Current Genetics

Henriques, João; Brendel, Martin

doi: 10.1007/BF00309906pmid: 2078865

Fujimura, Hiro-aki

doi: 10.1007/BF00309907pmid: 2078866

Mating pheromones, known as a and α-factors, arrest the division of cells of opposite mating types, α and a respectively, in Saccharomyces cerevisiae. I have cloned the DAC2 gene, which is required for both pheromone-induced division-arrest and cell-fusion during conjugation. The constructed dac2::LEU2 null mutation leads to defects in both pheromone-induced divisionarrest and cell-fusion during conjugation; it also suppresses the growth defect caused by the gpal mutation (a mutation in the α subunit of the S. cerevisiae G protein). These results indicate that DAC2 may be the same gene as FUS3, which was recently isolated by Elion et al. (1990) as a gene essential for cell-fusion during conjugation. The dac2::LEU2 null mutant also showed morphological alterations in response to mating pheromones. I show here that the DAC2 product plays an essential role in both the division-arrest signalling pathway of the yeast pheromone response and in cell-fusion during conjugation.

Oda, Yuji; Ouchi, Kozo

doi: 10.1007/BF00309908pmid: N/A

To investigate the leavening ability of yeast in dough, chromosome loss was induced by benomyl treatment in YOY1037, a diploid between a baking strain and a laboratory strain, and its effect on the leavening ability was studied. When benomyl-treated cells were spread on plates with a dye indicator for ploidy, about 20% of the visible colonies were stained dark blue or dark purple; the rest stained pale blue, similar to the diploid YOY1037. Strains showing the MATα phenotype, and non-galactose fermenting strains, apparently having lost particular chromosomes, were observed only in those with darkcoloured colonies. Strains with dark-coloured colonies showed a wider range of leavening ability than did those with pale-coloured colonies.

McNeil, J.; Dykshoorn, P.; Huy, J.; Small, S.

doi: 10.1007/BF00309909pmid: 2127735

We show by deletion mutagenesis, followed by in vivo and in vitro analysis, that the binding of a protein factor to the upstream activation sequence (USA) of the Saccharomyces cerevisiae glycolytic gene PYK, encoding pyruvate kinase, is required for efficient transcription of the corresponding coding region. In addition, gel electrophoretic mobility shift and DNase I protection studies, involving yeast gene products expressed in E. coli, suggest that this trans-acting DNA-binding protein is encoding by the RAP1 gene. The identification of RAP1 binding sites located within the UAS element of the yeast PYK, PGK (phosphoglycerate kinase) and ENO1 (enolase) genes, and in the 5′-upstream region of the ADHI (alcohol dehydrogenase) gene, suggests that a mechanism of coordinate gene expression involving several of the glycolytic genes may exist in yeast.

Tron, Thierry; Lemesle-Meunier, Danielle

doi: 10.1007/BF00309910pmid: 2078867

Two cytochrome b respiratory—deficient mutants were sequenced and their DNA base change identified, leading to the replacement of glycine (G137 by valine or glutamicacid. No variation in their cytochrome b content with regard to cytochrome oxidase and cytochrome (c+c1) was found to have occurred. Their cellular respiratory activity with various substrates was partly conserved and was totally inhibited by antimycin A. Their ubiquinol (QH2)-cytochrome c reductase/mole cytochrome b activity decreased by about 50%. Paradoxically their growth on respiratory substrate was abolished. Both mutants retained a high-affinity binding site for antimycin A, and exhibited a myxothiazol—resistance at the mitochondrial level. It seems likely that the mutated position (137), which belongs to the ubiquinol oxidizing domain of the bc1 complex, interferes, directly or indirectly, with the respiratory growth capacity of the cell.

Mittelmeier, Telsa; Dieckmann, Carol

doi: 10.1007/BF00309911pmid: 2150347

The nuclear gene product CBP1 stabilizes cytochrome b transcripts in yeast mitochondria. In cbp1 mutant strains, cytochrome b gene (cob) transcripts are not detectable by Northern blot analysis. The results of previous studies led to the hypothesis that CBP1 interacts with the 5′-untranslated sequence of the cob mRNA, or pre-mRNA, to stabilize the message. To determine what portion of the cob leader is sufficient for interaction with CBP1, we have investigated the stability of transcripts from a novel hybrid gene, cob-oli1, in which the 5′-terminal third of the cob leader sequence was fused to the coding sequence of the gene for ATP synthase subunit 9, oli1. The hybrid cob-oli1 transcript was stale in a strain wild-type at the CBP1 locus, but was undetectable in the cbp1 mutant background. That the cob-oli1 transcript was translated to produce ATP synthase subunit 9 in CBP1 strains containing the cob-oli1 gene was verified by 35S-methionine labeling of mitochondrial proteins. We conclude that the 5′-terminal portion of the cob message is sufficient for CBP1 function and discuss the hypothesis that CBP1 interacts directly with this region of the transcript to promote cob mRNA stability.

Radetzky, R.

doi: 10.1007/BF00309912pmid: N/A

The mitochondrial genome of several poplar clones has been characterized by restriction analysis and hybridization with gene probes from Oenothera. The mitochondrial (mt) DNA of Populus has a complex restriction fragment pattern and its genome size was estimated to be about 450 kilobase pairs (kb). Restriction fragment length polymorphisms (RFLPs) could be detected only between, and not within, species of Populus and were used as genetic markers to follow mitochondrial inheritance. Location of the apocytochrome b (cob) gene on different Eco RI or Hin dIII fragments in “Muhle Larsen” (P. trichocarpa) and “Androscoggin” (P. maximowiczii × P. trichocarpa) has been used for analysis of mitochondrial inheritance. All hybrids investigated exhibit a fragment pattern identical to that of the female parent. Hybridization with other gene probes (coxIII/HindIII, coxIII/SalI, coxIII/Xho I, atp6/Bgl1, atp6/Hco RI, atp6/Hin dIII, nad1/Hin dIII, nad1/Sal I, nad1/Xba I) showed the same results as given by cob hybridization. Thus, mtDNA seems to be inherited maternally in Populus.

Swart, Klaas; Vondervoort, Peter; Witteveen, Cor; Visser, Jaap

doi: 10.1007/BF00309913pmid: 2078868

A number of mutants of Aspergillus niger, affected in glucose oxidase (GOX) expression, are described. The overproducing mutants could be classified into seven complementation groups whereas two glucose oxidase-negative complementation groups were recognized. These nine gox loci were assigned to linkage groups using master strains with marked chromosomes. Three gox loci are in linkage group II, one is in III, two are in V and two are in linkage group VII. One weak glucose oxidase-overproducing mutant could not be assigned to one of the linkage groups. These genetically well characterized mutants will be used in a strain improvement program based on genetic recombination.

Fincham, J.

doi: 10.1007/BF00309914pmid: 2150348

In the further analysis of a cross in which the mis-sense allele, am 3, of the Neurospora crassa am (glutamate dehydrogenase) gene was present in one parent together with two ectopic wild-type gene copies, one ascus was identified in which the two ectopic copies had been inactivated by the RIP process whereas the am 3 allele continued to produce its characteristic enzyme variety in active, but heat-sensitive, form. The am 3 allele had also acquired a new HindIII restriction site. It had no detectable methylation. The mutations responsible respectively for the new restriction site and the modified enzyme properties were separated from each other, and from the original am 3 mutation, by selecting for intragenic recombination on either side of the am 3 site. In this way two new effectively wild-type alleles were generated, one characterised by its heat-sensitive and kinetically modified enzyme product and the other by a new HindIII site. These results demonstrate that the RIP phenomenon can be a source of new functional alleles.

Gruber, Franz; Visser, Jaap; Kubicek, Christian; Graaff, Leo

doi: 10.1007/BF00309915pmid: N/A

The Trichoderma reesei orotidine-5′-phosphate decarboxylase gene was isolated by heterologous hybridization with the corresponding Neurospora gene as a probe. A 2.7 kb SalI fragment, which exclusively hybridized to the Neurospora gene, was subcloned in pGEM-5Zf(+). This subclone was termed pFG1 and was used to transform a Trichoderma reesei pyrG- negative mutant to PYR+. The transformation frequency in this homologous system was up to 12000 transformants per μg DNA. About one-fifth of the transformants tested were abortive. Perfect mitotic stability was found in half of the non-abortive transformants, correlating with vector integration at homologous and ectopic loci. In the unstable transformants the transforming DNA appears to be present in the form of extrachromosomal elements.