Biomedical subjects

Serge Potier

Publications and source records attributed to Serge Potier.

13 recordsLinked to original sources

PubMed2005

Duplication processes in Saccharomyces cerevisiae haploid strains.

Duplication is thought to be one of the main processes providing a substrate on which the effects of evolution are visible. The mechanisms underlying this chromosomal rearrangement were investigated here in the yeast Saccharomyces cerevisiae. Spontaneous revertants containing a duplication event were selected and analyzed. In addition to the single gene duplication described in a previous study, we demonstrated here that direct tandem duplicated regions ranging from 5 to 90 kb in size can also occur spontaneously. To further investigate the mechanisms in the duplication events, we examined whether homologous recombination contributes to these processes. The results obtained show that the mechanisms involved in segmental duplication are RAD52-independent, contrary to those involved in single gene duplication. Moreover, this study shows that the duplication of a given gene can occur in S.cerevisiae haploid strains via at least two ways: single gene or segmental duplication.

Aspartate Carbamoyltransferase↗

PubMed2005

An evolutionary scenario for one of the largest yeast gene families.

The DUP gene family of Saccharomyces cerevisiae comprises 23 members that can be divided into two subfamilies--DUP240 and DUP380. The location of the DUP loci suggests that at least three mechanisms were responsible for their genomic dispersion: nonreciprocal translocation at chromosomal ends, tandem duplication and Ty-associated duplication. The data we present here suggest that these nonessential genes encode proteins that facilitate membrane trafficking processes. Dup240 proteins have three conserved domains (C1, C2 and C3) and two predicted transmembrane segments (H1 and H2). A direct repetition of the C1-H1-H2-C2 module is observed in Dup380p sequences. In this article, we propose an evolutionary model to account for the emergence of the two gene subfamilies.

Amino Acid Sequence↗

PubMed2005

Paleogenomics or the search for remnant duplicated copies of the yeast DUP240 gene family in intergenic areas.

Duplication, resulting in gene redundancy, is well known to be a driving force of evolutionary change. Gene families are therefore useful targets for approaching genome evolution. To address the gene death process, we examined the fate of the 10-member-large S288C DUP240 family in 15 Saccharomyces cerevisiae strains. Using an original three-step method of analysis reported here, both slightly and highly degenerate DUP240 copies, called pseudo-open reading frames (ORFs) and relics, respectively, were detected in strain S288C. It was concluded that two previously annotated ORFs correspond, in fact, to pseudo-ORFs and three additional relics were identified in intergenic areas. Comparative intraspecies analysis of these degenerate DUP240 loci revealed that the two pseudo-ORFs are present in a nondegenerate state in some other strains. This suggests that within a given gene family different loci are the target of the gene erasure process, which is therefore strain dependent. Besides, the variable positions observed indicate that the relic sequence may diverge faster than the flanking regions. All in all, this study shows that short conserved protein motifs provide a useful tool for detecting and accurately mapping degenerate gene remnants. The present results also highlight the strong contribution of comparative genomics for gene relic detection because the possibility of finding short conserved protein motifs in intergenic regions (IRs) largely depends on the choice of the most closely related paralog or ortholog. By mapping new genetic components in previously annotated IRs, our study constitutes a further refinement step in the crucial stage of genome annotation and provides a strategy for retracing ancient chromosomal reshaping events and, hence, for deciphering genome history.

Amino Acid Sequence↗

PubMed2004

Genome evolution in yeasts.

Identifying the mechanisms of eukaryotic genome evolution by comparative genomics is often complicated by the multiplicity of events that have taken place throughout the history of individual lineages, leaving only distorted and superimposed traces in the genome of each living organism. The hemiascomycete yeasts, with their compact genomes, similar lifestyle and distinct sexual and physiological properties, provide a unique opportunity to explore such mechanisms. We present here the complete, assembled genome sequences of four yeast species, selected to represent a broad evolutionary range within a single eukaryotic phylum, that after analysis proved to be molecularly as diverse as the entire phylum of chordates. A total of approximately 24,200 novel genes were identified, the translation products of which were classified together with Saccharomyces cerevisiae proteins into about 4,700 families, forming the basis for interspecific comparisons. Analysis of chromosome maps and genome redundancies reveal that the different yeast lineages have evolved through a marked interplay between several distinct molecular mechanisms, including tandem gene repeat formation, segmental duplication, a massive genome duplication and extensive gene loss.

Chromosomes, Fungal↗

PubMed2004

Integrated allosteric regulation in the S. cerevisiae carbamylphosphate synthetase - aspartate transcarbamylase multifunctional protein.

BACKGROUND: The S. cerevisiae carbamylphosphate synthetase - aspartate transcarbamylase multifunctional protein catalyses the first two reactions of the pyrimidine pathway. In this organism, these two reactions are feedback inhibited by the end product UTP. In the present work, the mechanisms of these integrated inhibitions were studied. RESULTS: The results obtained show that the inhibition is competitive in the case of carbamylphosphate synthetase and non-competitive in the case of aspartate transcarbamylase. They also identify the substrate whose binding is altered by this nucleotide and the step of the carbamylphosphate synthetase reaction which is inhibited. Furthermore, the structure of the domains catalyzing these two reactions were modelled in order to localize the mutations which, specifically, alter the aspartate transcarbamylase sensitivity to the feedback inhibitor UTP. Taken together, the results make it possible to propose a model for the integrated regulation of the two activities of the complex. UTP binds to a regulatory site located in the vicinity of the carbamylphosphate synthetase catalytic subsite which catalyzes the third step of this enzyme reaction. Through a local conformational change, this binding decreases, competitively, the affinity of this site for the substrate ATP. At the same time, through a long distance signal transmission process it allosterically decreases the affinity of the aspartate transcarbamylase catalytic site for the substrate aspartate. CONCLUSION: This investigation provides informations about the mechanisms of allosteric inhibition of the two activities of the CPSase-ATCase complex. Although many allosteric monofunctional enzymes were studied, this is the first report on integrated allosteric regulation in a multifunctional protein. The positions of the point mutations which specifically abolish the sensitivity of aspartate transcarbamylase to UTP define an interface between the carbamylphosphate synthetase and aspartate transcarbamylase domains, through which the allosteric signal for the regulation of aspartate transcarbamylase must be propagated.

Allosteric Regulation↗

PubMed2004

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.

Alcoholic Beverages↗

PubMed2004

Recovery of a function involving gene duplication by retroposition in Saccharomyces cerevisiae.

The duplication of DNA sequences contributes to genomic plasticity and is known to be one of the key factors responsible for evolution. The mechanisms underlying these rare events, which have been frequently mentioned by authors performing genomic analysis, have not yet been completely elucidated. These mechanisms were approached here in the yeast Saccharomyces cerevisiae, using a positive selection screen based on a particular mutated allele of the URA2 gene. Spontaneous revertants containing a duplication of the terminal part of the URA2 gene were selected and analyzed. Some important features of the duplicated regions, such as their chromosome location, size, and insertion sites, were characterized. The events selected correspond to a single inter- or intrachromosomal gene duplication process. The duplicated ATCase sequence is generally punctuated by a poly(A) tract and is always located in Ty1 sequences. In addition, the activation of a Ty1 transcription process increased the frequency of the duplication events. All in all, these data suggest that the duplication mechanism involves the reverse transcription of mRNA and the subsequent integration of the cDNA into a Ty1 area. The Ty1 elements and the retrotransposon-encoded function are key factors contributing to chromosomal reshaping. The genomic rearrangements described constitute experimental evidence for the recovery of a function involving duplication by retroposition.

3' Flanking Region↗

PubMed2004

Expansion and contraction of the DUP240 multigene family in Saccharomyces cerevisiae populations.

The influence of duplicated sequences on chromosomal stability is poorly understood. To characterize chromosomal rearrangements involving duplicated sequences, we compared the organization of tandem repeats of the DUP240 gene family in 15 Saccharomyces cerevisiae strains of various origins. The DUP240 gene family consists of 10 members of unknown function in the reference strain S288C. Five DUP240 paralogs on chromosome I and two on chromosome VII are arranged as tandem repeats that are highly polymorphic in copy number and sequence. We characterized DNA sequences that are likely involved in homologous or nonhomologous recombination events and are responsible for intra- and interchromosomal rearrangements that cause the creation and disappearance of DUP240 paralogs. The tandemly repeated DUP240 genes seem to be privileged sites of gene birth and death.

Base Sequence↗

PubMed2002

Role of the Nha1 antiporter in regulating K(+) influx in Saccharomyces cerevisiae.

NHA1 encodes a K(+) (Na(+))/H(+) antiporter in the plasma membrane of Saccharomyces cerevisiae. We report that cells expressing the NHA1 gene contained less K(+) than the mutant lacking the gene when grown without K(+) limitation. They also grew better at low K(+) and showed higher affinity of transport than the nha1 strain. In agreement with the function of an electroneutral cation/H(+) antiporter, the effect was only observed at acidic pH. The improved growth and transport depended on the presence of Trk1p (the main K(+) influx system) and did not require the product of TRK2. We propose that Nha1p regulates the potassium content of the cell and, as a consequence, can affect the activity of the main K(+) influx system (Trk1p).

Biological Transport↗

PubMed2002

Cation/H+ antiporters mediate potassium and sodium fluxes in Pichia sorbitophila. Cloning of the PsNHA1 and PsNHA2 genes and expression in Saccharomyces cerevisiae.

Pichia sorbitophila grows rapidly in the presence of very high NaCl concentrations. Under these conditions, even when the K(+) concentration is low, P. sorbitophila cells can maintain low Na(+) and high K(+) contents. This remarkable capacity of P. sorbitophila fails when the external pH is not acidic. This indicates that Na(+) efflux is mediated by an electroneutral Na(+)/H(+) antiporter. We have cloned and sequenced two genes designated as PsNHA1 and PsNHA2, which probably encode two antiporters of this type. The genes present high similarity with the corresponding genes from other yeasts. The heterologous expression of PsNHA1 or PsNHA2 in a Saccharomyces cerevisiae mutant lacking the Na(+) efflux systems and sensitive to high concentrations of Na(+) and K(+) rescued the tolerance and the ability to extrude both cations. The Accession Nos of the sequenced DNA fragments are: PsNHA1, AJ496431; PsNHA2, AJ496432. (TC 2.A.36)

Amino Acid Sequence↗

PubMed2002

Difference in substrate specificity divides the yeast alkali-metal-cation/H(+) antiporters into two subfamilies.

Yeast plasma membrane Na(+)/H(+) antiporters (TC 2.A.36) share a high degree of similarity at the protein level. Expression of four antiporters (Saccharomyces cerevisiae Nha1p, Candida albicans Cnh1p, Zygosaccharomyces rouxii ZrSod2-22p and Schizosaccharomyces pombe sod2p) in a SACCH: cerevisiae mutant strain lacking both Na(+)-ATPase and Na(+)/H(+) antiporter genes made it possible to study the transport properties and contribution to cell salt tolerance of all antiporters under the same conditions. The ZrSod2-22p of the osmotolerant yeast Z. rouxii has the highest transport capacity for lithium and sodium but, like the SCHIZ: pombe sod2p, it does not recognize K(+) and Rb(+) as substrates. The SACCH: cerevisiae Nha1p and C. albicans Cnh1p have a broad substrate specificity for at least four alkali metal cations (Na(+), Li(+), K(+), Rb(+)), but their contribution to overall cell tolerance to high external concentration of toxic Na(+) and Li(+) cations seems to be lower compared to the antiporters of SCHIZ: pombe and especially Z. rouxii.

Candida albicans↗

PubMed2002

Functional analysis of the Saccharomyces cerevisiae DUP240 multigene family reveals membrane-associated proteins that are not essential for cell viability.

The DUP240 gene family of Saccharomyces cerevisiae is composed of 10 members. They encode proteins of about 240 amino acids which contain two predicted transmembrane domains. Database searches identified only one homologue in the closely related species Saccharomyces bayanus, indicating that the DUP240 genes encode proteins specific to Saccharomyces sensu stricto. The short-flanking homology PCR gene-replacement strategy with a variety of selective markers for replacements, and classical genetic methods, were used to generate strains deleted for all 10 DUP240 genes. All of the knock-out strains were viable and had similar growth kinetics to the wild-type. Two-hybrid screens, hSos1p fusions and GFP fusions were carried out; the results indicated that the Dup240 proteins are membrane associated, and that some of them are concentrated around the plasma membrane.

Amino Acid Sequence↗

PubMed1998

The Nha1 antiporter of Saccharomyces cerevisiae mediates sodium and potassium efflux.

The NHA1 gene of Saccharomyces cerevisiae, transcribed into a 3.5 kb mRNA, encodes a protein mediating Na+ and K+ efflux through the plasma membrane that is required for alkali cation tolerance at acidic pH. Deletion of the gene in a wild-type strain resulted in higher sensitivity to both K+ and Na+ at acidic pH. Measurements of cation loss in strains carrying deleted or overexpressed alleles of NHA1 demonstrated its role in K+ and Na+ efflux. In addition, high K+ and Na+ efflux observed upon alkalinization of the cytoplasm implies a role of Nha1p in the regulation of intracellular pH. Moreover, the overexpression of ENA1 and NHA1 genes in an ena1-4 delta-nha1 delta strain showed that the Nha1 alkalication antiporter is responsible for growth on high concentrations of KCl and NaCl at acidic pH, and Ena alkali-cation ATPases are necessary at higher pH values. Both systems have a complementary action to maintain the intracellular steady-state concentration of K+ and Na+.

Adenosine Triphosphatases↗