A re-annotation of the Saccharomyces cerevisiae genome

Discrepancies in gene and orphan number indicated by previous analyses suggest that S. cerevisiae would benefit from a consistent re-annotation. In this analysis three new genes are identified and 46 alterations to gene coordinates are described. 370 ORFs are defined as totally spurious ORFs which should be disregarded. At least a further 193 genes could be described as very hypothetical, based on a number of criteria. It was found that disparate genes with sequence overlaps over ten amino acids (especially at the N-terminus) are rare in both S. cerevisiae and Sz. pombe. A new S. cerevisiae gene number estimate with an upper limit of 5804 is proposed, but after the removal of very hypothetical genes and pseudogenes this is reduced to 5570. Although this is likely to be closer to the true upper limit, it is still predicted to be an overestimate of gene number. A complete list of revised gene coordinates is available from the Sanger Centre (S. cerevisiae reannotation: ftp://ftp/pub/yeast/SCreannotation).     

Protosilencers in Saccharomyces cerevisiae subtelomeric regions

Saccharomyces cerevisiae subtelomeric repeats contain silencing elements such as the core X sequence, which is present at all chromosome ends. When transplaced at HML, core X can enhance the action of a distant silencer without acting as a silencer on its own, thus fulfilling the functional definition of a protosilencer. Here we show that an ACS motif and an Abf1p-binding site participate in the silencing capacity of core X and that their effects are additive. In addition, in a variety of settings, core X was found to bring about substantial gene repression only when a low level of silencing was already detectable in its absence. Adjoining an X-STAR sequence, which naturally abuts core X in subtelomeric regions, did not improve the silencing capacity of core X. We propose that protosilencers play a major role in a variety of silencing phenomena, as is the case for core X, which acts as a silencing relay, prolonging silencing propagation away from telomeres.     

Isoaccepting mitochondrial glutamyl-tRNA species transcribed from different regions of the mitochondrial genome of Saccharomyces cerevisiae

Mitochondrial glutamyl-tRNA isolated from mitochondria of Saccharomyces cerevisiae was separated into two distinct species by re versed-phase chromatography. The migration of the two mitochondrial glutamyl-tRNAs (tRNA_I^Glu and tRNA_II^Glu) differed from that of two glutamyl-tRNA species found in the cytoplasm of a mitochondrial DNA-less petite strain. Both mitochondrial tRNAs hybridized with mitochondrial DNA. Three lines of evidence demonstrate that mitochondrial tRNA_I^Glu and tRNA_II^Glu are transcribed from different mitochondrial cistrons. First the level of hybridization of a mixture of the two tRNAs to mitochondrial DNA was equal to the sum of the saturation hybridization levels of each glutamyl-tRNA alone. Second, the two mitochondrial glutamyl-tRNAs did not compete with each other in hybridization competition experiments. Finally the tRNAs showed individual hybridization patterns with different petite mitochondrial DNAs.Hybridization of the tRNAs to mitochondrial DNA of genetically defined petite strains localized each tRNA with respect to antibiotic resistance markers. The two glutamyl-tRNA cistrons were spatially separated on the genetic map.     

Evidence for the occurrence of nucleotide-activated oligosaccharides in Saccharomyces cerevisiae

Recently, nucleotide-activated oligosaccharides have been found to be involved in the biosynthesis of certain glycoconjugates in archaeal and bacterial procaryotes. This paper describes the isolation and partial chemical characterization of nucleotide-activated oligosaccharides from the eucaryotic microbe Saccharomyces cerevisiae. We purified four different nucleotide-activated oligosaccharides from cell extracts of Saccharomyces cerevisiae. Three of the oligosaccharides were UDP, and one was TDP-activated. D-Glucose was the only carbohydrate constituent, except for one oligosaccharide, which also contained glucosamine. The chain length varied between two and four sugar residues.     

Evidence for short-patch mismatch repair in Saccharomyces cerevisiae

Recombination events between non-identical sequences most often involve heteroduplex DNA intermediates that are subjected to mismatch repair. The well-characterized long-patch mismatch repair process, controlled in eukaryotes by bacterial MutS and MutL orthologs, is the major system involved in repair of mispaired bases. Here we present evidence for an alternative short-patch mismatch repair pathway that operates on a broad spectrum of mismatches. In msh2 mutants lacking the long-patch repair system, sequence analysis of recombination tracts resulting from exchanges between similar but non-identical (homeologous) parental DNAs showed the occurrence of short-patch repair events that can involve <12 nucleotides. Such events were detected both in mitotic and in meiotic recombinants. Confirming the existence of a distinct short-patch repair activity, we found in a recombination assay involving homologous alleles that closely spaced mismatches are repaired independently with high efficiency in cells lacking MSH2 or PMS1. We show that this activity does not depend on genes required for nucleotide excision repair and thus differs from the short-patch mismatch repair described in Schizosaccharomyces pombe.     

Decade of genomics--methods for genome investigation in yeast Saccharomyces cerevisiae

The last ten years, since yeast Saccharomyces cerevisiae genome was sequenced, brought a big impact in genome-wide techniques. The tenth anniversary of genomic era provokes the following resume: a lot of new methods were invented. The yeast strains libraries carrying transposon insertions, gene deletions or tagged proteins have been created. Using them, the phenotypes of gene deletions, as well as the biological activity, cellular localization and possible modifications of their protein products were elucidated. SAGE analysis and DNA microarray experiments showed gene expression profiles and allowed to build interaction networks of gene regulation. The two dimensional gels, mass spectrometry, protein arrays and two-hybrid system carry information about protein interactions, modifications, and biochemical activities. All these methods permit to increase the number of genes with known cellular functions. Moreover, testing these techniques on yeast S. cerevisiae--a model eukaryotic organism--opened the door for their usage in all other species.     

Evidence for dual physiological formsof ergosterol in Saccharomyces cerevisiae

Data obtained from acid hydrolysis and extraction of yeast have demonstrated that routine saponification does not recover total sterol from the cells. This suggests the existence of a form of ergosterol resistant to saponification. Time course analyses of sterol synthesis by resting cell suspensions reveal an inverse relationship between the amounts of base labile and acid labile forms of sterol. These data give strong presumptive evidence for dual forms of ergosterol which are interconvertible according to the respiratory state of the cell.     

Evidence for a new chromosome in Saccharomyces cerevisiae.

The current yeast map has 16 chromosomes, each originally defined by a centromere-linked gene unlinked to previously defined centromere markers. We examined four genes, cly2, KRB1, AMY2, and tsm0115, each centromere linked, but previously thought to be not on chromosomes I to XVI. We found that AMY2 is linked to cly2, and both are on chromosome II. tsm0115 is on the left arm of chromosome XVI. We confirm the earlier evidence that KRB1 is not on chromosomes I through XVI. This gene thus defines a new chromosome XVII. We also report meiotic linkage of met4 and pet8 (on chromosome XIV), confirming the connection between the petx-kex2 fragment of XIV and the centromere of XIV.     

The nucleotide sequence of the mitochondrial DNA genome of an abundant petite mutant of Saccharomyces cerevisiae carrying the ori1 replication origin

We have determined the 903 bp nucleotide sequence of the mitochondrial DNA genome of a Saccharomyces cerevisiae petite mutant BB5. This petite, containing the 265 nucleotide ori1 region, is representative of a class of petites arising at exceptionally high frequency within the population of spontaneous petites derived from a particular mit- strain Mb12. The DNA sequences of both the ori1 region and the flanking intergenic regions have been compared to those of the corresponding regions of mtDNA in a previously reported petite strain, a1/1R/1 of Bernardi's laboratory, that has a similar (880 bp) repeat unit. The BB5 petite genome carries a canonical ori1 sequence that is identical in both petite mtDNAs, but the flanking intergenic sequences show significant differences between the two petite strains. The divergence is considered to arise from differences in the sequences flanking ori1 in the respective parent strains.