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MetA and MetX are phylogenetically unrelated families of acyl-L-homoserine transferases. Experimental assignation of function and structural modeling of these families correct widespread misannotation, reveal convergence of function and uncover new functions in a subclass of MetX. Experimental validation of enzyme function is crucial for genome interpretation, but it remains challenging because it cannot be scaled up to accommodate the constant accumulation of genome sequences. We tackled this issue for the MetA and MetX enzyme families, phylogenetically unrelated families of acyl-L-homoserine transferases involved in L-methionine biosynthesis. Members of these families are prone to incorrect annotation because MetX and MetA enzymes are assumed to always use acetyl-CoA and succinyl-CoA, respectively. We determined the enzymatic activities of 100 enzymes from diverse species, and interpreted the results by structural classification of active sites based on protein structure modeling. We predict that >60% of the 10,000 sequences from these families currently present in databases are incorrectly annotated, and suggest that acetyl-CoA was originally the sole substrate of these isofunctional enzymes, which evolved to use exclusively succinyl-CoA in the most recent bacteria. We also uncovered a divergent subgroup of MetX enzymes in fungi that participate only in L-cysteine biosynthesis as O-succinyl-L-serine transferases.

The exponential increase in genome sequencing output has led to the accumulation of thousands of predicted genes lacking a proper functional annotation. Among this mass of hypothetical proteins, enzymes catalyzing new reactions or using novel ways to catalyze already known reactions might still wait to be identified. Here, we provide a structural and biochemical characterization of the 3-keto-5-aminohexanoate cleavage enzyme (Kce), an enzymatic activity long known as being involved in the anaerobic fermentation of lysine but whose catalytic mechanism has remained elusive so far. Although the enzyme shows the ubiquitous triose phosphate isomerase (TIM) barrel fold and a Zn2+ cation reminiscent of metal-dependent class II aldolases, our results based on a combination of x-ray snapshots and molecular modeling point to an unprecedented mechanism that proceeds through deprotonation of the 3-keto-5-aminohexanoate substrate, nucleophilic addition onto an incoming acetyl-CoA, intramolecular transfer of the CoA moiety, and final retro-Claisen reaction leading to acetoacetate and 3-aminobutyryl-CoA. This model also accounts for earlier observations showing the origin of carbon atoms in the products, as well as the absence of detection of any covalent acyl-enzyme intermediate. Kce is the first representative of a large family of prokaryotic hypothetical proteins, currently annotated as the “domain of unknown function” DUF849.

Aldosterone and vasopressin are responsible for the final adjustment of sodium and water reabsorption in the kidney. In principal cells of the kidney cortical collecting duct (CCD), the integral response to aldosterone and the long-term functional effects of vasopressin depend on transcription. In this study, we analyzed the transcriptome of a highly differentiated mouse clonal CCD principal cell line (mpkCCD cl4 ) and the changes in the transcriptome induced by aldosterone and vasopressin. Serial analysis of gene expression (SAGE) was performed on untreated cells and on cells treated with either aldosterone or vasopressin for 4 h. The transcriptomes in these three experimental conditions were determined by sequencing 169,721 transcript tags from the corresponding SAGE libraries. Limiting the analysis to tags that occurred twice or more in the data set, 14,654 different transcripts were identified, 3,642 of which do not match known mouse sequences. Statistical comparison (at P < 0.05 level) of the three SAGE libraries revealed 34 AITs (aldosterone-induced transcripts), 29 ARTs (aldosterone-repressed transcripts), 48 VITs (vasopressin-induced transcripts) and 11 VRTs (vasopressin-repressed transcripts). A selection of the differentially-expressed, hormone-specific transcripts (5 VITs, 2 AITs and 1 ART) has been validated in the mpkCCD cl4 cell line either by Northern blot hybridization or reverse transcription–PCR . The hepatocyte nuclear transcription factor HNF-3-α (VIT39), the receptor activity modifying protein RAMP3 (VIT48), and the glucocorticoid-induced leucine zipper protein (GILZ) (AIT28) are candidate proteins playing a role in physiological responses of this cell line to vasopressin and aldosterone.

This paper reports the genomic analysis of strain CBS732 of Zygosaccharomyces rouxii, a homothallic diploid yeast. We explored the sequences of 4934 random sequencing tags of about 1 kb in size and compared them to the Saccharomyces cerevisiae gene products. Approximately 2250 nuclear genes, 57 tRNAs, the rDNA locus, the endogenous pSR1 plasmid and 15 mitochondrial genes were identified. According to 18S and 25S rRNA cladograms and to synteny analysis, Z. rouxii could be placed among the S. cerevisiae sensu lato yeasts.

As part of the comparative genomics project ‘GENOLEVURES’, we studied the Kluyveromyces marxianus var. marxianus strain CBS712 using a partial random sequencing strategy. With a 0.2×genome equivalent coverage, we identified ca. 1300 novel genes encoding proteins, some containing spliceosomal introns with consensus splice sites identical to those of Saccharomyces cerevisiae, 28 tRNA genes, the whole rDNA repeat, and retrotransposons of the Ty1/2 family of S. cerevisiae with diverged Long Terminal Repeats. Functional classification of the K. marxianus genes, as well as the analysis of the paralogous gene families revealed few differences with respect to S. cerevisiae. Only 42 K. marxianus identified genes are without detectable homolog in the baker’s yeast. However, we identified several genetic rearrangements between these two yeast species.

We have analyzed the evolution of chromosome maps of Hemiascomycetes by comparing gene order and orientation of the 13 yeast species partially sequenced in this program with the genome map of Saccharomyces cerevisiae. From the analysis of nearly 8000 situations in which two distinct genes having homologs in S. cerevisiae could be identified on the sequenced inserts of another yeast species, we have quantified the loss of synteny, the frequency of single gene deletion and the occurrence of gene inversion. Traces of ancestral duplications in the genome of S. cerevisiae could be identified from the comparison with the other species that do not entirely coincide with those identified from the comparison of S. cerevisiae with itself. From such duplications and from the correlation observed between gene inversion and loss of synteny, a model is proposed for the molecular evolution of Hemiascomycetes. This model, which can possibly be extended to other eukaryotes, is based on the reiteration of events of duplication of chromosome segments, creating transient merodiploids that are subsequently resolved by single gene deletion events.

The generation of sequencing data for the hemiascomycetous yeast random sequence tag project was performed using the procedures established at GENOSCOPE. These procedures include a series of protocols for the sequencing reactions, using infra-red labelled primers, performed on both ends of the plasmid inserts in the same reaction tube, and their analysis on automated DNA sequencers. They also include a package of computer programs aimed at detecting potential assignation errors, selecting good quality sequences and estimating their useful length.

The identification of molecular evolutionary mechanisms in eukaryotes is approached by a comparative genomics study of a homogeneous group of species classified as Hemiascomycetes. This group includes Saccharomyces cerevisiae, the first eukaryotic genome entirely sequenced, back in 1996. A random sequencing analysis has been performed on 13 different species sharing a small genome size and a low frequency of introns. Detailed information is provided in the 20 following papers. Additional tables available on websites describe the ca. 20 000 newly identified genes. This wealth of data, so far unique among eukaryotes, allowed us to examine the conservation of chromosome maps, to identify the ‘yeast-specific’ genes, and to review the distribution of gene families into functional classes. This project conducted by a network of seven French laboratories has been designated ‘Genolevures’.

Random sequencing of the Kluyveromyces lactis genome allowed the identification of 2235–2601 open reading frames (ORFs) homologous to S. cerevisiae ORFs, 51 ORFs which were homologous to genes from other species, 64 tRNAs, the complete rDNA repeat, and a few Ty1- and Ty2-like sequences. In addition, the complete sequence of plasmid pKD1 and a large coverage of the mitochondrial genome were obtained. The global distribution into general functional categories found in Saccharomyces cerevisiae and as defined by MIPS is well conserved in K. lactis. However, detailed examination of certain subcategories revealed a small excess of genes involved in amino acid metabolism in K. lactis. The sequences are deposited at EMBL under the accession numbers AL424881–AL430960.

Root-knot nematodes are globally the most aggressive and damaging plant-parasitic nematodes. Chemical nematicides have so far constituted the most efficient control measures against these agricultural pests. Because of their toxicity for the environment and danger for human health, these nematicides have now been banned from use. Consequently, new and more specific control means, safe for the environment and human health, are urgently needed to avoid worldwide proliferation of these devastating plant-parasites. Mining the genomes of root-knot nematodes through an evolutionary and comparative genomics approach, we identified and analyzed 15,952 nematode genes conserved in genomes of plant-damaging species but absent from non target genomes of chordates, plants, annelids, insect pollinators and mollusks. Functional annotation of the corresponding proteins revealed a relative abundance of putative transcription factors in this parasite-specific set compared to whole proteomes of root-knot nematodes. This may point to important and specific regulators of genes involved in parasitism. Because these nematodes are known to secrete effector proteins in planta, essential for parasitism, we searched and identified 993 such effector-like proteins absent from non-target species. Aiming at identifying novel targets for the development of future control methods, we biologically tested the effect of inactivation of the corresponding genes through RNA interference. A total of 15 novel effector-like proteins and one putative transcription factor compatible with the design of siRNAs were present as non-redundant genes and had transcriptional support in the model root-knot nematode Meloidogyne incognita. Infestation assays with siRNA-treated M. incognita on tomato plants showed significant and reproducible reduction of the infestation for 12 of the 16 tested genes compared to control nematodes. These 12 novel genes, showing efficient reduction of parasitism when silenced, constitute promising targets for the development of more specific and safer control means., Author Summary Plant-parasitic nematodes are annually responsible for more than $100 billion crop yield loss worldwide and those considered as causing most of the damages are root-knot nematodes. These nematodes used to be controlled by chemicals that are now banned from use because of their poor specificity and high toxicity for the environment and human health. In the absence of sustainable alternative solutions, new control means, more specifically targeted against these nematodes and safe for the environment are needed. We searched in root-knot nematode genomes, genes conserved in various plant-damaging species while otherwise absent from the genomes of non target species such as those of chordates, plants, annelids, insect pollinators and mollusks. These genes are probably important for plant parasitism and their absence from non-target species make them interesting candidates for the development of more specific and safer control means. Further bioinformatics pruning of this set of genes yielded 16 novel candidates that could be biologically tested. Using RNA interference, we knocked down each of these 16 genes in a root-knot nematode and tested the effect on plant parasitism efficiency. Out of the 16 tested genes, 12 showed a significant and reproducible diminution of infestation when silenced and are thus particularly promising.

Motivation: Current computational approaches to function prediction are mostly based on protein sequence classification and transfer of annotation from known proteins to their closest homologous sequences relying on the orthology concept of function conservation. This approach suffers a major weakness: annotation reliability depends on global sequence similarity to known proteins and is poorly efficient for enzyme superfamilies that catalyze different reactions. Structural biology offers a different strategy to overcome the problem of annotation by adding information about protein 3D structures. This information can be used to identify amino acids located in active sites, focusing on detection of functional polymorphisms residues in an enzyme superfamily. Structural genomics programs are providing more and more novel protein structures at a high-throughput rate. However, there is still a huge gap between the number of sequences and available structures. Computational methods, such as homology modeling provides reliable approaches to bridge this gap and could be a new precise tool to annotate protein functions. Results: Here, we present Active Sites Modeling and Clustering (ASMC) method, a novel unsupervised method to classify sequences using structural information of protein pockets. ASMC combines homology modeling of family members, structural alignment of modeled active sites and a subsequent hierarchical conceptual classification. Comparison of profiles obtained from computed clusters allows the identification of residues correlated to subfamily function divergence, called specificity determining positions. ASMC method has been validated on a benchmark of 42 Pfam families for which previous resolved holo-structures were available. ASMC was also applied to several families containing known protein structures and comprehensive functional annotations. We will discuss how ASMC improves annotation and understanding of protein families functions by giving some specific illustrative examples on nucleotidyl cyclases, protein kinases and serine proteases. Availability: http://www.genoscope.fr/ASMC/. Contact: raquelcm@dcc.ufmg.br; kbastard@genoscope.cns.fr; artigue@genoscope.cns.fr Supplementary information: Supplementary data are available at Bioinformatics online.

Single nucleotide polymorphism (SNP) markers have been shown to be useful in genetic investigations of medically important parasites and their hosts. In this paper, we describe the prediction and validation of SNPs in ESTs of Schistosoma mansoni. We used 107,417 public sequences of S. mansoni and identified 15,614 high-quality candidate SNPs in 12,184 contigs. The presence of predicted SNPs was observed in well characterized antigens and vaccine candidates such as those coding for myosin; Sm14 and Sm23; cathepsin B and triosephosphate isomerase (TPI). Additionally, SNPs were experimentally validated for the cathepsin B. A comparative model of the S. mansoni cathepsin B was built for predicting the possible consequences of amino acid substitutions on the protein structure. An analysis of the substitutions indicated that the amino acids were mostly located on the surface of the molecule, and we found no evidence for a significant conformational change of the enzyme. However, at least one of the substitutions could result in a structural modification of an epitope.

In most bilaterian organisms so far studied, Hox genes are organized in genomic clusters and determine development along the anteroposterior axis. It has been suggested that this clustering, together with spatial and temporal colinearity of gene expression, represents the ancestral condition. However, in organisms with derived modes of embryogenesis and lineage-dependent mechanisms for the determination of cell fate, temporal colinearity of expression can be lost and Hox cluster organization disrupted, as is the case for the ecdysozoans Drosophila melanogaster and Caenorhabditis elegans and the urochordates Ciona intestinalis and Oikopleura dioica. We sought to determine whether a lophotrochozoan, the platyhelminth parasite Schistosoma mansoni, possesses a conserved or disrupted Hox cluster. Using a polymerase chain reaction (PCR)-based strategy, we have cloned and characterized three novel S. mansoni genes encoding orthologues of Drosophila labial (SmHox 1), deformed (SmHox4), and abdominal A (SmHox8), as well as the full-length coding sequence of the previously described Smox1, which we identify as an orthologue of fushi tarazu. Quantitative reverse transcriptase-PCR showed that the four genes were expressed at all life-cycle stages but that levels of expression were differentially regulated. Phylogenetic analysis and the conservation of "parapeptide'' sequences ('"-terminal to the homeodomains of SmHox8 and Smox1 support the grouping of platyhelminths within the lophotrochozoan clade. However, Bacterial Artificial Chromosome (BAC) library screening followed by genome walking failed to reconstitute a cluster. The BAC clones containing Hox genes were sequenced. and in no case were other Hox genes found on the same clone. Moreover, the SmHox4 and SmHox8 genes contained single very large introns (>40 kbp) further indicating that the schistosome Hox cluster is highly extended. Localization of the Hox genes to chromosomes using fluorescence in situ hybridization showed that SinHox4 and SmHox8 are on the long arm of chromosome 4, whereas SmHox1 and Smox1 are on chromosome 3. In silico screening of the available genome sequences corroborated results of Southern blotting and BAC library screening that indicate that there are no paralogues of SmHox1, SmHox4, or SmHox8. The schistosome Hox cluster is therefore not duplicated, but is both dispersed and disintegrated in the genome.

Saccharomyces bayanus var. uvarum investigated here is the species closest to Saccharomyces cerevisiae. Random sequence tags (RSTs) allowed us to identify homologues to 2789 open reading frames (ORFs) in S. cerevisiae, ORFs duplicated in S. uvarum but not in S. cerevisiae, centromeres, tRNAs, homologues of Ty1/2 and Ty4 retrotransposons, and a complete rDNA repeat. Only 13 RSTs seem to be homologous to sequences in other organisms but not in S. cerevisiae. As the synteny between the two species is very high, cases in which synteny is lost suggest special mechanisms of genome evolution. The corresponding RSTs revealed that S. uvarum can exist without any S. cerevisiae DNA introgression. Accession numbers are from AL397139 to AL402278 in the EMBL databank.

A genomic exploration of Kluyveromyces thermotolerans was performed by random sequence tag (RST) analysis. We sequenced 2653 RSTs corresponding to inserts sequenced from both ends. We performed a systematic comparison with a complete set of proteins from Saccharomyces cerevisiae, other completely sequenced genomes and SwissProt. We identified six mitochondrial genes and 1358–1496 nuclear genes by comparison with S. cerevisiae. In addition, 25 genes were identified by comparison with other organisms. This corresponds to about 24% of the estimated gene content of this organism. A lower level of conservation is observed with orthologues to genes of S. cerevisiae previously classified as orphans. Gene order was found to be conserved between S. cerevisiae and K. thermotolerans in 56.5% of studied cases.

We explored the biological diversity of hemiascomycetous yeasts using a set of 22 000 newly identified genes in 13 species through BLASTX searches. Genes without clear homologue in Saccharomyces cerevisiae appeared to be conserved in several species, suggesting that they were recently lost by S. cerevisiae. They often identified well-known species-specific traits. Cases of gene acquisition through horizontal transfer appeared to occur very rarely if at all. All identified genes were ascribed to functional classes. Functional classes were differently represented among species. Species classification by functional clustering roughly paralleled rDNA phylogeny. Unequal distribution of rapidly evolving, ascomycete-specific, genes among species and functions was shown to contribute strongly to this clustering. A few cases of gene family amplification were documented, but no general correlation could be observed between functional differentiation of yeast species and variations of gene family sizes. Yeast biological diversity seems thus to result from limited species-specific gene losses or duplications, and for a large part from rapid evolution of genes and regulatory factors dedicated to specific functions.

Random sequence tags were obtained from a genomic DNA library of Saccharomyces exiguus. The mitochondrial genome appeared to be at least 25.7 kb in size, with a different organization compared to Saccharomyces cerevisiae. An unusual putative 953 bp long terminal repeated element associated to Ty3 was found. A set of 1451 genes was identified homologous to S. cerevisiae open reading frames. Only five genes were identified outside the S. cerevisiae taxon, confirming that S. exiguus is phylogenetically closely related to S. cerevisiae. Unexpectedly, numerous duplicated genes were found whereas they are unique in S. cerevisiae. The sequences are deposited at EMBL under the accession numbers: AL407377–AL409955.

The genome of Saccharomyces servazzii was analyzed with 2570 random sequence tags totalling 2.3 Mb. BLASTX comparisons revealed a minimum of 1420 putative open reading frames with significant homology to Saccharomyces cerevisiae (58% aa identity on average), two with Schizosaccharomyces pombe and one with a human protein, confirming that S. servazzii is closely related to S. cerevisiae. About 25% of the S. servazzii genes were identified, assuming that the gene complement is identical in both yeasts. S. servazzii carries very few transposable elements related to Ty elements in S. cerevisiae. Most of the mitochondrial genes were identified in eight contigs altogether spanning 25 kb for a predicted size of 29 kb. A significant match with the Kluyveromyces lactis linear DNA plasmid pGKL-1 encoded RF4 killer protein suggests that a related plasmid exists in S. servazzii. The sequences have been deposited with EMBL under the accession numbers AL402279–AL404848.

By analyzing 2830 random sequence tags (RSTs), totalling 2.7 Mb, we explored the genome of the marine, osmo- and halotolerant yeast, Debaryomyces hansenii. A contig 29 kb in length harbors the entire mitochondrial genome. The genes encoding Cox1, Cox2, Cox3, Cob, Atp6, Atp8, Atp9, several subunits of the NADH dehydrogenase complex 1 and 11 tRNAs were unambiguously identified. An equivalent number of putative transposable elements compared to Saccharomyces cerevisiae were detected, the majority of which are more related to higher eukaryote copia elements. BLASTX comparisons of RSTs with databases revealed at least 1119 putative open reading frames with homology to S. cerevisiae and 49 to other genomes. Specific functions, including transport of metabolites, are clearly over-represented in D. hansenii compared to S. cerevisiae, consistent with the observed difference in physiology of the two species. The sequences have been deposited with EMBL under the accession numbers AL436045–AL438874.

A total of 4940 random sequence tags of the dimorphic yeast Yarrowia lipolytica , totalling 4.9 Mb, were analyzed. BLASTX comparisons revealed at least 1229 novel Y. lipolytica genes 1083 genes having homology with Saccharomyces cerevisiae genes and 146 with genes from various other genomes. This confirms the rapid sequence evolution assumed for Y. lipolytica . Functional analysis of newly discovered genes revealed that several enzymatic activities were increased compared to S. cerevisiae , in particular, transport activities, ion homeostasis, and various metabolism pathways. Most of the mitochondrial genes were identified in contigs spanning more than 47 kb. Matches to retrotransposons were observed, including a S. cerevisiae Ty3 and a LINE element. The sequences have been deposited with EMBL under the accession numbers AL409956 – AL414895 .

A method is described for mutagenesis of Pseudomonas fluorescens strains by electroporation with the transposon delivery vector pUT/mini-Tn5 Km. The transposition process was shown to be optimal at 12.5 kV cm-1 for a pulse time (Bowen and Koslak, 1992) of about 4 ms. The Pseudomonas fluorescens L6.5 target strain exhibited maximal electrocompetence when harvested at the middle of the exponential growth phase. As many as 7.7 10(5) mutants per picomole of delivery vector (7.5 kb) could be obtained, and these kanamycin-resistant mutants were shown to have lost the pUT plasmid. By external calibration with plasmids of increasing size (from 11.5 to 60.1 kb), the efficiency of the transformation process was evaluated to be approximately 1.31 x 10(8) transformants per picomole of delivery vector. Efficiency of the transposition process was 0.58%. This rapid method was used to tag for the cloning three independent chromosomal loci responsible for the Alk+ phenotype of Pseudomonas fluorescens L6.5 strain.

The genome of Saccharomyces kluyveri was explored through 2528 random sequence tags with an average length of 981 bp. The complete nuclear ribosomal DNA unit was found to be 8656 bp in length. Sequences homologous to retroelements of the gypsy and copia types were identified as well as numerous solo long terminal repeats. We identified at least 1406 genes homologous to Saccharomyces cerevisiae open reading frames, with on average 58.1% and 72.4% amino acid identity and similarity, respectively. In addition, by comparison with completely sequenced genomes and the SwissProt database, we found 27 novel S. kluyveri genes. Most of these genes belong to pathways or have functions absent from S. cerevisiae , such as the catabolic pathway of purines or pyrimidines, melibiose fermentation, sorbitol utilization, or degradation of pollutants. The sequences are deposited in EMBL under the accession numbers AL404849 – AL407376 .

As part of a comparative genomics project on 13 hemiascomycetous yeasts, the Pichia angusta type strain was studied using a partial random sequencing strategy. With coverage of 0.5 genome equivalents, about 2500 novel protein-coding genes were identified, probably corresponding to more than half of the P. angusta protein-coding genes, 6% of which do not have homologs in Saccharomyces cerevisiae. Some of them contain one or two introns, on average three times shorter than those in S. cerevisiae. We also identified 28 tRNA genes, a few retrotransposons similar to Ty5 of S. cerevisiae, solo long terminal repeats, the whole ribosomal DNA cluster, and segments of mitochondrial DNA. The P. angusta sequences were deposited in EMBL under the accession numbers AL430961 to AL436044.

Comparisons of the 6213 predicted Saccharomyces cerevisiae open reading frame (ORF) products with sequences from organisms of other biological phyla differentiate genes commonly conserved in evolution from ‘maverick’ genes which have no homologue in phyla other than the Ascomycetes. We show that a majority of the ‘maverick’ genes have homologues among other yeast species and thus define a set of 1892 genes that, from sequence comparisons, appear ‘Ascomycetes-specific’. We estimate, retrospectively, that the S. cerevisiae genome contains 5651 actual protein-coding genes, 50 of which were identified for the first time in this work, and that the present public databases contain 612 predicted ORFs that are not real genes. Interestingly, the sequences of the ‘Ascomycetes-specific’ genes tend to diverge more rapidly in evolution than that of other genes. Half of the ‘Ascomycetes-specific’ genes are functionally characterized in S. cerevisiae, and a few functional categories are over-represented in them.

We have evaluated the degree of gene redundancy in the nuclear genomes of 13 hemiascomycetous yeast species. Saccharomyces cerevisiae singletons and gene families appear generally conserved in these species as singletons and families of similar size, respectively. Variations of the number of homologues with respect to that expected affect from 7 to less than 24% of each genome. Since S. cerevisiae homologues represent the majority of the genes identified in the genomes studied, the overall degree of gene redundancy seems conserved across all species. This is best explained by a dynamic equilibrium resulting from numerous events of gene duplication and deletion rather than by a massive duplication event occurring in some lineages and not in others.

The genome of the diploid hemiascomycetous yeast Candida tropicalis, an opportunistic human pathogen and an important organism for industrial applications, was explored by the analysis of 2541 Random Sequenced Tags (RSTs) covering about 20% of its genome. Comparison of these sequences with Saccharomyces cerevisiae and other species permitted the identification and the analysis of a total of more than 1000 novel genetic elements of C. tropicalis. Moreover, the present study confirms that in C. tropicalis, the rare CUG codon is read as a serine and not a leucine. The sequences have been deposited at EMBL with the accession numbers AL438875–AL441602.

This paper reports the genomic analysis of the strain CBS7064 of Pichia sorbitophila, a homothallic diploid yeast. We sequenced 4829 random sequence tags of about 1 kb and compared them to the Saccharomyces cerevisiae gene products. Approximately 1300 nuclear genes, 22 tRNAs, the rDNA locus, and six mitochondrial genes have been identified. The analysis of the rDNA genes has permitted to classify this organism close to the Candida species. Accession numbers from AL414896 to AL419724 at EMBL databank.