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The breakdown of lignin by fungi is a key step during carbon recycling in terrestrial ecosystems. This process is of great interest for green and white biotechnological applications. Given the importance of these enzymatic processes, we have classified the enzymes potentially involved in lignin catabolism into sequence-based families and integrated them in a newly developed database, designated Fungal Oxidative Lignin enzymes (FOLy). Families were defined after sequence similarity searches starting from protein sequences and validated by the convergence of results with biochemical experiments reported in the literature. The resulting database was applied as a tool for the functional annotation of genomes from different fungi, namely (i) the Basidiomycota Coprinopsis cinerea, Phanerochaete chrysosporium and Ustilago maydis and (ii) the Ascomycota Aspergillus nidulans and Trichoderma reesei. Genomic comparison of the oxidoreductases of these fungi revealed significant differences in the putative enzyme arsenals. Two Ascomycota fungal genomes were annotated and new candidate genes were identified that could be useful for lignin degradation and (or) melanin synthesis, and their function investigated experimentally. This database efforts aims at providing the means to get new insights for the understanding and biotechnological exploitation of the lignin degradation. A WWW server giving access to the routinely updated FOLy classifications of enzymes potentially involved in lignin degradation can be found at http://foly.esil.univ-mrs.fr.

Several recent studies have shown that genes specifically expressed by the oocyte are subject to rapid evolution, in particular via gene duplication mechanisms. In the present work, we have focused our attention on a family of genes, specific to eutherian mammals, that are located in unstable genomic regions. We have identified two genes specifically expressed in the mouse oocyte: Khdc1a (KH homology domain containing 1a, also named Ndg1 for Nur 77 downstream gene 1, a target gene of the Nur77 orphan receptor), and another gene structurally related to Khdc1a that we have renamed Khdc1b. In this paper, we show that Khdc1a and Khdc1b belong to a family of several members including the so-called developmental pluripotency A5 (Dppa5) genes, the cat/dog oocyte expressed protein (cat OOEP and dog OOEP) genes, and the ES cell-associated transcript 1 (Ecat1) genes. These genes encode structurally related proteins that are characterized by an atypical RNA-binding KH domain and are specifically expressed in oocytes and/or embryonic stem cells. They are absent in fish, bird, and marsupial genomes and thus seem to have first appeared in eutherian mammals, in which they have evolved rapidly. They are located in a single syntenic region in all mammalian genomes studied, except in rodents, in which a synteny rupture due to a paracentric inversion has separated this gene family into two genomic regions and seems to be associated with increased instability in these regions. Overall, we have identified and characterized a novel family of oocyte and/or embryonic stem cell-specific genes encoding proteins that share an atypical KH RNA-binding domain and that have evolved rapidly since their emergence in eutherian mammalian genomes.

By using in silico methods in a previous study, we identified 100 oocyte-specific genes and 150 genes, enriched in the mouse oocyte. Interestingly, approximately half of the oocyte-specific genes tend to cluster on mouse chromosomes as if they have recently duplicated during evolution. In this study, we focused our attention on mouse BRDT, which belongs to a family of four structurally related proteins characterized by two N-terminal bromodomains and one C-terminal extraterminal domain (ET domain), defining the BET family. In mammals, BRD2, -3, and -4 are ubiquitously expressed, whereas BRDT expression was shown to be restricted to the testis. We were interested to know whether there was a correlation between the evolutionary rate and the specificity of expression of these four paralogous genes. First, we show by RT-PCR and in situ hybridization that BRDT is also expressed in mouse oocyte. Moreover, phylogenetic analyses show that the BRDT germ cell-specific orthology group clearly evolves faster than its ubiquitously expressed paralogs BRD2, BRD3, and BRD4. This suggests that there is a relationship between the evolution of these four groups of orthology and their tissue specificity of expression.

In animals, the tetraspanins are a large superfamily of membrane proteins that play important roles in organizing various cell-cell and matrix-cell interactions and signal pathways based on such interactions. However, their origin and evolution largely remain elusive and most of the family's members are functionally unknown or less known due to difficulties of study, such as functional redundancy. In this study, we rebuilt the family's phylogeny with sequences retrieved from online databases and our cDNA library of amphioxus. We reveal that, in addition to in metazoans, various tetraspanins are extensively expressed in protozoan amoebae, fungi, and plants. We also discuss the structural evolution of tetraspanin's major extracellular domain and the relation between tetraspanin's duplication and functional redundancy. Finally, we elucidate the coevolution of tetraspanins and eukaryotes and suggest that tetraspanins play important roles in the unicell-to-multicell transition. In short, the study of tetraspanin in a phylogenetic context helps us understand the evolution of intercellular interactions.

Human families in which recombinant meiotic event(s) are known to have occurred are powerful tools with which to analyze more precisely the structures of defined genomic regions, especially unstable areas. Such families allow the determination of the haplotypes of each member and, taking into account the recombinant event, it is possible to localize very precisely the point of crossover. Using families in which crossovers between the genes HLA-A and -B have occurred, we have constructed a meiotic map localizing the meiotic breakpoint events with respect to both anonymous markers and the principal genes of the region. Such mapping, which depends on the direct analysis of genomic DNA, is essential for fine structural analysis and is a powerful means of verification of the order and the localization of markers: physical mapping alone, using yeast artificial chromosomes, presents some uncertainties due to the numerous chimeras and inversions that can be produced. The establishment of this map will allow us to determine efficiently the precise location for new markers already localized to the map region. Three microsatellites (D6S265, D6S276, and D6S306), localized in the HLA region by linkage analysis, have been precisely located with respect to the points of recombination in the class I region. The sites of meiotic recombination in the MHC class I region seem to be not randomly distributed but in the majority of cases occurred between HLA-C and the microsatellite D6S265. This study also shows two cases of abnormal segregation of alleles. We discuss how these mutations correspond to a spontaneous mutation event at the somatic or germinal level.

Human myelin oligodendrocyte glycoprotein (MOG), a myelin component of the central nervous system, is a candidate target antigen for autoimmune-mediated demyelination. We have isolated and sequenced part of a cosmid clone that contains the entire human MOG gene. The primary nuclear transcript, extending from the putative start of transcription to the site of poly(A) addition, is 15,561 nucleotides in length. The human MOG gene contains 8 exons, separated by 7 introns; canonical intron/exon boundary sites are observed at each junction. The introns vary in size from 242 to 6484 bp and contain numerous repetitive DNA elements, including 14 Alu sequences within 3 introns. Another Alu element is located in the 3'-untranslated region of the gene. Alu sequences were classified with respect to subfamily assignment. Seven hundred sixty-three nucleotides 5' of the transcription start and 1214 nucleotides 3' of the poly(A) addition sites were also sequenced. The 5'-flanking region revealed the presence of several consensus sequences that could be relevant in the transcription of the MOG gene, in particular binding sites in common with other myelin gene promoters. Two polymorphic intragenic dinucleotide (CA)n and tetranucleotide (TAAA)n repeats were identified and may provide genetic marker tools for association and linkage studies.

We have refined and extended the map of the distal half of the human major histocompatibility complex. The map is continuous from HLA-E to 1000 kb telomeric of HLA-F and includes six new markers and genes. In addition, the corresponding sequences that were not previously mapped in the mouse genome have been located. The human and the mouse organizations have therefore been compared. This comparison allows us to demonstrate that the structure of the distal part of the MHC is similar in the two species. In addition, this comparison shows the presence of a breakpoint of synteny telomeric of the distal part of the H-2 region. Indeed, the region telomeric of HLA in human is found on a chromosome different from that carrying H-2 in mouse. The mapping analysis of paralogous genes (structurally related genes) around the breakpoint shows that the human organization probably represents the putative human/mouse ancestral one. This evolutionary breakpoint was precisely mapped in human, and the surrounding region was cloned into yeast artificial chromosomes. Finally, we show that the region found around the breakpoint was involved several times in chromosome recombinations in the mouse lineage, as it seems to correspond also to the t-complex distal inversion point.

OTF3 (octamer transcription factor 3) is a transcription factor containing a POU-specific domain and a homeodomain that could play a role in early development. In situ hybridization and pairwise linkage analysis showed that OTF3 gene maps close to the human MHC (major histocompatibility complex). In this paper, we define its localization within the MHC, around 100 kb telomeric to HLA-C, using a combination of physical and genetic analyses.

We report the successful use of whole yeast artificial chromosomes (YACs) as probes for direct positional cloning of novel expressed sequences in a given genomic fragment. The class I region of the human major histocompatibility complex, in particular the chromosomal fragment spanning the HLA-E locus, was investigated. The screening of a cDNA library with a 210-kb-long YAC clone led to the identification of a new gene, positionally conserved in the major histocompatibility complex of the mouse genome and encoding a putative GTP binding protein. Although its precise function remains unknown, the interspecies conservation of both sequence and map position suggests a regulatory or functional link with the histocompatibility cluster.

CD28 and CTLA4 are structurally homologous single-V-domain molecules of the Ig superfamily, the genes of which comap on the same chromosomal bands in mouse and man. Using polymerase chain reactions, we isolated six yeast artificial chromosome (YAC) clones positive for CTLA4 and/or CD28 from a human-DNA-containing YAC library. Two double-positive clones, 365 and 550 kb long, respectively, were further studied. Detailed restriction enzyme maps showed that one of these YACs was nested in the other, that they both bore the same CD28- and CTLA4-hybridizing fragments, that similar fragments were seen in genomic DNA, and that the distance between the CD28 and CTLA4 genes was at most 150 kb and at least 25 kb. A CpG island was found between these genes. These results provide a high-resolution estimate of the physical distance between the CD28 and CTLA4 genes and constitute a basis for the isolation of neighboring structures.