: catalase T deficiency, defective glycogen synthesis and defective trehalose accumulation. These three deficiencies appear to be determined by a single, nuclear gene. The possibility that glc1 mutations alter a protein kinase is considered.
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  • 128
    ISSN: 1432-0983
    Keywords: Arginine catabolism ; Regulation ; Ornithine transaminase ; Double induction ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Yeast ornithine transaminase is known to be induced by arginine and ornithine, through the action of regulatory elements common to arginase induction. We show here that it is subject to a second induction circuit, that which is responsible for urea amidolyase and urea permease induction by allophanate and defined by the regulatory mutants durL − and durM −
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  • 129
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    Current genetics 7 (1983), S. 69-72 
    ISSN: 1432-0983
    Keywords: Yeast ; Protoplast ; Cybrid ; Plasmid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Data presented here demonstrate that fusion of protoplasts of two different haploid strains of Saccharomyces cerevisiae having the same mating type leads to the formation of “fusants” and “cytoplasmic hybrids”. The nuclear and cytoplasmic genome of a “fusant” combine those of the parent haploid strains. The “cytoplasmic hybrid” possesses the haploid genome of one parent and the combined cytoplasmic genomes of both. In mouse cells lines such products have been termed “cybrids” and this term has therefore been adopted here (Bunn and Wallace 1974).
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  • 130
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    Current genetics 7 (1983), S. 93-100 
    ISSN: 1432-0983
    Keywords: Yeast ; RAD genes ; Cloning
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Plasmids that complement the yeast mutations rad50-1, rad51-1, rad54-3 and rad55-3 were obtained by transforming strains that carried a leu2 marker and the particular rad mutation, with YEp13 plasmids containing near random yeast DNA inserts. Integration of these plasmids or of fragments of these plasmids was accomplished. Genetic studies using the integrants established the presence of the genes RAD51, RAD54 and RAD55 in the respective plasmids. However, a BamHI subclone of the rad50-1 complementing plasmid failed to integrate at the RAD50 locus, indicating that no homology exists between this fragment and the RAD50 gene. A BamHI fragment from the RAD54 plasmid was shown to be internal to the RAD54 gene: its integration within a wild type copy of RAD54 causes the cell to become Rad−; its excision is X-ray inducible and restores the Rad+ phenotype. Since cells bearing a disrupted copy of RAD54 are able to survive, we conclude that this gene is not essential.
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  • 131
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    Current genetics 6 (1982), S. 99-103 
    ISSN: 1432-0983
    Keywords: Ultraviolet light mutagenesis ; Mitochondrial genome ; Meiosis ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Clones derived from ascospores from ultraviolet irradiated diploid cells were examined for the genetic determinants or respiratory properties. Approximately 10% of the cells produced petites of mitochondrial origin at the dose applied. Among 13 asci which produced mitochondrial petites with high frequencies, 6 asci of uniparental type, 0 grandes : 4 petites, were observed. Furthermore, most of the petite spore clones from each individual uniparental ascus showed similar levels of suppressiveness and of mitochondrial gene retention. From these results it is suggested that a single mitochondrial genome participates meiosis in yeast.
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  • 132
    ISSN: 1432-0983
    Keywords: Yeast ; Mutant ; Uncoupler ; Resistance ; Permeability
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    Topics: Biology
    Notes: Summary One mutant resistant to carbonylcyanide m-chlorophenylhydrazone (CCCP), an uncoupler of oxidative phosphorylation, was isolated in Saccharomyces cerevisiae. Genetic analysis showed that a single nuclear gene is responsible for increased resistance; this gene was dominant. The mutant showed cross-resistance or collateral sensitivity to several chemically-unrelated inhibitors (cycloheximide, dinitrophenol, tributhyltin chloride, chloramphenicol). The resistance of the mutant is related to a decreased uptake of CCCP which is not expressed in glucose-starved cells. It was shown that glucose induced a CCCP efflux which was more efficient in the mutant than in the wild-type cells. This effect was correlated to a greater acidification of the internal pH by glucose addition in the mutant cells. It was proposed that resistance was not due to a change of permeability of the plasmic membrane itself but to the change of internal pH which determines the extent of accumulation of weak acids or bases.
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  • 133
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    Current genetics 8 (1984), S. 567-573 
    ISSN: 1432-0983
    Keywords: Non-Mendelian ; Yeast ; Suppressor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Omnipotent suppressors cause translational ambiguity and have been associated with poor growth and inviability. We now report that a non-Mendelian element, [eta+], causes this inviability. In [eta−] strains the suppressors are not inviable. The [eta+] genetic element segregates to about 70% of the meiotic progeny, although almost all of the spores probably have the [eta+] phenotype for the first few divisions. Growth on 5 mM guanidine hydrochloride efficiently causes [eta+] strains to become [eta−]. The [eta+] factor has many similarities with the previously described [psi+] factor (Cox 1965, 1971). However, [eta+] and [psi+] differ in their patterns of inheritance, and by the fact that [psi+] affects ochre specific and not omnipotent suppressors, while the converse is true of [eta+].
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  • 134
    ISSN: 1432-0983
    Keywords: Yeast ; Ribosomal protein gene ; Sequence analysis ; Conserved elements
    Source: Springer Online Journal Archives 1860-2000
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    Notes: Summary Computer analysis has previously revealed the presence of a 12-nucleotide common sequence element (AACATC CA TG T A G CA; HOMOL1) in the upstream regions of several yeast ribosomal protein genes. By extending the sequence analysis of the 5′-flanking regions of a number of other ribosomal protein genes (including those encoding S10-1, S10-2, S33 and L16-2) we could establish that HOMOL1 occurs upstream of most but not all yeast ribosomal protein genes. Apart from HOMOL1 an additional conserved sequence (ACCCATACATT A T ; RPG-box) was detected in front of nearly all yeast ribosomal protein genes, although in some cases it is present in the opposite orientation in the other strand. There seems to be no correlation between the occurrence of one box and that of the other. However when both boxes are present the RPG-box is always located 3′ to the HOMOL1-sequence mostly at a distance of only a few nucleotides. A further one-to-one comparison of the upstream regions of several yeast ribosomal protein genes revealed extensive additional sequence homologies that are suggested to be involved in the coordinate control of ribosomal protein gene expression in yeast.
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  • 135
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    Current genetics 6 (1982), S. 179-188 
    ISSN: 1432-0983
    Keywords: Yeast ; Mitochondria ; var1 gene
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Several mit mutants mapping within or near the var1 determinant region have been characterized genetically and biochemically. These mutants were isolated using a new enrichment protocol which simplifies the isolation and identification of rare respiration-deficient mutants of yeast. Two of the mutants, PZ200L and PZ206, map in genome segments which flank the known varl gene reading frame; nevertheless, both belong to the same complementation group, apparently that of the varl gene. A third mutant, PZ200R is closely linked to one of the varl allelic determinants now known to be a short insertion within the gene. All three var1 mutants exhibit decreased levels of mitochondrial protein synthesis and negligible activity of the respiratory enzyme complexes. Another cluster of mutants belonging to a separate complementation group from that defined by PZ200L and PZ206 was also mapped and it contains mutants in the nearby serine tRNA gene. The isolation of these mutants in the varl region shows that the varl locus contains information essential for the maintenance of respiration-competent mitochondria. Because these mutants affect mitochondrial protein synthesis, their existence supports the previous hypothesis that the varl protein is an integral component of mitochondrial ribosomes. Furthermore, the mutant sites are present in a DNA sequence that is highly, rich in A+T residues that also contains a gene. Since approximately 50% of the yeast mitochondrial genome is similarly rich in A+T and since most of those regions have not yet been sequenced it is quite possible that other A+T-rich genes may exist.
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  • 136
    ISSN: 1432-0983
    Keywords: Nitrosoguanidine ; Comutation ; Yeast ; Chromosome replication pattern
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Contrary to what happens in bacteria, mutations induced by nitrosoguanidine in yeast are not accompanied by an excess of mutations in nearby genes. We have investigated nitrosoguanidine mutagenesis in three regions of the yeast genome: the contiguous DNA segments HIS4A, HIS4B and HIS4C, located on chromosome III; ADE1 and CDC15 separated by about 3 map units on chromosome I; and CAN1, some 50 map units away from the centromere on chromosome V. Revertants at HIS4C never suffered mutations at HIS4A or HIS4B. Reversion at CDC15 did not affect the frequency of mutation at ADE1. No tsm mutations, leading to thermonsensitivity, were found in the immediate vicinity of the locus CAN1 after selecting for canavanine resistant mutants. However, as expected from nitrosoguanidine mutagenesis of replication points and the fixed pattern of chromosome replication, the induced tsm mutations seem not to map randomly over the yeast genome; in fact, two out of the three groups of such tsm mutations studied are located in the same chromosome arm as CAN1, indicating that these two regions are replicated at the same time as CAN1. Replication synchrony is less than perfect, since the tsm mutations of each group affect many different genes.
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  • 137
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    Keywords: Canavanine ; Yeast ; Plasmids
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    Topics: Biology
    Notes: Summary We have found that the application of the amino acid analog canavanine to a culture of yeast cells transformed with chimeric plasmids based on the yeast 2-µm DNA plasmid increases the percentage of cells which have lost the transforming plasmid. This effect is found whether the plasmid carries the CAN1 sensitive allele and the yeast strain carries a can1 mutation confering resistance, or the plasmid contains no CAN1 allele and the yeast strain carries the wild-type CAN1 sensitive allele. Canavanine exerts this effect on yeast strains transformed with chimeric plasmids containing either a portion or the entire 2-µm DNA plasmid, yet canavanine does not appear to effect the maintenance of the native 2-µm DNA plasmid complement within the cell. The effect of canavanine on strains transformed with chimeric plasmids is the same whether or not the yeast strain also contains native 2-µm plasmid DNA. Neither the amino acid analog ethionine, the protein synthesis inhibitor cycloheximide, nor the DNA replication inhibitor hydroxyurea exhibit this effect. Some of the experimental results suggest that canavanine may be a curing agent rather than an agent which selects for spontaneous plasmid loss.
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  • 138
    ISSN: 1432-0983
    Keywords: Yeast ; Polyamines ; Termination ; In vitro Translation
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    Notes: Summary The effects of polyamines (spermidine and putrescine) on yeast suppressor tRNA-mediated readthrough of amber and UGA termination codons, in a homologous cell-free system, was examined. The efficiency of readthrough in a [psi+] lysate, mediated by exogenous suppressor tRNA, was significantly increased by polyamines as was the efficiency of endogenous UGA readthrough. The addition of polyamines, in the absence of exogenous suppressor tRNA, did not induce amber or ochre readthrough, nor could polyamines restore efficient termination readthrough in [psi−] lysates. It is concluded that polyamines interact with tRNA to increase the strength and specificity of the codon: anticodon interaction.
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  • 139
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    Current genetics 8 (1984), S. 265-270 
    ISSN: 1432-0983
    Keywords: DHF Reductase ; Cloning ; Trimethoprim ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The dihydrofolate reductase gene of Saccharomyces cerevisiae has been isolated by selection of trimethoprim resistant Escherichia coli transformed with a gene bank of yeast DNA in plasmid pBR322. From 9.2 kilobase pair BamHI DNA fragment this gene has been localized to a 1.76 kb fragment, the restriction map of which appears different from those reported for the E. coli and the mouse dihydrofolate reductase genes. The enzyme encoded by the chimeric plasmid was established as yeast dihydrofolate reductase by its sensitivity to antifolates in vivo through growth studies and in vitro by enzyme assay. Since, the expression of this gene occurs independent of its orientation within the chimeric plasmid, the 1.76 kb fragment may contain functional regulatory sequences in addition to the structural sequences for yeast dihydrofolate reductase.
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  • 140
    ISSN: 1432-0983
    Keywords: Yeast ; Ribosomal protein gene ; Sequence analysis ; Northern blot
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    Notes: Summary Sequence analysis revealed that a gene coding for yeast ribosomal protein L34 comprises an amino acid coding region of 339 nucleotides which is interrupted by an intron after the 19th codon. Like for other yeast ribosomal protein genes analyzed thus far a strong codon bias was observed. The flanking and intervening sequences of this gene encoding L34 show several elements that are conserved in a number of split ribosomal protein genes in yeast. Northern blot analysis using an intron-specific probe demonstrated that the sequenced gene copy coding for L34 is transcribed in vivo.
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  • 141
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    Current genetics 9 (1985), S. 529-532 
    ISSN: 1432-0983
    Keywords: Yeast ; UV-inducible proteins ; rad mutants
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    Notes: Summary Two UV-inducible proteins have been detected in the yeast, Saccharomyces cerevisiae. The proteins have molecular weights of 78,000 Daltons and 23,000 Daltons. This induction is specific for UV-irradiation as exposure to X-rays, mitomycin C and heat shock does not result in the synthesis of the proteins. The larger (78 kD) protein is induced in various rad strains and in a π° cir° strain. Attempts are being made to isolate the genes coding for these inducible proteins.
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  • 142
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    Current genetics 9 (1985), S. 653-660 
    ISSN: 1432-0983
    Keywords: Gene expression ; Yeast ; Transcription
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Efficient expression of theEscherichia coli ZeuB (ß-isopropylmalate dehydrogenase) gene occured in yeast after in vitro DNase digestion and religation of plasmid bound ZeuB and the yeastIIIS3 DNA which placed the 5′ end of the yeastHIS3 gene immediately adjacent to the coding region of theE. coli leuB gene. Two structurally distinct classes of gene fusions were constructed, each involved portions of the yeastHIS3 gene which contributed DNA sequences responsible forleuB expression in yeast. The first class involved fusion of theHIS3 coding region to bacterial DNA resulting in the production of a fusion protein with ß-isopropylmalate dehydrogenase activity. The second class consisted of bacterial DNA, including theleuB coding region, fused to theHIS3 promotor region with the absence of any portion of theIIIS3 coding region. In both constructions theIIIS3 promotor region is required for transcription, however, translation of the class two fusion is initiated at a bacterial DNA coded AUG, and the 5′ end of the mRNA coded by theleuB gene mapped predominantly at bacterial DNA sequences. The DNA sequence responsible for the 5′ end of theHIS3 mRNAs remain in the class two gene fusions but this did not preclude the initiation of transcription at bacterial DNA sequences. The pattern of mRNA initiation at bacterial DNA suggests that DNA sequences at, or adjacent to, the site of transcription initiation are involved in the determination of the sites of initiation, and perhaps the frequency at which initiation occurs.
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  • 143
    ISSN: 1432-0983
    Keywords: Yeast ; Cycloheximide ; Ribosomal mutations
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    Notes: Summary At least four different mutations at the cyh2 locus (rp1X; gene product: YL24) of Saccharomyces cerevisiae confer cycloheximide resistance. The mutant YL24 proteins are either more basic (high-level resistant phenotype), more acidic (low-level resistant phenotype), or unchanged in their electrophoretic mobility (both low-and high-level resistant phenotypes). None of the mutations at other loci seem to induce high-level resistance to cycloheximide.
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  • 144
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    Current genetics 5 (1982), S. 171-180 
    ISSN: 1432-0983
    Keywords: Yeast ; Mitochondrial genes ; Vegetative segregation
    Source: Springer Online Journal Archives 1860-2000
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    Notes: Summary A three-factor cross of Saccharomyces cerevisiae involving the cap1, ery1, and oli1 loci was done, with partial pedigree analyses of 117 zygotes. First, second, and third buds were removed and the genotypes of their diploid progeny determined, along with those of the residual zygote mother cell. Results were analyzed in terms of frequencies of individual alleles and of recombinant genotypes in the dividing cells. There is a gradual increase in the frequency of homoplasmic cells and in gene frequency variance during these three generations, as would result from stochastic partitioning of mtDNA molecules between mother and bud, probably coupled with random drift of gene frequencies in interphase cells. These phenomena are more pronounced for buds than for mothers, suggesting that buds receive a smaller sample of molecules. End buds are more likely to be homoplasmic and have a lower frequency of recombinant genotypes than do central buds; an end bud is particularly enriched in alleles contributed by the parent that formed that end of the zygote. Zygotes with first central buds produce clones with a higher recombination frequency than do those with first end buds. These results confirm previous studies and suggest that mixing of parental genotypes occurs first in the center of the zygote. If segregation were strictly random, the number of segregating units would have to be much smaller than the number of mtDNA molecules in the zygote. On the other hand, there is no evidence for a region of the molecule (“attachment point”) which segregates deterministically.
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  • 145
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    Current genetics 6 (1982), S. 29-30 
    ISSN: 1432-0983
    Keywords: Yeast ; Plasmid ; Repair ; Ligase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We show that the DNA ligase encoded or controlled by the cdc9 gene in Saccharomyces cerevisiae is required for replication of plasmid DNA but that excision repair of pyrimidine dimers in plasmid DNA can be completed in its absence.
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  • 146
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    Current genetics 6 (1982), S. 195-201 
    ISSN: 1432-0983
    Keywords: Benomyl resistance ; Yeast ; Mitosis ; Cell cycle mutants
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We have isolated 150 benomyl resistant mutants of the fission yeast Schizosaccharomyces pombe. Seven of these mutants were found to be cold sensitive for mitosis. These mutants were the subject of physiological, cytological and genetical characterisation. Growth and division of the seven mutants were similar to the wild type strain at 35 °C. After shift from the permissive (35 °C) to the restrictive temperature (20 °C) the mutants became blocked in mitosis whilst cellular growth continued. Consequently, elongate cells were formed. Six of the seven benomyl resistant mutants became blocked in mitosis at 20 °C with a single aberrant nucleus. In every case the benomyl resistant and cold sensitive phenotype was due to a mutation in a single nuclear gene. These mutants were found to comprise a single genetic linkage group (ben4) and were unlinked to existing TBZ/MBC resistant mutants of S. pombe. Whilst no cross resistance was found in our mutants to TBZ, six of the seven mutants were super sensitive to the spindle poison CIPC. We believe that the phenotype exhibited by these mutants is consistent with a defective tubulin subunit.
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  • 147
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    Current genetics 7 (1983), S. 79-84 
    ISSN: 1432-0983
    Keywords: ars sequences ; Ustilago ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary U. maydis chromosomal DNA sequences which promote the autonomous replication of plasmid YIp5 in S. cerevisiae YNN27 have been isolated and three of them characterised in some detail. Their properties are idential to yeast ars sequences in that plasmids containing them are maintained extrachromosomally as circular double-stranded DNA molecules, are mitotically unstable in yeast transformants and transform yeast at high frequencies. There is no sequence homology between the three U. maydis sequences and they are not reiterated in the U. maydis genome.
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  • 148
    ISSN: 1432-0983
    Keywords: Yeast ; Recombination ; Conversion ; Restoration
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    Notes: Summary Heteroduplex repair models of recombination predict that restoration of the parental genotype from heteroduplex will have the same frequency as conversion to the genotype of the homologue. We have reported previously (Savage and Hastings 1981) that the proportion of intragenic reciprocal events in the his1 locus of yeast is too low for this expectation. In this study, two classes of recombination event involving longer lengths of his1 are compared in a series of crosses using a constant right-hand marker. This comparison gives a value for conversion: restoration for the left-hand markers which is independant of the method used before. The values obtained by the two methods are significantly correlated, confirming that there is a disparity in the ratio of conversion to restoration, and that this disparity is seen as a deficiency of restoration for five alleles of his1.
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    Current genetics 8 (1984), S. 429-437 
    ISSN: 1432-0983
    Keywords: Yeast ; Heat-shock response ; Mitochondrial inhibition ; 2D electrophoresis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Conventional and two dimensional (2D) electrophoresis on ultrathin horizontal slab gels shows that heat shock proteins are synthesized and heat stroke proteins are curtailed after the transfer of Saccharomyces cerevisiae strain Z270 from 23 °C to 37 °C. Upon addition of the mitochondrial translation inhibitor erythromycin to cell cultures which incorporated labelled methionine at 23 °C, and after the transfer to 37 °C, we have shown that: a) in extracts of cells labelled at 23 °C, translational products sensitive to erythromycin could be observed on 2D gels; the synthesis of some of these proteins was enhanced, whereas the synthesis of some others declined when the labelling was carried out 20 min after the transfer to 37 °C; b) there are heat shock proteins whose induction at 37 °C was prevented by erythromycin; c) labelling of a number of proteins became weaker at 37 °C, but not at 23 °C, when this was done in the presence of erythromycin; d) two proteins were detectable only in samples labelled at 37 °C in the presence of erythromycin.
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  • 150
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    Keywords: Yeast ; Mitochondrial ochre mutations ; Specificity of suppressors ; Mitoribosomal misreading ; Intron-encoded maturases
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We describe studies on the action spectra of the mitochondrial suppressor mim3-1 and the three alleles of nuclear suppressor nam3. Their specificity of action was tested on 516 mit − mutations located in different mitochondrial genes. The degree of suppression was quantified by the extent of cytochrome oxidase and cytochrome b synthesis. We show that the four suppressors are allele-specific gene-nonspecific informational suppressors. They would act by changing the structure of the small mitoribosomal subunit which would decrease fidelity of translation enabling misreading of some but not all ochre codons. The implications of the results on the role of intron encoded maturases are discussed.
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  • 151
    ISSN: 1432-0983
    Keywords: Yeast ; Mitochondrial ochre mutations ; Nuclear informational suppressors ; Mitochondrial informational suppressors
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    Topics: Biology
    Notes: Summary A systematic search for suppressors of mutations which cause a deficiency in the splicing of mitochondrial RNA has been undertaken. These splicing mutations were localized in the mRNA-maturase coding sequence of the second intron of the cob-boxgene, i.e. in the box3locus. A total of 953 revertants (mostly spontaneous in origin) were isolated and their genetic nature (nuclear vs. mitochondrial) and phenotype characterized. Most revertants were mitochondrially determined and displayed a wild-type phenotype. A mitochondrial suppressor unlinked with the box3 −target mutation was uncovered among the revertants displaying a pseudo-wild phenotype: out of 26 revertants analyzed, derived from 7 different box3− mutants only one such suppressor mutation mim3-1 was found. It was localized by rho− deletion mapping in the region between the oxi2 and oxi3 gene, within (or in the vicinity) the gene specifying the 15S ribosomal RNA. Nuclear suppressors were isolated from seven different box3 −mutants. All were recessive and had a pseudo-wild phenotype. Three such suppressors nam3-1, nam3-2 and nam3-3 were investigated more extensively. Tetrad analysis has shown that they are alleles of the same nuclear locus NAM3 and mitotic analysis has shown that they do not segregate mitotically.
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    Current genetics 9 (1985), S. 147-155 
    ISSN: 1432-0983
    Keywords: Killer ; Yeast ; Linear plasmid ; Sequence
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Some strains of the yeast Kluyveromyces lactis contain a pair of linear DNA plasmids, k1 and k2, 8.8 and 13.8 kilobase pairs long, respectively. Simultaneous presence of the two plasmids confer a killer phenotype on the cell by producing a toxin which blocks the growth of sensitive yeast species. Previous genetic studies have suggested that the toxin protein is coded by the k1 plasmid. We have now determined the total nucleotide sequence of k1 DNA. The genome is 8,874 base pairs in length. It contains four protein-coding reading frames, three transcribed from one strand and the fourth transcribed from the complementary strand and has terminal inverted repeats of 202 base pairs. Nuclease S1 mapping confirmed this arrangement and showed that these genes are transcribed. The terminal repeats and the four genes form an extremely compact genome, with some overlapping of genes. All four genes use highly biased codons, 86% of them having A or T at the wobble position, reminiscent of yeast mitochondrial genes. Three genes share a very similar 5′ leader sequence. The nature of gene products is discussed in the light of what is known of the excreted toxin protein.
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    Current genetics 9 (1985), S. 179-181 
    ISSN: 1432-0983
    Keywords: Blasticidin S ; Yeast ; Resistant mutants
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Blasticidin S-resistant mutants of S. cerevisiae were isolated and characterized. Resistant mutations were found to fall into two complementation groups. A single recessive nuclear gene was responsible for each group, donated as bls1 and bls2, respectively. A gene bls1 was linked to an ilv3 gene located on the right arm of chromosome X. The resistant phenotypes from both genes were not associated with ribosomes known to be target sites of Blasticidin S, when analyzed by poly(U)-directed polyphenylalanine synthesis. The resistant mechanisms of the mutations are discussed in this paper.
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  • 154
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    Current genetics 9 (1985), S. 427-433 
    ISSN: 1432-0983
    Keywords: Yeast ; Mitochondrial mutations ; Informational suppressors
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Ten nuclear suppressors (nam mutations) of the mitochondrial oxi1-V25 ochre mutation are characterized. They restore to some extent the functional form of cytochrome oxidase, as judged by the results of growth tests, cytochrome spectra, cytochrome oxidase activities, and electrophoresis of the products of mitochondrial translation. The nam mutants can suppress some mit − mutations mapping in four mitochondrial genes. They act on a number of chain-terminating mit − mutations. When grown on glycerol medium some double mutants nam x-V25 show an increased sensitivity to paromomycin, while the growth of others is stimulated by the drug. The nam mutants are probably omnipotent suppressors resulting from mutations in nuclear gene(s) specifying mitoribosomal protein(s).
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    Current genetics 9 (1985), S. 553-560 
    ISSN: 1432-0983
    Keywords: Yeast ; Ty element ; Recombination ; Gene conversion ; Regulation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A haploid yeast mutant carrying a reciprocal translocation was analyzed. Cloning and comparison of sequences involved in the translocation event in wildtype and mutant revealed that the crossover between non-homologous chromosomes has occured within Ty sequences. By DNA sequence analysis it could be demonstrated that the reciprocal recombination event is accompanied by a short segment of non-reciprocal exchange (gene conversion) in the immediate vicinity of the crossover. Analysis of the translocation mutant and revertant isolates also indicated that the regulatory effect of Ty elements on adjacent genes can be modified by discrete changes within a Ty element.
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  • 156
    ISSN: 1432-0983
    Keywords: Mitochondria ; Mutation ; Yeast ; Selection ; Random drift
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Haploid yeast cells have about 50 copies of the mitochondrial genome, and a mutational event is unlikely to affect more than one of these at a time. This raises the question of how such cells, or their progeny, become fixed (homoplasmic) for the mutant alele. We have tested the roles of six hypothetical mechanisms in producing erythromycin-resistant mutant cells: (i) random partitioning of mitochondrial genomes at cell division; (ii) intracellular selection for mtDNA molecules of one genotype; (iii) intracellular random drift of mitochondrial allele frequencies; (iv) intercellular selection for cells of a particular mitochondrial genotype; (v) induction of mitochondrial gene mutations by the antibiotic used to select mutants; and (vi) reduction in the number of mitochondrial genomes per cell by the antibiotic. Our experiments indicate that intracellular selection plays the major role in producing erythromycin-resistant mutant cells in the presence of the antibiotic. In the absence of the antibiotic, the combined effects of random drift and random partitioning are most important in determining the fate of new mutations, most of which are lost rather than fixed. Our experiments provide no evidence for mutation induction or ploidy reduction by erythromycin.
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  • 157
    ISSN: 1432-072X
    Keywords: Mating tube ; Microtubule ; Tremella ; Ultrastructure ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Ultrastructure of the mating tube formed in yeast haplont of the heterobasidiomycete Tremella mesenterica was studied by electron microscopy. Cell wall of the mating tube emerged as evagination of the inner layers, rupturing outer layers of the mother cell wall. Comparison with budding cells suggested that the tube emergence place at bud scar and the process of tube emergence was the same as that of bud emergence. Electron transparent vesicles of 0.1 μm diameter were scattered in the cytoplasm of the mating tube. Nucleus-associated organelle was located at one side of the nuclear envelope which extended towards the mating tube. A few microtubules were detected in the mating tube, but their association with a nucleus was not clear. The cytoplasmic structure of the mating tube was discussed in comparison with that of hyphae of the filamentous fungi.
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  • 158
    ISSN: 1432-072X
    Keywords: Pyrophosphate ; Polymerie acid-soluble poly-phosphates ; Budding process ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In the cells of hybrid yeast strain Saccharomyces N.C.Y.C. 644 SU3 (Karlsberg collection), a large amount of pyrophosphate (30–300 μmol/g of dry weight) accumulates whatever the aeration conditions and the content of glucose in the medium. The content of pyrophosphate is 10–1000 times higher than that of ATP. At the early and mid-exponential growth phases two maxima of pyrophosphate accumulation are observable. The periods of maximal pyrophosphate accumulation in yeast coincide with those of the minimal content of polymeric acid-soluble polyphosphates and intense budding. In the light of the data obtained, the question is discussed as to the relationship between the metabolism of pyrophosphates and acid-soluble polyphosphates in yeast.
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  • 159
    ISSN: 1432-072X
    Keywords: Germination ; Glycogen ; Outgrowth ; Schizosaccharomyces pombe ; Spore ; Trehatase ; Trehalose ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Quantitative changes in various carbohydrates of Schizosaccharomyces, pombe spores during germination and outgrowth were studied. Trehalose decreased rapidly, shortly after onset of germination, while glycogen remained constant throughout germination and outgrowth. Alkali-insoluble carbohydrates decreased after the lag period of about 40 min. The content of alkali-soluble carbohydrates was constant during germination, but increased remarkably in parallel with germtube formation. The mechanism of rapid degradation of trehalose during germination was also studied. The activity of trehalase (EC 3.2.1.28) was detected only in the cell wall fraction of isolated spores. Trehalase activity in the cell wall fraction was not enhanced during germination. Trehalose was not found in the isolated spore walls, but in the soluble fraction. These facts strongly suggested that trehalose, and trehalase were spatially separated in dormant spores and that trehalase became accessible to trehalose upon germination.
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  • 160
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    Archives of microbiology 129 (1981), S. 47-48 
    ISSN: 1432-072X
    Keywords: Adenylate energy charge ; Phosphate ; Saccharomyces ; Sporulation ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The induction of sporulation in yeast is generally accompanied by a sharp increase in energy metabolism which is evidenced by a rise of the adenylate energy charge by that time. The energy charge can be held at a low level by limitation of the phosphate supply in the growth medium. Ascus formation remains unaffected by this treatment. This suggests that the rise in ATP production normally encountered during early sporulation is not essential for the initiation of sporulation.
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  • 161
    ISSN: 1432-072X
    Keywords: Ions ; Concentration ; Regulation ; Cytoplasm ; Vacuole ; Yeast ; Saccharomyces carlsbergensis
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    Topics: Biology
    Notes: Abstract Intracellular distributions of K+, Mg2+ and orthophosphate under various conditions of cultivation or incubation of the yeast Saccharomyces carlsbergensis were studied by differential extraction of ion pools. The decisive role of vacuolar compartmentation of ions in regulation of K+, Mg2+ and orthophosphate levels in the yeast cytoplasm was shown. The content of intracellular K+ and Mg2+ in yeast increased or decreased primarily depending on the increase or decrease in the vacuolar ion pool. The levels of K+ and Mg2+ in the cytoplasm were practically unchanged. Vacuoles were involved in regulation of Mn2+ concentration in the cytoplasm of the yeast S. carlsbergensis accumulating this ion in the presence of glucose. Alongside the vacuolar compartmentation, the chemical compartmentation, i. e. formation of bound Mg2+, Mn2+ and K+ was, evidently, also involved in the control of ion levels in the cytoplasm. The orthophosphate level in the yeast cytoplasm was regulated by its accumulation in vacuoles and biosynthesis of inorganic polyphosphates in these organelles. The biosynthesis of low-molecular weight polyphosphates occurred parallel to the accumulation of Mg2+ or Mn2+ in vacuoles, thus confirming the availability of the other mechanism for the transport of these ions through the tonoplast differing from the transport mechanism through the plasmalemma.
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  • 162
    ISSN: 1432-072X
    Keywords: Yeast ; Phosphate uptake ; Phosphate-binding protein ; Anti-phosphate-binding protein antibodies
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    Topics: Biology
    Notes: Abstract The purification of a phosphate-binding protein (PiBP2) by immunoadsorption is described. The entire anti phosphate-binding protein 2 antibodies as well as the Fab fragments obtained from these antibodies inhibit Pi uptake by whole cells. The inhibition is a mixed type of inhibition (V m and K m are affected). These results should be regarded as a possible involvement of phosphate-binding protein 2 in Pi uptake. The binding of 125I-labelled fragments prepared from anti phosphate-binding protein 2 antibodies to whole cells, to shocked cells and to protoplasts has been investigated. The results confirm the release of phosphate-binding protein by osmotic shock and during protoplast formation. From these findings, a cell-wall localisation, near the cell surface of the phosphate-binding protein should be proposed.
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  • 163
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    Archives of microbiology 137 (1984), S. 357-361 
    ISSN: 1432-072X
    Keywords: Yeast ; Saccharomyces cerevisiae ; Killer toxin ; Extracellular glycoprotein
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    Topics: Biology
    Notes: Abstract A total of 13 killer toxin producing strains belonging to the genera Saccharomyces, Candida and Pichia were tested against each other and against a sensitive yeast strain. Based on the activity of the toxins 4 different toxins of Saccharomyces cerevisiae, 2 different toxins of Pichia and one toxin of Candida were recognized. The culture filtrate of Pichia and Candida showed a much smaller activity than the strains of Saccharomyces. Extracellular killer toxins of 3 types of Saccharomyces were concentrated and partially purified. The pH optimum and the isoelectric point were determined. The killer toxins of S. cerevisiae strain NCYC 738, strain 399 and strain 28 were glycoproteins and had a molecular weight of Mr=16,000. The amino acid composition of the toxin type K2 of S. cerevisiae strain 399 was determined and compared with the composition of two other toxins.
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  • 164
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    Archives of microbiology 138 (1984), S. 183-186 
    ISSN: 1432-072X
    Keywords: Yeast ; Nitrogen assimilation ; Nitrate reductase ; Nitrite reductase ; Candida utilis ; Food yeast ; Nitrate reduction ; Nitrogenous oxide
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Assimilation of nitrate and various other inorganic nitrogen compounds by different yeasts was investigated. Nitrate, nitrite, hydroxylamine, hydrazine, ammonium sulphate, urea and L-asparagine were tested as sole sources of nitrogen for the growth of Candida albicans, C. pelliculosa, Debaryomyces hansenii, Saccharomyces cerevisiae, C. tropicalis, and C. utilis. Ammonium sulphate and L-asparagine supported the growth of all the yeasts tested except D. hansenii while hydroxylamine and hydrazine failed to support the growth of any. Nitrate and nitrite were assimilated only by C. utilis. Nitrate utilization by C. utilis was also accompanied by the enzymatic activities of NAD(P)H: nitrate oxidoreductase (EC 1.6.6.2) and NAD(P)H: nitrite oxidoreductase (EC 1.6.6.4), but not reduced methyl viologen-or FAD-nitrate oxidoreductases (EC 1.7.99.4). It is demonstrated here that nitrate and nitrite reductase activities are responsible for the ability of C. utilis to assimilate primary nitrogen.
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  • 165
    ISSN: 1432-072X
    Keywords: β-Glucosidase ; Yeast ; Genetic engineering ; Biosynthesis regulation
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    Topics: Biology
    Notes: Abstract The biosynthesis of the β-glucosidase enzyme was studied in a transformed yeast obtained by cloning in Saccharomyces cerevisiae the structural gene coding for β-glucosidase in Kluyveromyces fragilis. The enzyme biosynthesis was found to be non-adaptative, and repressed by glucose. These features are similar to those observed in K. fragilis. β-Glucosidase activity in the transformed yeast was much higher than in K. fragilis. We attempted to ferment cellobiose with the transformed yeast: practically no cellobiose was consumed, growth and ethanol production were negligible. Warburg experiments showed that cellobiose fermentation did not occur when the respiratory chain was not functioning.
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  • 166
    ISSN: 1432-072X
    Keywords: Extracellular proteins ; Surface fibrils ; Algae-fungi-Chrysochromulina ; Immunocytochemistry ; Agglutination ; Fimoriae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract An extensive network of extracellular fibrils was revealed by negative staining in the greenish gold algal flagellate, Chrysochromulina breviturrita. These fibrils were of uniform diameter (4–5 nm), sometimes exceeding 5 μm in length. In addition there were short, narrower fibrils (2–3 nm) on the surface of the flagella. Six protein bands were isolated from spent culture medium by SDS-PAGE and one of 80,000 Da was found to polymerize after dialysis into 4–5 nm fibrils identical to those found on the cell surface. Two other proteins of 58,000 Da and 65,000 Da also formed 4–5 nm fibrils but these were either rare or of a shorter length and different appearance. An antiserum directed against the surface 7 nm fibrils (fimbriae) of fungi agglutinated cells of C. breviturrita and some other Prymnesiophyceae and Chrysophyceae, but did not agglutinate cells of algal species in other groups. Immunofluorescence and protein A gold labelling confirmed that antigens related to fungal fimbriae were present on the surface of cells of C. breviturrita. Only the 80,000 and 58,000 Da proteins labelled heavily following protein A gold labelling. Some individual 4–5 nm fibrils labelled with gold were observed in the material prepared from the 80,000 Da band. These results therefore establish that C. breviturrita produces a surface network of fibrils that are serologically related to the fimbriae of fungi, and suggest a previously unrecognized relationship between members of the Prymnesiophyceae, Chrysophyceae and fungal groups.
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  • 167
    ISSN: 1432-072X
    Keywords: Hansenula polymorpha ; Yeast ; Peroxisomes ; Proton-translocating ATPase ; Cell fractionation ; Fluorescence quenching studies ; Cytochemistry
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The association of an ATPase with the yeast peroxisomal membrane was established by both biochemical and cytochemical procedures. Peroxisomes were purified from protoplast homogenates of the methanol-grown yeast Hansenula polymorpha by differential and sucrose gradient centrifugation. Biochemical analysis revealed that ATPase activity was associated with the peroxisomal peak fractions which were identified on the basis of alcohol oxidase and catalase activity. The properties of this ATPase closely resembled those of the mitochondrial ATPase of this yeast. The enzyme was Mg2+-dependent, had a pH optimum of approximately 8.5 and was sensitive to N,N′-dicyclohexylcarbodiimide (DCCD), oligomycin and azide, but not to vanadate. A major difference was the apparent K m for ATP which was 4–6 mM for the peroxisomal ATPase compared to 0.6–0.9 mM for the mitochondrial enzyme. Cytochemical experiments indicated that the peroxisomal ATPase was associated with the membranes surrounding these organelles. After incubations with CeCl3 and ATP specific reaction products were localized on the peroxisomal membrane, both when unfixed isolated peroxisomes or formaldehyde-fixed protoplasts were used. This staining was strictly ATP-dependent; in controls performed i) in the absence of substrate, ii) in the presence of glycerol 2-phosphate instead of ATP, or iii) in the presence of DCCD, staining was invariably absent. Similar staining patterns were observed in subcellular fractions and protoplasts of Candida utilis and Trichosporon cutaneum X4, grown in the presence of ethanol/ethylamine or ethylamine, respectively.
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  • 168
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    Archives of microbiology 147 (1987), S. 105-108 
    ISSN: 1432-072X
    Keywords: Ozone ; Yeast ; Saccharomyces cerevisiae ; ATP ; Nucleotides ; Permeability ; Cytosolic enzymes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Treatment of a yeast suspension with ozone inactivates a number of cytosolic enzymes. Among 15 studied, the most drastic inactivation was found for glyceraldehyde-3-phosphate dehydrogenase and to lesser extents: NAD-glutamate dehydrogenase, pyruvate decarboxylase, phosphofructokinase-1 and NAD-alcohol dehydrogenase. Ozone treatment also effects the quantity of ATP and of other nucleoside triphosphates, reducing to about 50% of the initial value. The ATP missing in the cells appears in the medium. NAD and protein also accumulate in the medium suggesting that the yeast cells have been permeabilized. Permeabilization of the yeast cells by treatment with ozone preceeds the inactivation of glyceraldehyde-3-phosphate dehydrogenase and other cytosolic enzymes.
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  • 169
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    Keywords: Methanobacterium thermoautotrophicum ; Methyl-CoM reductase ; Immunocytochemistry ; Colloidal gold ; Energy conservation
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    Topics: Biology
    Notes: Abstract Cells of Methanobacterium thermoautotrophicum were fixed with glutaraldehyde, sectioned and labeled with antibodies against the β subunit of component C (=methyl-CoM reductase) of methyl-CoM reductase system and with colloidal gold-labeled protein A. It was found that the gold particles were located predominantly in the vicinity of the cytoplasmic membrane, when the cells were grown under conditions where methyl-CoM reductase was not overproduced. This finding confirms the recent data obtained with Methanococcus voltae showing via the same immunocytochemical localization technique that in this organism methyl-CoM reductase is membrane associated.
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  • 170
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    Archives of microbiology 147 (1987), S. 235-239 
    ISSN: 1432-072X
    Keywords: Chloroquine ; Yeast ; Fructose-1,6-bisphosphatase ; Phosphorylation ; Protein kinase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The rapid phase of fructose-1,6-bisphosphatase (FBPase) inactivation following glucose addition to starved yeast cells [reported previously] is inhibited on addition of 10 mM chloroquine (CQ) at about pH 8. This inhibition of inactivation was shown to be due to the prevention of phosphorylation of the enzyme. CQ was also found to inhibit general protein phosphorylation in the yeast cells. Glycolysis, as observed by changes in intracellular glucose-6-phosphate and extracellular glucose and ethanol concentrations, was shown to be significantly inhibited in cells treated with CQ. Similarly, a decrease in ATP concentrations was observed. However, during the early stages of phosphorylation of FBPase, levels of ATP were similar in cells containing CQ as in those without CQ. Thus, decrease in ATP levels is not thought to be significantly responsible for the inhibition of protein phosphorylation. However, the phosphorylating activity of cyclic AMP-dependent protein kinases is inhibited in vitro by relatively low concentrations of CQ. Thus, prevention of protein phosphorylation by CQ is believed to be due to inhibition of protein kinases in yeast cells.
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  • 171
    ISSN: 1432-072X
    Keywords: Saccharomyces cerevisiae ; Yeast ; Phospholipase B ; Lysophospholipase ; Enzyme inhibition ; AMP ; Unesterified fatty acids
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Divalent cations activate the lysophospholipase and transacylase reactions catalyzed by the same enzymes in the yeast Saccharomyces cerevisiae. The activation was observed at neutral pH, but not at the pH optimum of lysophospholipase/transacylase, near 3.5. Adenine nucleotides, especially AMP and ADP, are strong inhibitors of the same group of enzymes. Half maximal inhibition by AMP was found at a concentration of about 20 μM. The inhibition by nucleotides in low concentrations is enhanced by divalent cations.
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  • 172
    ISSN: 1432-072X
    Keywords: Acidaminococcus fermentans ; Glutamate fermentation ; Electron microscopy ; Immunocytochemistry ; Post-embedding labelling ; Antibody-gold complexes ; Protein A-gold complexes
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    Topics: Biology
    Notes: Abstract We have investigated the in situ location of glutaconyl-CoA decarboxylase and 2-htdroxyglutaryl-CoA dehydratase in Acidaminococcus fermentans using the antibody-gold and protein A-gold techniques carried out as a post-embedding immunoelectron microscopic procedure. Polyclonal antisera were raised in rabbits against homogeneous fractions of the enzymes. Anaerobically grown cells of A. fermentans of the late exponential growth phase were fixed with 0.2% glutaraldehyde and 0.3% formaldehyde (final concentrations) in the growth medium. Dehydration of the cells was achieved with methanol. The cells were embedded in the low temperature embedding resin Lowicryl K4M. The markers indicative for antigenic sites of the two enzymes unequivocally demonstrate that the sodium pump glutaconyl-CoA decarboxylase is located at the cell periphery being a membrane-bound enzyme as expected whereas 2-hydroxyglutaryl-CoA dehydratase is a soluble cytoplasmic enzyme.
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  • 173
    ISSN: 1432-072X
    Keywords: Peroxisome ; Methanol ; Methylamine ; Yeast ; Hansenula polymorpha ; Alcohol oxidase ; Amino oxidase ; Catalase ; Catabolite inactivation ; Turnover ; Cytochemical localization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Growth of Hansenula polymorpha in shake flasks and chemostat cultures in the presence of methanol as the sole source of carbon and methylamine as the sole source of nitrogen was associated with the development of peroxisomes in the cells. The organelles were involved in the concurrent oxidation of these two compounds, since they contained both alcohol oxidase and amine oxidase, which are key enzymes in methanol and methylamine metabolism, respectively. In addition catalase was present. Peroxisomes with a completely crystalline substructure were observed in methanol-limited chemostat-grown cells. Amine oxidase probably formed an integral part of these crystalloids, whereas catalase was present in a freely diffusable form. Transfer of cells, grown in a methanol-limited chemostat in the presence of methylamine into glucose/ammonium sulphate media resulted in the loss of both alcohol oxidase and amine oxidase activity from the cells. This process was associated with degradation of the crystalline peroxisomes. However, when cells were transferred into glucose/methylamine media, amine oxidase activity only declined during 2 h after the transfer and thereafter increased again. This subsequent rise in amine oxidase activity was associated with the development of new peroxisomes in the cells in which degradation of the crystalline peroxisomes, originally present, continued. These newly formed organelles probably originated from peroxisomes which had not been affected by degradation. When in the methanollimited chemostat methylamine was replaced by ammonium sulphate, repression of the synthesis of amine oxidase was observed. However, inactivation of this enzyme or degradation of peroxisomes was not detected. The decrease of amine oxidase activity in the culture was accounted for by dilution of enzyme as a result of growth and washout.
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  • 174
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    Archives of microbiology 137 (1984), S. 104-108 
    ISSN: 1432-072X
    Keywords: Chloroquine ; Yeast ; Proteolysis ; ATP hydrolysis ; Glucose consumption ; Ethanol formation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In yeast cells, degradation of cellular proteins was inhibited by addition of chloroquine to the medium as shown by a decrease of the release of trichloroacetic acid-soluble radioactivity from prelabelled cell protein. Penetration of chloroquine into the cells was strongly enhanced with increasing pH value of the medium. The concentration in the cells reached 5–14 times that in the medium of pH 8.0. Fluorescence microscopy showed that chloroquine was concentrated in the vacuoles of the cells. Chloroquine, at concentrations attained in the cells, inhibited the activities of vacuolar proteinases in vitro. Furthermore, chloroquine caused a rapid and drastic decrease of the ATP content of the cells and prevented the fermentation of glucose and formation of ethanol under aerobic conditions.
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  • 175
    ISSN: 1432-072X
    Keywords: Agglutination substance ; Cell-cell recognition ; Glycoprotein ; Mating ; Saccharomyces cerevisiae ; Sexual agglutinability ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract An a-mating-type-specific substance responsible for sexual agglutination was purified to 397-times in specific activity (units/mg protein) from the cytoplasm of a-mating type cells. The purified substance gave a single band stained with PAS reagent but not with both Coomassie brilliant blue and silver staining reagent by polyacrylamide gel electrophoresis in the presence of 8 M urea. However, incorporation of [35S]methionine and Lowry reaction clearly indicate that the substance is a glycoprotein. The substance specifically masked sexual agglutinability of cells of the opposite mating type α, indicating univalent action. The substance is a glycoprotein with a carbohydrate content of 90%, a pI of 4.5, and a molecular weight of 130,000. The substance was inactivated by 2-mercaptoethanol and proteolytic enzymes but not by glycolytic enzymes. The substance formed a complementary complex having no biological activity when mixed with α-agglutination substance from the wall or cytoplasm of α-cells in vitro.
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  • 176
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    Archives of microbiology 131 (1982), S. 298-301 
    ISSN: 1432-072X
    Keywords: Yeast ; Protoplast ; Compartmentation ; Vacuole ; Trehalose ; Trehalase ; Carbohydrate metabolism ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Protoplasts of Saccharomyces cerevisiae synthesized and degraded trehalose when they were incubated in a medium containing traces of glucose and acetate. Such protoplasts were gently lyzed by the polybase method and a particulate and soluble fraction was prepared. Trehalose was found in the soluble fraction and the trehalase activity mostly in the particulate fraction which also contained the vacuoles besides other cell organelles. Upon purification of the vacuoles, by density gradient centrifugation, the specific activity of trehalase increased parallel to the specific content of vacuolar markers. This indicates that trehalose is located in the cytosol and trehalase in the vacuole. It is suggested that trehalose, in addition to its role as a reserve may also function as a protective agent to maintain the cytosolic structure under conditions of stress.
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  • 177
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    Keywords: Hansenula polymorpha ; Peroxisomes ; Methanol ; Dihydroxyacetone synthase ; Cell fractionation ; Immunocytochemistry
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The subcellular localization of dihydroxyacetone synthase (DHAS) in the methylotrophic yeast Hansenula polymorpha was studied by various biochemical and immunocytochemical methods. After cell fractionation involving differential and sucrose gradient centrifugation of protoplast homogenates prepared from methanol-grown cells, DHAS cosedimented with the peroxisomal enzymes alcohol oxidase and catalase. Electron microscopy of this fraction showed that it contained mainly intact peroxisomes, whereas SDS-polyacrylamide gel electrophoresis revealed two major protein bands (75 and 78 kDa) which were identified as alcohol oxidase and DHAS, respectively. The localization of DHAS in peroxisomes was further established by immunocytochemistry. After immuno-gold staining carried out on ultrathin sections of methanol-grown H. polymorpha using DHAS-specific antibodies, labelling was confined to the peroxisomal matrix.
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  • 178
    ISSN: 1432-072X
    Keywords: Yeast ; Phosphate uptake ; Antigenic relationships ; Cell wall proteins ; Candida tropicalis
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    Topics: Biology
    Notes: Abstract Immunological cross-reactivity between cell wall proteins obtained from two yeast genera (Candida tropicalis and Saccharomyces cerevisiae) is reported. Specific retention of two cell wall proteins from Saccharomyces cerevisiae by an immunoabsorbent column coupled with antibodies against phosphate binding protein 2 (PiBP2) from Candida tropicalis allowed to generate antibodies against the proteins from S. cerevisiae. These antibodies were effective in inhibiting phosphate uptake by S. cerevisiae cells. The proteins from S. cerevisiae displayed a phosphate binding activity which was inhibited in the presence of the forementioned antibodies. These results and the observation that the amount of these proteins in the shock fluid was dependent of the growth conditions (i.e., in the presence or in the absence of phosphate) support the idea that these proteins are involved in the high affinity phosphate transport system.
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  • 179
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    Archives of microbiology 150 (1988), S. 309-312 
    ISSN: 1432-072X
    Keywords: Chloroquine ; Glycolytic enzymes ; Yeast ; Chloroquine and ATP/ADP
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Chloroquine at pH 8.0 and 10 mM concentration inhibits about 30% glucose consumption and ethanol formation in yeast cells. Out of the 11 glycolytic enzymes assayed, phosphoglycerate kinase and pyruvate decarboxylase have been found to be most sensitive to chloroquine. Next sensitive are hexokinase, glyceraldehyde-3-phosphate dehydrogenase and pyruvate kinase. Kinetic studies with the three kinases studied revealed competitive inhibition of chloroquine with ATP (hexokinase, phosphoglycerate kinase) or ADP (pyruvate kinase).
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  • 180
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    Archives of microbiology 151 (1988), S. 20-25 
    ISSN: 1432-072X
    Keywords: Yeast ; Saccharomyces cerevisiae ; Mating ; Zygote formation ; Chloroquine ; Lysosomotropic agent ; Plasma membrane ; Cell fusion
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Haploid cells of opposite mating type of Saccharomyces cerevisiae conjugate to form zygote. During the conjugation process, the degradation or reorganization of the cell wall and the fusion of the two plasma membranes take place. Since chloroquine inhibits cellular events associated with the reorganization of the plasma membrane, the effect of the drug on conjugation was studied. Chloroquine at a concentration, at which cell growth was not retarded, inhibited zygote formation, while it did not affect other mating functions, such as sexual agglutination, production of and response to mating pheromone. Cells in a mating culture containing chloroquine formed no “prezygote” suggesting that they were not prepared for entering into fusion process. The inhibitory effect of chloroquine was reversible as cells formed zygote when they were washed after treatment with chloroquine. Zygote formation was unaffected in cells possessing chlorquine within vacuoles after incubation with the drug in complete medium (YPD) at pH 7.5, followed by washing. This suggests that chloroquine inhibits zygote formaton by adsorbing to the plasma membrane of S. cerevisiae.
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  • 181
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    Archives of microbiology 132 (1982), S. 144-148 
    ISSN: 1432-072X
    Keywords: Yeast ; Cell wall ; Mannoproteins ; Envelope turnover ; Concanavalin A
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract By pulse and chase labeling experiments, two independent mannoprotein pools have been found associated with the Saccharomyces cerevisiae envelope. One of them probably corresponds to mannoproteins localized in the periplasmic space. These molecules showed a high turnover rate at 28° C. The second pool is formed by intrinsic wall mannoproteins which are apparently stable for long periods of time, after a small initial turnover. These results suggest that at least part of the mannoproteins initially found in the periplasmic space may move into the wall. The time lag between the addition of the radioactive precursors and their incorporation in the cell envelope (20–30 min for amino acids and about 10 min for carbohydrate) indicates that protein formation and carbohydrate incorporation take place in succession. Moreover, bulk glycosylation of mannoproteins seems to occur close in time to the moment of secretion into the periplasmic space.
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  • 182
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    Keywords: Carbon catabolite inactivation ; Yeast ; Malate dehydrogenase ; Fructose-1,6-bisphosphatase ; Glycolytic block mutants
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Experimental conditions have been elaborated to test for reversibility of the malate dehydrogenase inactivation (E.C.1.1.1.37) after addition of glucose to derepressed yeast cells. Malate dehydrogenase inactivation was shown to be irreversible at all stages of inactivation. In contrast fructose-1,6-bisphosphatase inactivation (E.C.3.1.11) remained reversible for at least 30 min after addition of glucose. Rapid reversible inactivation of fructose-1,6-bisphosphatase and irreversible inactivation of malate dehydrogenase were additionally investigated in glycolytic block mutants. Normal inactivation kinetics were observed in mutants without catalytic activity of phosphoglucoseisomerase (E.C.5.3.1.9), phosphofructokinase (E.C.2.7.1.11), triosephosphate isomerase (E.C.5.3.1.1) and phosphoglycerate kinase (E.C.2.7.2.3). Hence, neither type of inactivation depended on the accumulation of any glucose metabolite beyond glucose-6-phosphate. Under anaerobic conditions irreversible inactivation was completely abolished in glycolytic block mutants. In contrast rapid reversible inactivation was independent of energy provided by respiration or fermentation. Reversibility of fructose-1,6-bisphosphatase inactivation was tested under conditions which prevented irreversible malate dehydrogenase inactivation. In these experiments, fructose-1,6-bisphosphatase inactivation remained reversible for at least 120 min, whereas reversibility was normally restricted to about 30 min. This indicated a common mechanism between the irreversible part of fructose-1,6-bisphosphatase inactivation and irreversible malate dehydrogenase inactivation.
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  • 183
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    Keywords: Induction ; Catabolic repression ; galactose metabolism ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Induction and repression kinetics of alphagalactosidase, galactose uptake system and Leloir pathway enzymes were studied in chemostat cultures by changing the medium feed from glucose (11 mM) to glucose and galactose (11 mM; 17 mM respectively) in the induction experiments; and from galactose (11 mM) or (111 mM) to galactose plus glucose (83 mM) in the repression experiments. Basal levels of alpha-galactosidase and glucose uptake could be estimated in glucose-limited yeast cells, but it was not possible to detect any glactose pathway enzyme activity. In the repression experiments under galactose-limited or galactose-sufficient yeast cells, alpha-galactosidase and galactokinase decayed with K d=-0.21h-1=-D; that is, synthesis of these enzymes ceased (catabolite repression). In contrast transferase and epimerase activities and galactose uptake, decreased with K d values of-0.33 and-0.54h-1, showing that these activities were also subject to catabolite inactivation.
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  • 184
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    Archives of microbiology 146 (1987), S. 327-331 
    ISSN: 1432-072X
    Keywords: Actinomycetes ; Nitrogen fixation ; Symbiosis ; Immunocytochemistry ; Ultracryotomy
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Immunogoldlabelling on ultrathin cryosections of Frankia sp. Cc1.17 showed specific labelling of nitrogenase in the spherical cells called vesicles. No label was found in the hyphae in any cells grown on a medium with combined nitrogen, nor in those to which no specific antiserum was added. Similar results were obtained with cultures grown under high (20%) and low (2%) oxygen tension in the gas phase.
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  • 185
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    Archives of microbiology 150 (1988), S. 37-41 
    ISSN: 1432-072X
    Keywords: Yeast ; Hexose transport ; Sugar ; Malate uptake ; 2,4-DNP ; Zygosaccharomyces bailii
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract When grown in fructose or glucose the cells of Zygosaccharomyces bailii were physiologically different. Only the glucose grown cells (glucose cells) possessed an additional transport system for glucose and malate. Experiments with transport mutants had lead to the assumption that malate and glucose were transported by one carrier, but further experiments proved the existence of two separate carrier systems. Glucose was taken up by carriers with high and low affinity. Malate was only transported by an uptake system and it was not liberated by starved malate-loaded cells, probably due to the low affinity of the intracellular anion to the carrier. The uptake of malate was inhibited by fructose, glucose, mannose, and 2-DOG but not by non metabolisable analogues of glucose. The interference of malate transport by glucose, mannose or 2-DOG was prevented by 2,4-dinitrophenol, probably by inhibiting the sugar phosphorylation by hexokinase. Preincubation of glucose-cells with metabolisable hexoses promoted the subsequent malate transport in a sugar free environment. Preincubation of glucose-cells with 2-DOG, but not with 2-DOG/2,4-DNP, decreased the subsequent malate transport. The existence of two separate transport systems for glucose and malate was demonstrated with specific inhibitors: malate transport was inhibited by sodium fluoride and glucose transport by uranylnitrate. A model has been discussed that might explain the interference of hexoses with malate uptake in Z. bailii.
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  • 186
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    Archives of microbiology 151 (1989), S. 198-202 
    ISSN: 1432-072X
    Keywords: Sexual agglutination ; Mating ; Saccharomyces cerevisiae ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Genetic regulation of the inducibility of sexual agglutination ability in the yeast Saccharomyces cerevisiae was studied. Detailed analysis of the degree of sexual agglutination was carried out; it showed that a greater number of genes are involved in the regulation of inducible sexual agglutination in strain H1-0 than previously assumed. Although dominancy of inducible phenotype over constitutive was confirmed, the effectiveness of one gene changing the constitutive phenotype to the inducible seemed to be somewhat low. Quantity per cell of agglutination substances responsible for sexual agglutination increased as the agglutination ability became greater.
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  • 187
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    Archives of microbiology 124 (1980), S. 285-287 
    ISSN: 1432-072X
    Keywords: Yeast ; Saccharomyces cerevisiae ; Heat killing ; Membrane damage ; Genetic damage ; Growth temperature
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The resistance of exponentially growing yeast cells to killing by exposure to 52°C increased markedly as the growth temperature was increased. Identical killing curves were obtained for cells suspended in growth medium or in 0.9% saline. Cells resistant to killing at 52°C were quite sensitive to killing at slightly higher temperatures. These results suggest a primary role for membrane damage in the mechanism of heat killing.
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  • 188
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    Archives of microbiology 133 (1982), S. 155-161 
    ISSN: 1432-072X
    Keywords: Microtubule ; Nocodazole ; Yeast ; Cell cycle ; Dimorphism ; Fungus ; Wangiella dermatitidis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The microtubule inhibitor nocodazole {methyl-5-[2-(thienylcarbonyl)-1H-benzimidazol-2-yl]-carbamate} prevented nuclear migration and nuclear division in yeasts and developing multicellular forms of the polymorphic fungus Wangiella dermatitidis. It did not prevent yeast bud formation during at least two or three budding cycles, and caused yeasts to accumulate as premitotic forms with one to three buds. The effects of the drug suggested that at least three control pathways were involved in the yeast cell cycle; that the nocodazole block point was separate from the execution points of two temperature-sensitive mutations which lead to multicellularity; and that microtubules were controlling neither the yeast budding process nor the development of multicellular forms.
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  • 189
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    Archives of microbiology 133 (1982), S. 131-136 
    ISSN: 1432-072X
    Keywords: Fructose-bisphosphatase deficient mutants ; Yeast ; Schizosaccharomyces pombe ; Gluconeogenesis ; Glucose repression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We showed that in the yeast Schizosaccharomyces pombe, fructose-bisphosphatase is not subject to catabolite inactivation as it was observed in Saccharomyces cerevisiae. However, this enzyme activity is sensitive to catabolite repression in both yeasts. Two mutants lacking completely fructose-bisphosphatase activity were found. They were unable to grow on glycerol medium. They were still respiratory competent and exhibited the ability to derepress partially malate dehydrogenase activity. In glucose exponential phase culture, the parental strain lacks completely the fructosebisphosphatase activity due to catabolite repression. In these conditions, the growth is slowed down only in the mutants eventhough both mutants and their parental strain lack this enzyme activity. Normal sporulation and poor spore germination were observed for one mutant whereas, only in the presence of glucose, normal sporulation and normal spore germination were observed for the second mutant. Mendelian segregation of glycerol growth was found for the well germinating mutant. It is of nuclear heredity. The two mutations appeared to be closely linked.
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  • 190
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    Archives of microbiology 134 (1983), S. 270-275 
    ISSN: 1432-072X
    Keywords: Candida utilis ; Brettanomyces intermedius ; Yeast ; Glycolysis ; Vacuole ; Cytoplasm ; Phosphate compartmentation ; Phosphate transport ; Polyphosphate
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract 31P NMR was used to study the dynamics of phosphate pools during substrate utilization by aerobic and anaerobic suspensions of the yeast Candida utilis and by aerobic suspensions of the yeast Brettanomyces intermedius. In both yeast, the cytoplasmic pH was monitored; in C. utilis also the vacuolar pH could be measured. When glucose was used as a substrate for C. utilis, the vacuolar store of inorganic phosphorus (both orthophosphate and polyphosphate) was mobilized to replenish cytoplasmic phosphate which had become very low due to the build-up of high sugar phosphate levels. The hydrolysis of polyphosphate was glucose-dependent; it did not occur with ethanol as the substrate. After glucose depletion resynthesis of polyphosphate occurred only under aerobic conditions; anaerobic C. utilis cells continued to hydrolyze vacuolar polyphosphate. This difference between the aerobic and anaerobic suspension could be related to differences in cellular ATP levels. When ethanol was employed as a substrate, both Candida utilis and Brettanomyces intermedius exhibited a substantial increase in polyphosphate levels. These observations suggested a dual role for polyphosphate in yeasts both as a phosphate and an energy store. The cytoplasmic pH in C. utilis displayed characteristic responses to metabolic changes during glucose degradation. B. intermedius experienced a strong cytoplasmic acidification upon ethanol utilization due to acetic acid formation. The mechanism of transport of Pi across the vacuolar membrane in C. utilis appeared to be different from that reported for the plasma membrane.
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  • 191
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    Archives of microbiology 130 (1981), S. 87-89 
    ISSN: 1432-072X
    Keywords: Yeast ; Kluyveromyces ; 6-Deoxyglucose ; Glucose transport
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The inducible glucose transport system of the yeast, Kluyveromyces lactis, was studied using the nonmetabolizeable glucose analogue, 6-deoxyglucose. The free sugar analogue is transported into glucose-grown cells via a facilitated diffusion system as determined by the nonconcentrative uptake of the sugar analogue, by the failure of energy inhibitors to reduce the rate of transport and by exchange diffusion across the membrane. Free 6-deoxyglucose is also transported into succinate-grown cells passively. Induction experiments revealed that 6-deoxyglucose serves as a gratuitous inducer for the glucose transport system in this yeast.
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  • 192
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    Keywords: Yeast ; Saccharomyces cerevisiae ; Mating reaction ; Zygote formation ; Mating pheromone ; Fatty acid ; Arachidonic acid
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    Topics: Biology
    Notes: Abstract Effect of exogenous fatty acids on zygote formation in Saccharomyces cerevisiae was studied. Arachidonic and oleic acids considerably stimulated zygote formation, but other fatty acids tested, linoleic, linolenic, stearic and palmitic acids, did not. Pretreatment experiments with arachidonic acid showed that the stimulation of zygote formation by the fatty acid required the presence of mating pheromone.
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  • 193
    ISSN: 1432-072X
    Keywords: d-Xylose fermentation ; Aeration level ; Xylose reductase ; Xylitol dehydrogenase ; Yeast ; Candida shehatae ; Candida tenuis ; Pichia stipitis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The relationship between the degree of aerobiosis, xylitol production and the initial two key enzymes of d-xylose metabolism were investigated in the yeasts Pichia stipitis, Candida shehatae and C. tenuis. Anoxic conditions severely curtailed growth and retarded ethanol productivity. This, together with the inverse relationship between xylitol accumulation and aeration level, suggested a degree of redox imbalance. The ratios of NADH- to NADPH-linked xylose reductase were similar in all three yeasts and essentially independent of the degree of aerobiosis, and thus did not correlate with their differing capacities for ethanol production, xylitol accumulation or growth under the different conditions of aerobiosis. Under anoxic conditions the enzyme activity of Pichia stipitis decreased significantly, which possibly contributed to its weaker anoxic fermentation of xylose compared to C. shehatae.
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  • 194
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    Archives of microbiology 152 (1989), S. 564-566 
    ISSN: 1432-072X
    Keywords: l-Malate ; Schizosaccharomyces malidevorans ; Yeast
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    Topics: Biology
    Notes: Abstract The yeast Schizosaccharomyces malidevorans utilizes l-malate when grown on glucose as the carbon source. A mutant of this yeast has been isolated which is dependent on the presence of both l-malate and glucose for growth. The mutant utilizes l-malate as rapidly as the wildtype and the utilization of glucose is greatly reduced. Other TCA cycle intermediates do not relieve the malate dependence.
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  • 195
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    Archives of microbiology 149 (1988), S. 261-267 
    ISSN: 1432-072X
    Keywords: Yeast ; Hanseniaspora uvarum ; Pichia kluyveri ; Killer toxin ; dsRNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract By heat treatment killer strains of the type K1 of Saccharomyces cerevisiae that are known to harbour dsRNA plasmids were completely cured, whereas only a small fraction of the clones of the killer type K2 had lost the dsRNA dependent killer character. The K2 killers but not the strains of killer type K1 were easily cured by cycloheximide. Killer strains of Hanseniaspora uvarum were not curable by heat treatment. Curing was successfull with cycloheximide or 5-fluorouracil. Two double-stranded RNA plasmids were detected in the killer strains of H. uvarum. The smaller dsRNA plasmid was absent in the strains that were cured of their killer character by 5-fluorouracil. The killer character of H. uvarum was transferred to S. cerevisiae by spheroplast fusion. The fusion products showing the killer character contained both dsRNA plasmids, obviously the smaller plasmid (M-dsRNA) carries the genes for killer toxin formation. Killer strains of Pichia kluyveri were not curable of their killer character, in these strains no dsRNA plasmids were detected.
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  • 196
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    Journal of industrial microbiology and biotechnology 2 (1987), S. 59-62 
    ISSN: 1476-5535
    Keywords: Yeast ; Zygosaccharomyces ; Spoilage ; Synergism ; pH ; °Brix
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Summary A quantitative method for the prediction of growth of the food spoilage yeastZygosaccharomyces bailii in a model fruit-drink system is described. A factorially designed experiment was employed to produce polynomial equations relating pH and sugar concentration (°brix) to the lag period and doubling time of this yeast. Low pH values (〈3.0) and high °brix values (〉40) show a strong synergistic action on the extension of lag period, which could be used, along with the model presented, in the formulation of product preservation systems.
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  • 197
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    Journal of industrial microbiology and biotechnology 2 (1987), S. 159-165 
    ISSN: 1476-5535
    Keywords: Yeast ; Genetic stability ; Saccharomyces cerevisiae ; Selection ; Reproductive fitness
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Summary The potential for changes in allele frequencies in yeast populations by selection was examined. Cells from the wine yeastSaccharomyces cerevisiae (strain Montrachet) were grown over a large number of generations using two different culturing techniques, each with two variations: serial transfers on WLN agar plates with and without UV irradiation, and continuous culture in autoclaved and in filter-sterilized grape must. A low frequency of variant isozyme patterns was found in samples taken at the end of the experiment. Growth rates in must and on agar plates were also examined, and it was found that all samples were faster-growing than the original strain, to varying degrees. Applications for the selection system developed are discussed.
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  • 198
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    Journal of industrial microbiology and biotechnology 2 (1988), S. 365-372 
    ISSN: 1476-5535
    Keywords: Osmotic pressure ; Intracellular ethanol ; Yeast ; Nutrient ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Summary The intracellular accumulation of ethanol in yeast and its potential effects on growth and fermentation have been topics of controversy for the past several years. The determination of intracellular ethanol based on the exclusion of [14C]sorbitol to estimate aqueous cell volume was used to examine the question of intracellular ethanol accumulation. An intracellular accumulation of ethanol inSaccharomyces cerevisiae was observed during the early stages of fermentation. However, as fermentation continued, the intracellular and extracellular concentrations of ethanol became similar. Increasing the osmotic pressure of the medium with glucose or sorbitol was observed to cause an increase in the intracellular ethanol concentration. Associated with this was a decrease in yeast growth and fermentation rates. In addition, increasing the osmotic pressure of the medium was observed to cause an increase in glycerol production. Supplementation of the media with excess peptone, yeast extract, magnesium sulfate and potassium phosphate was found to relieve the detrimental effects of high osmotic pressure. Under these conditions, though, no effect on the intracellular and extracellular ethanol distribution was observed. These results indicate that nutrient limitation, and not necessarily intracellular ethanol accumulation, plays a key role during yeast fermentations in media of high osmolarity.
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  • 199
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    Planta 165 (1985), S. 522-526 
    ISSN: 1432-2048
    Keywords: Albumin (localisation) ; Cotyledon ; Pisum (albumin protein) ; Immunocytochemistry
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The major albumin protein in storage parenchyma tissue of developing peas has been localised at an ultrastructural level by immunocytochemistry. Tissue was fixed in buffered aldehyde and embedded in LR White resin which was polymerised by addition of catalyst. Sections were labelled by the indirect method of absorption of Protein A-gold to specifically bound antibodies. This method gives high levels of specific labelling on sections which retain good ultrastructural preservation and have high contrast after conventional staining. The albumin is located throughout the cytoplasm although no labelling was found associated with the endoplasmic reticulum, Golgi apparatus, vacuoles-protein bodies or other organelles.
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  • 200
    ISSN: 1432-2048
    Keywords: Cotyledon ; Golgi complex ; Immunocytochemistry ; Phaseolus (seed proteins) ; Phaseolin ; Phytohemagglutinin ; Protein (seeds)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Development of legume seeds is accompanied by the synthesis of storage proteins and lectins, and the deposition of these proteins in protein-storage vacuoles (protein bodies). We examined the subcellular distribution, in developing seeds of the common bean, Phaseolus vulgaris L., of the major storage protein (phaseolin) and the major lectin (phytohemagglutinin, PHA). The proteins were localized using an indirect immunocytochemical method in which ultrathin frozen sections were immunolabeled with rabbit antibodies specific for either PHA or phaseolin. Bound antibodies were then localized using goat-anti-rabbit immunoglobulin G adsorbed onto 4- to 5-nm colloidal gold particles. The sections were post-fixed with OsO4, dehydrated, and embedded in plastic on the grids. Both PHA and phaseolin exhibited a similar distribution in the storage-parenchyma cells, being found primarily in the developing protein bodies. Endoplasmic reticulum and Golgi complexes (cisternal stacks and associated vesicles) also were specifically labeled for both proteins, whereas the cytosol and other organelles, such as mitochondria, were not. We interpret these observations as supporting the hypothesis that the transport of storage proteins and lectins from their site of synthesis, the rough endoplasmic reticulum, to their site of deposition, the protein bodies, is mediated by the Golgi complex.
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