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We have used a combination of freeze-substitution electron microscopy and specific reaction for polysaccharides to re-evaluate glycogen structures in Saccharomyces cerevisiae. We also used mutant cells devoid of glycogen to confirm the glycogenic nature of the structures described. Previously described cytoplasmic aggregates were confirmed as a glycogen granules. Moreover, an original structure was discovered. This is a ring of glycogen surrounding a finger- or pleat-like plasma membrane invagination. This structure could be physiologically significant in terms of channelling glucose to or from glycogen reserves.
Keywords:glycogen, yeast, Thiery reaction, plasma membrane, invaginations glycogenin, Saccharomyces cerevisiae, freeze-substitution

Photoluminescence (PL) experiments have been performed on self-organized PbSe/Pb1—xEuxTe (x = 0.05 to 0.1) quantum dot superlattices grown by molecular beam epitaxy (MBE). At a temperature of 1.7 K, several PL lines were observed in the spectral range from 200 to 230 meV with a relatively narrow width of typically 1 to 2 meV. These lines emerge on top of a weak, broader band. Comparison of the luminescence photon energy with that of bulk PbSe (146 meV) shows the strong influence of carrier confinement. Studies of the temperature and magnetic field dependence demonstrate that the narrow lines are not significantly shifted up to 50 K and 7 T, respectively, whereas the broader band exhibits a shift comparable to that of bulk PbSe material. Studies of the PL intensity as a function of excitation power and the preferred emission of radiation from the sample edge indicate stimulated emission.

Using genetic and flow cytometric analyses, we showed that wine strain S6U is an allotetraploid of S. cerevisiae× S. bayanus. Hybrid constitution of the strain and its meiotic segregants was confirmed by Southern hybridization analysis of their chromosomal DNAs using four S. cerevisiaecloned genes: LYS2(chr. II), TRK1 (chr. X), ARG4 (chr. VIII), ACT1 (chr. VI) and PCR/RFLP analysis of the MET2 gene (chr. XIV). Monosporic progeny of strain S6U was highly viable in first generation but completely non-viable in the second one. According to the genetic analysis, sherry strain S. cerevisiaeSBY 2592 was found to be an autotetraploid heterozygous for homo-heterothallism.

In this report, we have shown that the yeast amphiphysin-like protein Rvs167p was localized mainly in small cortical patches throughout the cell in unbudding cells. During budding, the patches were polarized at bud emergence site. During mating, Rvs167p was concentrated at the tip of the shmoo. Rvs167p colocalized with actin patches during yeast vegetative growth and mating. Complete disruption of the actin cytoskeleton using Latrunculin-A did not affect Rvs167p localization in patches throughout the cell. In rvs167 mutant cells, actin patches are mislocalized and in rvs161 or abp1 mutant cells, Rvs167p localization is not affected. These observations suggest that Rvs167p may localize the actin cortical complex properly. Finally, the amphiphysin-conserved N-terminal domain of Rvs167p, called the BAR domain, was required but not sufficient for the correct localization of the protein.

In the framework of the EU genome‐sequencing programmes, the complete DNA sequence of the yeast Saccharomyces cerevisiae chromosome II (807 188 bp) has been determined. At present, this is the largest eukaryotic chromosome entirely sequenced. A total of 410 open reading frames (ORFs) were identified, covering 72% of the sequence. Similarity searches revealed that 124 ORFs (30%) correspond to genes of known function, 51 ORFs (12.5%) appear to be homologues of genes whose functions are known, 52 others (12.5%) have homologues the functions of which are not well defined and another 33 of the novel putative genes (8%) exhibit a degree of similarity which is insufficient to confidently assign function. Of the genes on chromosome II, 37‐45% are thus of unpredicted function. Among the novel putative genes, we found several that are related to genes that perform differentiated functions in multicellular organisms of are involved in malignancy. In addition to a compact arrangement of potential protein coding sequences, the analysis of this chromosome confirmed general chromosome patterns but also revealed particular novel features of chromosomal organization. Alternating regional variations in average base composition correlate with variations in local gene density along chromosome II, as observed in chromosomes XI and III. We propose that functional ARS elements are preferably located in the AT‐rich regions that have a spacing of approximately 110 kb. Similarly, the 13 tRNA genes and the three Ty elements of chromosome II are found in AT‐rich regions. In chromosome II, the distribution of coding sequences between the two strands is biased, with a ratio of 1.3:1. An interesting aspect regarding the evolution of the eukaryotic genome is the finding that chromosome II has a high degree of internal genetic redundancy, amounting to 16% of the coding capacity.

In the framework of the EU genome‐sequencing programmes, the complete DNA sequence of the yeast Saccharomyces cerevisiae chromosome II (807 188 bp) has been determined. At present, this is the largest eukaryotic chromosome entirely sequenced. A total of 410 open reading frames (ORFs) were identified, covering 72% of the sequence. Similarity searches revealed that 124 ORFs (30%) correspond to genes of known function, 51 ORFs (12.5%) appear to be homologues of genes whose functions are known, 52 others (12.5%) have homologues the functions of which are not well defined and another 33 of the novel putative genes (8%) exhibit a degree of similarity which is insufficient to confidently assign function. Of the genes on chromosome II, 37‐45% are thus of unpredicted function. Among the novel putative genes, we found several that are related to genes that perform differentiated functions in multicellular organisms of are involved in malignancy. In addition to a compact arrangement of potential protein coding sequences, the analysis of this chromosome confirmed general chromosome patterns but also revealed particular novel features of chromosomal organization. Alternating regional variations in average base composition correlate with variations in local gene density along chromosome II, as observed in chromosomes XI and III. We propose that functional ARS elements are preferably located in the AT‐rich regions that have a spacing of approximately 110 kb. Similarly, the 13 tRNA genes and the three Ty elements of chromosome II are found in AT‐rich regions. In chromosome II, the distribution of coding sequences between the two strands is biased, with a ratio of 1.3:1. An interesting aspect regarding the evolution of the eukaryotic genome is the finding that chromosome II has a high degree of internal genetic redundancy, amounting to 16% of the coding capacity.

Abstract: When over-expressed, RTM yeast genes confer resistance to the toxicity of molasses. They are found in distiller's and baker's industrial yeasts in multiple copies, scattered on the telomeres and physically linked to the telomeric SUC genes. Because these genes are absent from some laboratory strains, we explored the genomes of other industrial yeasts (brewing strains) and wine wild strains. A collection of 47 wine yeast strains (S. cerevisiae and S. bayanus) and 15 brewing strains, lager, ale and possible ancestors (S. monacensis, S. paradoxus and S. carlsbergensis) were screened for the presence of RTM genes. Only three wine strains and all brewing strains proved to contain RTM sequences in different copy numbers. PCR and chromosome blotting confirm the presence of SUC sequences in tandem with RTM. Moreover, analysis of the entire S. cerevisiae genome sequence shows that three other, non-telomeric, genes related to RTM are scattered on different chromosomes.

Only a few yeast strains produce pectin‐degrading enzymes such as pectin esterases and depolymerases (hydrolases and lyases). Strain SCPP is the only known Saccharomycesstrain to produce these pectinases. One of these pectolytic enzymes, PGL1‐encoded endopolygalacturonase (EC 3.2.1.15), hydrolyses the α‐1,4‐glycosidic bonds within the rhamnogalacturonan chains in pectic substances. This paper presents the cloning and sequencing of the first S. cerevisiaegene involved in pectin degradation. Few differences were found between the two deduced amino acid sequences encoded by PGL1‐1from a pectolytic (PG+) strain (SCPP) and PGL1‐2from a non‐pectolytic (PG−) strain (X2180‐1B). Similarities were found with other polygalacturonases from plants and other microorganisms. Of the two S. cerevisiaegenes, only the one isolated from strain SCPP was able, by overexpression, to confer endopolygalacturonase activity to a laboratory strain of S. cerevisiae. Overexpression of PGL1‐1gene in a non‐pectolytic strain resulted in halo formation on polygalacturonic acid‐containing agar plates stained with ruthenium red. Copyright © 1999 John Wiley & Sons, Ltd.

Only a few yeast strains produce pectin-degrading enzymes such as pectin esterases and depolymerases (hydrolases and lyases). Strain SCPP is the only known Saccharomyces strain to produce these pectinases. One of these pectolytic enzymes, PGL1-encoded endopolygalacturonase (EC 3.2.1.15), hydrolyses the α-1,4-glycosidic bonds within the rhamnogalacturonan chains in pectic substances. This paper presents the cloning and sequencing of the first S. cerevisiae gene involved in pectin degradation. Few differences were found between the two deduced amino acid sequences encoded by PGL1-1 from a pectolytic (PG⁺ ) strain (SCPP) and PGL1-2 from a non-pectolytic (PG⁻ ) strain (X2180-1B). Similarities were found with other polygalacturonases from plants and other microorganisms. Of the two S. cerevisiae genes, only the one isolated from strain SCPP was able, by overexpression, to confer endopolygalacturonase activity to a laboratory strain of S. cerevisiae . Overexpression of PGL1-1 gene in a non-pectolytic strain resulted in halo formation on polygalacturonic acid-containing agar plates stained with ruthenium red. Copyright © 1999 John Wiley & Sons, Ltd.

Mutations in genes necessary for survival in stationary phase were isolated to understand the ability of wild-type Saccharomyces cerevisiae to remain viable during prolonged periods of nutritional deprivation. Here we report results concerning one of these mutants, rvs167, which shows reduced viability and abnormal cell morphology upon carbon and nitrogen starvation. The mutant exhibits the same response when cells are grown in high salt concentrations and other unfavorable growth conditions. The RVS167 gene product displays significant homology with the Rvs161 protein and contains a SH3 domain at the C-terminal end. Abnormal actin distribution is associated with the mutant phenotype. In addition, while the budding pattern of haploid strains remains axial in standard growth conditions, the budding pattern of diploid mutant strains is random. The gene RVS167 therefore could be implicated in cytoskeletal reorganization in response to environmental stresses and could act in the budding site selection mechanism.

Cells of Saccharomyces cerevisiae can choose a bud site in one of two different spatial patterns (axial or bipolar) determined by their mating type. Genes important for bud-site selection have been identified and a linear model describing the hierarchy of these genes was proposed. We have uncovered a new class of genes which is required only for the bipolar pattern. The phenotype of the corresponding mutants coupled with epistasis experiments with some budding mutants already described suggest the existence of specific genes for the bipolar pathway.

A modified cDNA of mature human apolipoprotein A-II (apoA-II) was expressed by a sterol-uptake yeast strain (erg 10) of Saccharomyces cerevisiæ. ApoA-II cDNA was fused with a modified yeast alpha factor leader peptide coding sequence under phosphoglycerate kinase promoter control in a 2 micron-based plasmid construction. The use of cholesterol allowed apoA-II secretion in the culture medium. The mono and dimeric forms of apoA-II (approximately 3 micrograms per ml) were detected.

The gag-myc oncogenic sequence of the avian retrovirus MC29 was first inserted in a multicopy expression vector allowing its expression in Saccharomyces cerevisiae. The oncogene transcripts were detected in yeast by Northern blot hybridization and gag-myc proteins were revealed by immunoprecipitation. On solid medium, the average size of gag-myc transformant colonies was smaller than control. In liquid cultures, the gag-myc strains had a doubling time of 4.7 h compared with 3.1 h in the controls. In one of the recipient strains, and after an initial transient period of 5 days, the gag-myc transformants became physiologically indistinguishable from control. In another recipient strain, the slow-growth phenotype is permanent. Plasmid instability is increased in gag-myc transformants. When a single copy of the gag-myc gene was inserted in a yeast chromosome, no phenotype was observed, showing that slow growth is the consequence of plasmid loss.

Three mutants of Saccharomyces cerevisiae resistant to triethyltin (an inhibitor of mitochondrial ATPase) on non-fermentative media, and non-resistant to this drug on fermentative media, were isolated and named TTR1, TTR2 and TTR3. Apart from triethyltin resistance, these mutants show the following common characteristics: (1) Increased intracellular cytochrome c concentration. (2) Increased respiration rate. (3) Decreased growth yield. (4) Increased growth sensitivity to several drugs inhibiting oxidative phosphorylation: namely, CCCP (permeabilizing inner mitochondrial membrane to protons), valinomycin (permeabilizing inner mitochondrial membrane to potassium) and oligomycin (inhibitor of mitochondrial ATPase). (5) Increased sensitivity to carbon source starvation. For each mutant, these characteristics appeared to be due to a single pleiotropic nuclear mutation. Mutation TTR1 causes additional phenotypic characteristics which do not appear in mutants TTR2 and TTR3: (1) Pinkish coloration of colonies which is more pronounced after a long growth period. (2) Inability of the cells to store glycogen. (3) Growth defect of the cells on a galactose-containing medium. (4) Inability of a diploid homozygote mutant strain to sporulate. All these phenotypic characteristics have already been described in yeast mutants deregulated in cAMP-dependant protein phosphorylation. Crossing of a strain bearing the TTR1 mutation with a strain mutated in the adenylate cyclase structural gene suggested that the TTR1 phenotype is due to a modification in regulation of cAPK by cAMP, making cell multiplication possible without intracellular cAMP.

An optically-pumped lead salt-based vertical cavity surface emitting laser is presented. The laser structure grown by molecular-beam epitaxy consists of a Pb0.99Eu0.01Te/PbTe λ/2 microcavity with Pb0.99Eu0.01Te/Pb0.94Eu0.06Te Bragg mirrors. Stimulated emission was observed at 6073 nm below 25 K.

We have used a combination of freeze-substitution electron microscopy and specific reaction for polysaccharides to re-evaluate glycogen structures in Saccharomyces cerevisiae. We also used mutant cells devoid of glycogen to confirm the glycogenic nature of the structures described. Previously described cytoplasmic aggregates were confirmed as a glycogen granules. Moreover, an original structure was discovered. This is a ring of glycogen surrounding a finger- or pleat-like plasma membrane invagination. This structure could be physiologically significant in terms of channelling glucose to or from glycogen reserves. Keywords:glycogen, yeast, Thiery reaction, plasma membrane, invaginations glycogenin, Saccharomyces cerevisiae, freeze-substitution