Influence of genetic background on the occurrence of chromosomal rearrangements in Saccharomyces cerevisiae

<p>Abstract</p> <p>Background</p> <p>Chromosomal rearrangements such as duplications and deletions are key factors in evolutionary processes because they promote genomic plasticity. Although the genetic variations in the <it>Saccharomyces cerevisiae </it>species have been well documented, there is little known to date about the impact of the genetic background on the appearance of rearrangements.</p> <p>Results</p> <p>Using the same genetic screening, the type of rearrangements and the mutation rates observed in the S288c <it>S. cerevisiae </it>strain were compared to previous findings obtained in the FL100 background. Transposon-associated rearrangements, a major chromosomal rearrangement event selected in FL100, were not detected in S288c. The mechanisms involved in the occurrence of deletions and duplications in the S288c strain were also tackled, using strains deleted for genes implicated in homologous recombination (HR) or non-homologous end joining (NHEJ). Our results indicate that an Yku80p-independent NHEJ pathway is involved in the occurrence of these rearrangements in the S288c background.</p> <p>Conclusion</p> <p>The comparison of two different <it>S</it>. <it>cerevisiae </it>strains, FL100 and S288c, allowed us to conclude that intra-species genomic variations have an important impact on the occurrence of chromosomal rearrangement and that this variability can partly be explained by differences in Ty1 retrotransposon activity.</p

Ploidy influences cellular responses to gross chromosomal rearrangements in saccharomyces cerevisiae

<p>Abstract</p> <p>Background</p> <p>Gross chromosomal rearrangements (GCRs) such as aneuploidy are key factors in genome evolution as well as being common features of human cancer. Their role in tumour initiation and progression has not yet been completely elucidated and the effects of additional chromosomes in cancer cells are still unknown. Most previous studies in which <it>Saccharomyces cerevisiae </it>has been used as a model for cancer cells have been carried out in the haploid context. To obtain new insights on the role of ploidy, the cellular effects of GCRs were compared between the haploid and diploid contexts.</p> <p>Results</p> <p>A total number of 21 haploid and diploid <it>S. cerevisiae </it>strains carrying various types of GCRs (aneuploidies, nonreciprocal translocations, segmental duplications and deletions) were studied with a view to determining the effects of ploidy on the cellular responses. Differences in colony and cell morphology as well as in the growth rates were observed between mutant and parental strains. These results suggest that cells are impaired physiologically in both contexts. We also investigated the variation in genomic expression in all the mutants. We observed that gene expression was significantly altered. The data obtained here clearly show that genes involved in energy metabolism, especially in the tricarboxylic acid cycle, are up-regulated in all these mutants. However, the genes involved in the composition of the ribosome or in RNA processing are down-regulated in diploids but up-regulated in haploids. Over-expression of genes involved in the regulation of the proteasome was found to occur only in haploid mutants.</p> <p>Conclusion</p> <p>The present comparisons between the cellular responses of strains carrying GCRs in different ploidy contexts bring to light two main findings. First, GCRs induce a general stress response in all studied mutants, regardless of their ploidy. Secondly, the ploidy context plays a crucial role in maintaining the stoichiometric balance of the proteins: the translation rates decrease in diploid strains, whereas the excess protein synthesized is degraded in haploids by proteasome activity.</p

The monomeric glutamyl-tRNA synthetase of Escherichia coli. Purification and relation between its structural and catalytic properties.

The glutamyl-tRNA synthetase has been purified to homogeneity from Escherichia coli with a yield of about 50%. It is a monomer with a molecular weight of 56,000 and has the same kinetic properties as those of the alpha chain of the dimeric alphabeta-glutamyl-tRNA synthetase described previously (Lapointe, J., and Söll, D. (1972) J. Biol. Chem. 247, 4966-4974). It is the smallest amino-acyl-tRNA synthetase purified from E. coli and contains no important sequence repetition. It is also the only monomeric aminoacyl-tRNA synthetase reported so far to contain no major sequence duplication. Considering its structural and mechanistic similarities with the glutaminyl- and the arginyl-tRNA synthetases of E. coli, we propose the existence of a relation between the true monomeric character of the glutamyl-tRNA synthetase (as opposed to monomers with sequence duplications) and its requirement for tRNA in the activation of glutamate. A single sulfhydryl group of the native enzyme reacts with 5,5'-dithiobis(2-nitrobenzoic acid) causing no loss of enzymatic activity, whereas four such groups per enzyme react in the presence of 4 M guanidine HCl.journal article1979 Jan 25importe

The primary structure of the aspartate transcarbamylase region of the URA2 gene product in Saccharomyces cerevisiae. Features involved in activity and nuclear localization.

The yeast URA2 locus encodes a multifunctional protein which possesses the carbamylphosphate synthetase and aspartate transcarbamylase activities and which catalyzes the first two reactions of the pyrimidine pathway. We report here the nucleotide sequence of the central and the 3' region of this locus. The latter encodes that part of the multifunctional protein which has the aspartate transcarbamylase activity. The deduced amino acid sequence shows a high degree of homology with the known aspartate transcarbamylases of various organisms from Escherichia coli to mammals. The amino acid residues that have been shown to be involved in the catalytic site of the E. coli enzyme are all conserved suggesting that, in the more complex structure of the yeast protein, the catalytic sites are also located at subunit interfaces. There is also an important conservation of the amino acid pairs that, in E. coli, are implicated in intra- and interchain interactions. As well as the oligomeric structure suggested by these two features, the three-dimensional structure of the yeast enzyme must also be organized to account for the channeling of carbamylphosphate, from the carbamylphosphate synthetase catalytic site to that of aspartate transcarbamylase, and for the concomitant feedback inhibition of the two activities by the end product UTP. The URA2 gene product was shown to be localized in the nucleus. With the aim of identifying the regions that may be involved in this transport, we have determined by electron microscopy the subcellular distribution of aspartate transcarbamylase in three strains expressing different fragments of the URA2 locus. In the first strain the protein lacks 190 residues at the N terminus, but accumulates normally in the nucleus. In the second strain the protein lacks 382 residues in the central part and seems impaired in the nuclear transport process. In the third strain the 476-residue protein encoded by the 3' region of URA2 locus and catalyzing the aspartate transcarbamylase reaction is able by itself to migrate to and accumulate in the nucleus. This suggests that two regions are involved in the nuclear accumulation. On the basis of their conservation in analogous proteins of other eukaryotes and their similarity to sequences already identified as nuclear location signals, a sequence in the central region of the protein and two short sequences in the C-terminal region are good candidates for the nuclear location signal involved in the targeting of the URA2 product.comparative studyjournal article1989 May 15importe

Playing By the Rules: Gambling and Social Identity in Early Modern German Towns

We have constructed viable Saccharomyces cerevisiae strains containing a reciprocal translocation between the URA2 site of chromosome X and the HIS3 site of chromosome XV. Our methodology is an extension of the method originally developed to introduce an altered cloned sequence at the chromosomal location from which the parent sequence was derived (S. Scherer and R.W. Davis, Proc. Natl. Acad. Sci. U.S.A. 76:4951-4955, 1979). It comprises three essential steps. First, a nonreverting ura2- strain was constructed by deleting a 3.7-kilobase fragment from the coding sequence of the wild-type URA2 gene. Second, part of the coding sequence of the wild-type URA2 gene (without promotor) was inserted at the HIS3 locus of the ura2- strain. Third, after several generations of growth on uracil-supplemented medium, ura2+ colonies were selected which resulted from mitotic recombination between the nonoverlapping deletions of URA2 located on chromosomes X and XV.journal articleresearch support, non-u.s. gov't1982 Sepimporte

Structural studies on aminoacyl-tRNA synthetases. A tentative correlation between the subunit size and the occurrence of repeated sequences.

Recent studies have shown that those synthetases with subunits greater than 85,000 daltons contain extensive repeated sequences, whilst those with small subunits (40,000 daltons) do not. We have undertaken a comparative study of four aminoacyl-tRNA synthetases (glutamyl-, arginyl-, valyl-, and phenylalanyl-tRNA synthetases) with subunit sizes ranging from 56,000 to 130,000 daltons in an attempt to correlate the occurrence and extent of the repeats with the length of the polypeptide chain. Our results show that monomeric glutamyl-tRNA synthetase from Escherichia coli (56,000 daltons) contains few repeated sequences, whereas both subunits of yeast phenylalanyl-tRNA synthetase (alpha, 73,000 daltons; beta, 62,000 daltons) and yeast arginyl-tRNA synthetase (74,000 daltons) do have a significant amount of repeats. Thus 56,000 dalton appears to be the minimum size compatible with the existence of such repeats.journal article1980 Jul 24importe

The Zygosaccharomyces rouxii strain CBS732 contains only one copy of the HOG1 and the SOD2 genes.

The osmotolerant yeast Zygosaccharomyces rouxii CBS732 contains only one copy of the ZrHOG1 and ZrSOD2-22 genes. Both genes were cloned and sequenced (Acc. Nos. AJ132606 and AJ252273, respectively) and their sequences were compared to homologous pairs of genes from Z. rouxii ATCC42981 (genes Z-HOG1, Z-HOG2, Z-SOD2, Z-SOD22). The CBS732 ZrHog1p is shorter than its ATCC42981 counterparts (380 aa residues vs. 407 and 420 aa, respectively) and is more similar to ATCC42981 Z-Hog2p than to Z-Hog1p. Also its promoter region corresponds to that one of Z-HOG2. The CBS732 ZrHOG1 promoter region is recognised by Saccharomyces cerevisiae, and the gene product (MAP kinase ZrHog1p) presence fully complements the osmosensitivity of a S. cerevisiae hog1 mutant strain. The CBS ZrSOD2-22 gene is highly similar to ATCC42981 Z-SOD2 but it contains also a segment of 15 aa residues specific for Z-SOD22. Z. rouxii ZrSod2-22 Na(+)/H(+) antiporter expressed in S. cerevisiae shows better activity toward toxic Na(+) and Li(+) cations than does S. cerevisiae's own Nha1 antiporter, and is efficient in improving the halotolerance of some S. cerevisiae wild types.journal articleresearch support, non-u.s. gov't2001 Jun 15importe

Isolation and characterization of the TIM10 homologue from the yeast Pichia sorbitophila: a putative component of the mitochondrial protein import system.

The Saccharomyces cerevisiae TIM10 gene encodes one of the few essential mitochondrial proteins that are required for the import of nuclear-encoded precursor proteins from the cytosol and their subsequent sorting into the different mitochondrial compartments. We have isolated and characterized a putative homologue of TIM10 from the halotolerant yeast Pichia sorbitophila. The Pichia TIM10 gene encodes a protein of 90 amino acids with 66% identity to S. cerevisiae Tim10p. It was capable of suppressing the temperature sensitivity of tim10-1 mutant in S. cerevisiae, suggesting that Pichia TIM10 is both a functional and structural homologue of S. cerevisiae TIM10. The putative Pichia TIM10 gene product contains all the four conserved cysteine residues and the two CX(3)C motifs typical of the Tim family proteins in the mitochondrial intermembrane space. Using anti-Tim10p serum, Western blots detected a protein of about 10 kDa, suggesting that the Pichia Tim10p is a mitochondrial protein. The results suggest that mitochondrial import and sorting systems might be also strongly conserved in other fungi. The coding sequence of the P. sorbitophila TIM10 has been deposited in the EMBL Nucleotide Sequence Database under Accession No. AJ243940.journal articleresearch support, non-u.s. gov't2000 Mayimporte

Two-dimensional protein map of an "ale"-brewing yeast strain: proteome dynamics during fermentation.

The first protein map of an ale-fermenting yeast is presented in this paper: 205 spots corresponding to 133 different proteins were identified. Comparison of the proteome of this ale strain with a lager brewing yeast and the Saccharomyces cerevisiae strain S288c confirmed that this ale strain is much closer to S288c than the lager strain at the proteome level. The dynamics of the ale-brewing yeast proteome during production-scale fermentation was analysed at the beginning and end of the first and the third usage of the yeast (called generation in the brewing industry). During the first generation, most changes were related to the switch from aerobic propagation to anaerobic fermentation. Fewer changes were observed during the third generation but certain stress-response proteins such as Hsp26p, Ssa4p and Pnc1p exhibited constitutive expression in subsequent generations. The ale brewing yeast strain appears to be quite well adapted to fermentation conditions and stresses.journal articleresearch support, non-u.s. gov't2004 Decimporte

Cloning of Arabidopsis thaliana glutathione synthetase (GSH2) by functional complementation of a yeast gsh2 mutant.

Glutathione (L-gamma-glutamyl-L-cysteinylglycine, GSH) plays an important role in the protection of plants against various types of stress caused by reactive oxygen species, gazeous pollutants, heavy metals and xenobiotics. A cDNA fragment containing the entire coding unit for glutathione synthetase (GSH2) of Arabidopsis thaliana was cloned by complementation of the methylglyoxal sensitivity of a gsh2 mutant of the yeast Saccharomyces cerevisiae. The cDNA encodes a protein of 478 amino acids (deduced Mr: 53783), bearing clear sequence similarities to GSH2 products from frog embryos (Xenopus laevis), rat kidney (Rattus norvegicus) and from the fission yeast (Schizosaccharomyces pombe). A highly conserved glycine-rich domain close to the carboxy-terminus was found in the GSH2 product and appears to be typical for eukaryotic glutathione synthetases. The Mr is similar to those of soluble animal enzymes, suggesting that the Arabidopsis gene also codes for a cytosolic protein. Genomic DNA-blot analysis indicates the presence of a single GSH2 gene. The yeast gsh2 mutant becomes resistant to methylglyoxal and cadmium after transformation with the plasmid bearing the Arabidopsis GSH2 cDNA. Moreover, this increased resistance is correlated to the restoration of GSH content from below detectability in mutants to about 50% of the wild-type levels in transformed cells.comparative studyjournal article1996 Mar 01importe