Up-regulation of apoptotic- and cell survival-related gene pathways following exposures of western corn rootworm to B. thuringiensis crystalline pesticidal proteins in transgenic maize roots.

BACKGROUND: Resistance of pest insect species to insecticides, including B. thuringiensis (Bt) pesticidal proteins expressed by transgenic plants, is a threat to global food security. Despite the western corn rootworm, Diabrotica virgifera virgifera, being a major pest of maize and having populations showing increasing levels of resistance to hybrids expressing Bt pesticidal proteins, the cell mechanisms leading to mortality are not fully understood. RESULTS: Twenty unique RNA-seq libraries from the Bt susceptible D. v. virgifera inbred line Ped12, representing all growth stages and a range of different adult and larval exposures, were assembled into a reference transcriptome. Ten-day exposures of Ped12 larvae to transgenic Bt Cry3Bb1 and Gpp34/Tpp35Ab1 maize roots showed significant differential expression of 1055 and 1374 transcripts, respectively, compared to cohorts on non-Bt maize. Among these, 696 were differentially expressed in both Cry3Bb1 and Gpp34/Tpp35Ab1 maize exposures. Differentially-expressed transcripts encoded protein domains putatively involved in detoxification, metabolism, binding, and transport, were, in part, shared among transcripts that changed significantly following exposures to the entomopathogens Heterorhabditis bacteriophora and Metarhizium anisopliae. Differentially expressed transcripts in common between Bt and entomopathogen treatments encode proteins in general stress response pathways, including putative Bt binding receptors from the ATP binding cassette transporter superfamily. Putative caspases, pro- and anti-apoptotic factors, as well as endoplasmic reticulum (ER) stress-response factors were identified among transcripts uniquely up-regulated following exposure to either Bt protein. CONCLUSIONS: Our study suggests that the up-regulation of genes involved in ER stress management and apoptotic progression may be important in determining cell fate following exposure of susceptible D. v. virgifera larvae to Bt maize roots. This study provides novel insights into insect response to Bt intoxication, and a possible framework for future investigations of resistance mechanisms.

Biases of STRUCTURE software when exploring introduction routes of invasive species.

Population genetic methods are widely used to retrace the introduction routes of invasive species. The unsupervised Bayesian clustering algorithm implemented in STRUCTURE is amongst the most frequently used of these methods, but its ability to provide reliable information about introduction routes has never been assessed. We simulated microsatellite datasets to evaluate the extent to which the results provided by STRUCTURE were misleading for the inference of introduction routes. We focused on an invasion scenario involving one native and two independently introduced populations, because it is the sole scenario that can be rejected when obtaining a particular clustering with a STRUCTURE analysis at K = 2 (two clusters). Results were classified as "misleading" or "non-misleading". We investigated the influence of effective size, bottleneck severity and number of loci on the type and frequency of misleading results. We showed that misleading STRUCTURE results were obtained for 10% of all simulated datasets. Our results highlighted two categories of misleading output. The first occurs when the native population has a low level of diversity. In this case, the two introduced populations may be very similar, despite their independent introduction histories. The second category results from convergence issues in STRUCTURE for K = 2, with strong bottleneck severity and/or large numbers of loci resulting in high levels of differentiation between the three populations. Overall, the risk of being misled by STRUCTURE in the context of introduction routes inferences is moderate, but it is important to remain cautious when low genetic diversity or genuine multimodality between runs are involved.

Comparative venomics of Psyttalia lounsburyi and P. concolor, two olive fruit fly parasitoids: a hypothetical role for a GH1 ß-glucosidase.

Venom composition of parasitoid wasps attracts increasing interest - notably molecules ensuring parasitism success on arthropod pests - but its variation within and among taxa is not yet understood. We have identified here the main venom proteins of two braconid wasps, Psyttalia lounsburyi (two strains from South Africa and Kenya) and P. concolor, olive fruit fly parasitoids that differ in host range. Among the shared abundant proteins, we found a GH1 ß-glucosidase and a family of leucine-rich repeat (LRR) proteins. Olive is extremely rich in glycoside compounds that are hydrolyzed by ß-glucosidases into defensive toxic products in response to phytophagous insect attacks. Assuming that Psyttalia host larvae sequester ingested glycosides, the injected venom GH1 ß-glucosidase could induce the release of toxic compounds, thus participating in parasitism success by weakening the host. Venom LRR proteins are similar to truncated Toll-like receptors and may possibly scavenge the host immunity. The abundance of one of these LRR proteins in the venom of only one of the two P. lounsburyi strains evidences intraspecific variation in venom composition. Altogether, venom intra- and inter-specific variation in Psyttalia spp. were much lower than previously reported in the Leptopilina genus (Figitidae), suggesting it might depend upon the parasitoid taxa.

The LBP/BPI multigenic family in invertebrates: Evolutionary history and evidences of specialization in mollusks.

LBPs (lipopolysaccharide binding proteins) and BPIs (bactericidal permeability increasing proteins) are important proteins involved in defense against bacterial pathogens. We recently discovered a novel biocidal activity of a LBP/BPI from the gastropod Biomphalaria glabrata and demonstrated its role in parental immune protection of eggs, highlighting the importance of LBP/BPIs in invertebrate immunity. Here we characterize four additional LBP/BPI from B. glabrata, presenting conserved sequence architecture and exon-intron structure. Searches of invertebrate genomes revealed that existence of LBP/BPIs is not a conserved feature since they are absent from phyla such as arthropods and platyhelminths. Analyses of LBP/BPI transcripts from selected mollusk species showed recent parallel duplications in some species, including B. glabrata. In this snail species, LBP/BPI members vary in their expression tissue localization as well as their change in expression levels after immune challenges (Gram-negative bacterium; Gram-positive bacterium or yeast). These results, together with the predicted protein features provide evidences of functional specialization of LBP/BPI family members in molluscs.

Diversification of MIF immune regulators in aphids: link with agonistic and antagonistic interactions.

BACKGROUND: The widespread use of genome sequencing provided evidences for the high degree of conservation in innate immunity signalling pathways across animal phyla. However, the functioning and evolutionary history of immune-related genes remains unknown for most invertebrate species. A striking observation coming from the analysis of the pea aphid Acyrthosiphon pisum genome is the absence of important conserved genes known to be involved in the antimicrobial responses of other insects. This reduction in antibacterial immune defences is thought to be related to their long-term association with beneficial symbiotic bacteria and to facilitate symbiont maintenance. An additional possibility to avoid elimination of mutualistic symbionts is a fine-tuning of the host immune response. To explore this hypothesis we investigated the existence and potential involvement of immune regulators in aphid agonistic and antagonistic interactions. RESULTS: In contrast to the limited antibacterial arsenal, we showed that the pea aphid Acyrthosiphon pisum expresses 5 members of Macrophage Migration Inhibitory Factors (ApMIF), known to be key regulators of the innate immune response. In silico searches for MIF members in insect genomes followed by phylogenetic reconstruction suggest that evolution of MIF genes in hemipteran species has been shaped both by differential losses and serial duplications, raising the question of the functional importance of these genes in aphid immune responses. Expression analyses of ApMIFs revealed reduced expression levels in the presence, or during the establishment of secondary symbionts. By contrast, ApMIFs expression levels significantly increased upon challenge with a parasitoid or a Gram-negative bacteria. This increased expression in the presence of a pathogen/parasitoid was reduced or missing, in the presence of facultative symbiotic bacteria. CONCLUSIONS: This work provides evidence that while aphid's antibacterial arsenal is reduced, other immune genes widely absent from insect genomes are present, diversified and differentially regulated during antagonistic or agonistic interactions.

Transcriptome dynamics of Arabidopsis thaliana root penetration by the oomycete pathogen Phytophthora parasitica.

BACKGROUND: Oomycetes are a group of filamentous microorganisms that includes both animal and plant pathogens and causes major agricultural losses. Phytophthora species can infect most crops and plants from natural ecosystems. Despite their tremendous economic and ecologic importance, few effective methods exist for limiting the damage caused by these species. New solutions are required, and their development will require improvements in our understanding of the molecular events governing infection by these pathogens. In this study, we characterized the genetic program activated during penetration of the plant by the soil-borne pathogen Phytophthora parasitica. RESULTS: Using all the P. parasitica sequences available in public databases, we generated a custom oligo-array and performed a transcriptomic analysis of the early events of Arabidopsis thaliana infection. We characterized biological stages, ranging from the appressorium-mediated penetration of the pathogen into the roots to the occurrence of first dead cells in the plant. We identified a series of sequences that were transiently modulated during host penetration. Surprisingly, we observed an overall down regulation of genes encoding proteins involved in lipid and sugar metabolism, and an upregulation of functions controlling the transport of amino acids. We also showed that different groups of genes were expressed by P. parasitica during host penetration and the subsequent necrotrophic phase. Differential expression patterns were particularly marked for cell wall-degrading enzymes and other proteins involved in pathogenicity, including RXLR effectors. By transforming P. parasitica with a transcriptional fusion with GFP, we showed that an RXLR-ecoding gene was expressed in the appressorium and infectious hyphae during infection of the first plant cell. CONCLUSION: We have characterized the genetic program activated during the initial invasion of plant cells by P. parasitica. We showed that a specific set of proteins, including effectors, was mobilized for penetration and to facilitate infection. Our detection of the expression of an RXLR encoding gene by the appressorium and infection hyphae highlights a role of this structure in the manipulation of the host cells.

Identification of the main venom protein components of Aphidius ervi, a parasitoid wasp of the aphid model Acyrthosiphon pisum.

BACKGROUND: Endoparasitoid wasps are important natural enemies of the widely distributed aphid pests and are mainly used as biological control agents. However, despite the increased interest on aphid interaction networks, only sparse information is available on the factors used by parasitoids to modulate the aphid physiology. Our aim was here to identify the major protein components of the venom injected at oviposition by Aphidius ervi to ensure successful development in its aphid host, Acyrthosiphon pisum. RESULTS: A combined large-scale transcriptomic and proteomic approach allowed us to identify 16 putative venom proteins among which three γ-glutamyl transpeptidases (γ-GTs) were by far the most abundant. Two of the γ-GTs most likely correspond to alleles of the same gene, with one of these alleles previously described as involved in host castration. The third γ-GT was only distantly related to the others and may not be functional owing to the presence of mutations in the active site. Among the other abundant proteins in the venom, several were unique to A. ervi such as the molecular chaperone endoplasmin possibly involved in protecting proteins during their secretion and transport in the host. Abundant transcripts encoding three secreted cystein-rich toxin-like peptides whose function remains to be explored were also identified. CONCLUSIONS: Our data further support the role of γ-GTs as key players in A. ervi success on aphid hosts. However, they also evidence that this wasp venom is a complex fluid that contains diverse, more or less specific, protein components. Their characterization will undoubtedly help deciphering parasitoid-aphid and parasitoid-aphid-symbiont interactions. Finally, this study also shed light on the quick evolution of venom components through processes such as duplication and convergent recruitment of virulence factors between unrelated organisms.

Characterization of PPMUCL1/2/3, three members of a new oomycete-specific mucin-like protein family residing in Phytophthora parasitica biofilm.

The plant pathogen Phytophthora parasitica forms a biofilm on the host surface. The biofilm transcriptome is characterized by the expression of PPMUCL1/2/3 (PHYTOPHTHORA PARASITICA MUCIN-LIKE) genes, which we report here to be members of a new, large mucin-like gene family restricted to the oomycete lineage. These genes encode secreted proteins organized into two domains. The NH2-terminal domain is highly conserved, but of unknown function. The second domain is a mucin-like domain enriched in threonine and serine residues, with a large number of putative O-glycosylation sites and a repeated motif defining 15 subgroups among the 315 members of the family. The second domain was found to be glycosylated in the recombinant rPPMUCL1 and rPPMUCL2 proteins. An analysis of PPMUCL1/2/3 gene expression indicated that these genes were expressed in a specific and coordinated manner in the biofilm. A novel cis-motif (R) bound to nuclear proteins, suggesting a possible role in PPMUCL1/2/3 gene regulation. Immunohistochemical staining revealed that the PPMUCL1/2 proteins were secreted and accumulated on the surface of the biofilm. Our data demonstrate that PPMUCL1/2/3 belong to a new oomycete-specific family of mucin-like proteins playing a structural role in the biofilm extracellular matrix.

Transcriptome analysis of the scleractinian coral Stylophora pistillata.

The principal architects of coral reefs are the scleractinian corals; these species are divided in two major clades referred to as "robust" and "complex" corals. Although the molecular diversity of the "complex" clade has received considerable attention, with several expressed sequence tag (EST) libraries and a complete genome sequence having been constructed, the "robust" corals have received far less attention, despite the fact that robust corals have been prominent focal points for ecological and physiological studies. Filling this gap affords important opportunities to extend these studies and to improve our understanding of the differences between the two major clades. Here, we present an EST library from Stylophora pistillata (Esper 1797) and systematically analyze the assembled transcripts compared to putative homologs from the complete proteomes of six well-characterized metazoans: Nematostella vectensis, Hydra magnipapillata, Caenorhabditis elegans, Drosophila melanogaster, Strongylocentrotus purpuratus, Ciona intestinalis and Homo sapiens. Furthermore, comparative analyses of the Stylophora pistillata ESTs were performed against several Cnidaria from the Scleractinia, Actiniaria and Hydrozoa, as well as against other stony corals separately. Functional characterization of S. pistillata transcripts into KOG/COG categories and further description of Wnt and bone morphogenetic protein (BMP) signaling pathways showed that the assembled EST library provides sufficient data and coverage. These features of this new library suggest considerable opportunities for extending our understanding of the molecular and physiological behavior of "robust" corals.

Extensive inter- and intraspecific venom variation in closely related parasites targeting the same host: the case of Leptopilina parasitoids of Drosophila.

The arms race between immune suppressive parasites that produce virulence factors and hosts that evolve resistance to these factors is suggested to be a key driver for the diversification of both partners. However, little is known regarding the diversity of virulence factors in closely related parasites or the mechanisms underlying the variation of virulence. One of the best-described model to address this issue is the interaction between Leptopilina parasitic wasps and their Drosophila hosts, in which variation of virulence is well documented. Thanks to a combined transcriptomic and proteomic approach, we have identified the main secreted proteins in the venom of Leptopilina heterotoma (Gotheron strain, 66 proteins) and of two well-characterized strains of Leptopilina boulardi, ISm and ISy (65 and 49 proteins, respectively). Results revealed significant quantitative differences in venom components between the L. boulardi strains, in agreement with their different virulence properties. Strikingly, the two related Leptopilina species did not share any abundant venom protein. The main identified proteins in L. boulardi were RhoGAPs and serpins while an aspartylglucosaminidase (AGA) was found abundant in L. heterotoma. The extensive quantitative variation observed between these species may be related with their use of different virulence strategies and/or to differences in their host range (specialist versus generalist). Altogether, our data suggests that parasitoid venom can quickly evolve, mainly through rapid changes in regulation of gene expression. It also evidences venom evolutionary processes largely described in other venomous animals i.e. the convergent recruitment of venom proteins between phylogenetically unrelated organisms, and the role of duplications in the emergence of multigenic families of virulence factors.

A root-knot nematode-secreted protein is injected into giant cells and targeted to the nuclei.

Root-knot nematodes (RKNs) are obligate endoparasites that maintain a biotrophic relationship with their hosts over a period of several weeks and induce the differentiation of root cells into specialized feeding cells. Nematode effectors synthesized in the oesophageal glands and injected into the plant tissue through the syringe-like stylet certainly play a central role in these processes. In a search for nematode effectors, we used comparative genomics on expressed sequence tag (EST) datasets to identify Meloidogyne incognita genes encoding proteins potentially secreted upon the early steps of infection. We identified three genes specifically expressed in the oesophageal glands of parasitic juveniles that encode predicted secreted proteins. One of these genes, Mi-EFF1 is a pioneer gene that has no similarity in databases and a predicted nuclear localization signal. We demonstrate that RKNs secrete Mi-EFF1 within the feeding site and show Mi-EFF1 targeting to the nuclei of the feeding cells. RKNs were previously shown to secrete proteins in the apoplasm of infected tissues. Our results show that nematodes sedentarily established at the feeding site also deliver proteins within plant cells through their stylet. The protein Mi-EFF1 injected within the feeding cells is targeted at the nuclei where it may manipulate nuclear functions of the host cell.

Molecular cloning and characterization of first organic matrix protein from sclerites of red coral, Corallium rubrum.

We report here for the first time the isolation and characterization of a protein from the organic matrix (OM) of the sclerites of the alcyonarian, Corallium rubrum. This protein named scleritin is one of the predominant proteins extracted from the EDTA-soluble fraction of the OM. The entire open reading frame (ORF) was obtained by comparing amino acid sequences from de novo mass spectrometry and Edman degradation with an expressed sequence tag library dataset of C. rubrum. Scleritin is a secreted basic phosphorylated protein which exhibits a short amino acid sequence of 135 amino acids and a signal peptide of 20 amino acids. From specific antibodies raised against peptide sequences of scleritin, we obtained immunolabeling of scleroblasts and OM of the sclerites which provides information on the biomineralization pathway in C. rubrum.

Specific versus non-specific immune responses in an invertebrate species evidenced by a comparative de novo sequencing study.

Our present understanding of the functioning and evolutionary history of invertebrate innate immunity derives mostly from studies on a few model species belonging to ecdysozoa. In particular, the characterization of signaling pathways dedicated to specific responses towards fungi and Gram-positive or Gram-negative bacteria in Drosophila melanogaster challenged our original view of a non-specific immunity in invertebrates. However, much remains to be elucidated from lophotrochozoan species. To investigate the global specificity of the immune response in the fresh-water snail Biomphalaria glabrata, we used massive Illumina sequencing of 5'-end cDNAs to compare expression profiles after challenge by Gram-positive or Gram-negative bacteria or after a yeast challenge. 5'-end cDNA sequencing of the libraries yielded over 12 millions high quality reads. To link these short reads to expressed genes, we prepared a reference transcriptomic database through automatic assembly and annotation of the 758,510 redundant sequences (ESTs, mRNAs) of B. glabrata available in public databases. Computational analysis of Illumina reads followed by multivariate analyses allowed identification of 1685 candidate transcripts differentially expressed after an immune challenge, with a two fold ratio between transcripts showing a challenge-specific expression versus a lower or non-specific differential expression. Differential expression has been validated using quantitative PCR for a subset of randomly selected candidates. Predicted functions of annotated candidates (approx. 700 unisequences) belonged to a large extend to similar functional categories or protein types. This work significantly expands upon previous gene discovery and expression studies on B. glabrata and suggests that responses to various pathogens may involve similar immune processes or signaling pathways but different genes belonging to multigenic families. These results raise the question of the importance of gene duplication and acquisition of paralog functional diversity in the evolution of specific invertebrate immune responses.

Data-mining of the Meloidogyne incognita degradome and comparative analysis of proteases in nematodes.

Proteases perform essential physiological functions in all living organisms. In parasitic helminths, they are of particular importance for tissue penetration, digestion of host tissues for nutrition, and evasion of host immune responses. The recent availability of the genome sequence of the nematode Meloidogyne incognita has allowed the analysis of the protease repertoire of this major crop pathogen. The M. incognita degradome consists of at least 334 proteases that are distributed into 43 families of the five known catalytic classes. Expression profiling identified protease genes with a differential transcript level between eggs and infective juveniles. Comparing the M. incognita degradome with those of five other nematodes showed discrepancies in the distribution of some protease families, including large expansion in some families, that could reflect specific aspects of the parasitic lifestyle of this organism. This comparative study should provide a framework for deciphering the diversity of protease-mediated functions in nematodes.

Peroxiredoxins from the plant parasitic root-knot nematode, Meloidogyne incognita, are required for successful development within the host.

Root-knot nematodes, Meloidogyne spp., are sedentary biotrophic parasites which are able to infest > 2000 plant species. After root invasion they settle sedentarily inside the vascular cylinder and maintain a compatible interaction for up to 8 weeks. Plant cells respond to pathogen attacks by producing reactive oxygen species (ROS). These ROS, in particular hydroperoxides, are important regulators of host-parasite interactions and partly govern the success or failure of disease. ROS producing and ROS scavenging enzymes from both the pathogen and the host finely tune the redox state at the host-pathogen interface. We have analysed the gene structure and organization of peroxiredoxins (prx) in Meloidogyne incognita and analysed their role in the establishment of the nematode in its host. Meloidogyne incognita has seven prx genes that can be grouped with other nematode prx into three clades. Clade B prx genes are more actively transcribed in parasitic stages compared with free-living pre-parasitic juveniles. We confirmed in vitro the activity of one of these, Mi-prx2.1, on hydrogen peroxide and butylhydroperoxide. We showed by ultrastructural immunocytochemistry the expression of clade B PRX proteins in the hypodermis and pseudocoelum beneath the tissues directly in contact with the environment, both in free-living and parasitic stages. Finally, knock-down of clade B prx genes led to a significant reduction in the ability of the nematodes to complete their life cycle in the host. The expression of clade B PRX proteins in the tissues in close contact with plant cells during parasitism and the impaired development of nematodes inside the host after clade B prx knock-down suggest an important role for these genes during infection.

Genome-wide survey and analysis of microsatellites in nematodes, with a focus on the plant-parasitic species Meloidogyne incognita.

BACKGROUND: Microsatellites are the most popular source of molecular markers for studying population genetic variation in eukaryotes. However, few data are currently available about their genomic distribution and abundance across the phylum Nematoda. The recent completion of the genomes of several nematode species, including Meloidogyne incognita, a major agricultural pest worldwide, now opens the way for a comparative survey and analysis of microsatellites in these organisms. RESULTS: Using MsatFinder, the total numbers of 1-6 bp perfect microsatellites detected in the complete genomes of five nematode species (Brugia malayi, Caenorhabditis elegans, M. hapla, M. incognita, Pristionchus pacificus) ranged from 2,842 to 61,547, and covered from 0.09 to 1.20% of the nematode genomes. Under our search criteria, the most common repeat motifs for each length class varied according to the different nematode species considered, with no obvious relation to the AT-richness of their genomes. Overall, (AT)n, (AG)n and (CT)n were the three most frequent dinucleotide microsatellite motifs found in the five genomes considered. Except for two motifs in P. pacificus, all the most frequent trinucleotide motifs were AT-rich, with (AAT)n and (ATT)n being the only common to the five nematode species. A particular attention was paid to the microsatellite content of the plant-parasitic species M. incognita. In this species, a repertoire of 4,880 microsatellite loci was identified, from which 2,183 appeared suitable to design markers for population genetic studies. Interestingly, 1,094 microsatellites were identified in 801 predicted protein-coding regions, 99% of them being trinucleotides. When compared against the InterPro domain database, 497 of these CDS were successfully annotated, and further assigned to Gene Ontology terms. CONCLUSIONS: Contrasted patterns of microsatellite abundance and diversity were characterized in five nematode genomes, even in the case of two closely related Meloidogyne species. 2,245 di- to hexanucleotide loci were identified in the genome of M. incognita, providing adequate material for the future development of a wide range of microsatellite markers in this major plant parasite.

Comprehensive EST analysis of the symbiotic sea anemone, Anemonia viridis.

BACKGROUND: Coral reef ecosystems are renowned for their diversity and beauty. Their immense ecological success is due to a symbiotic association between cnidarian hosts and unicellular dinoflagellate algae, known as zooxanthellae. These algae are photosynthetic and the cnidarian-zooxanthellae association is based on nutritional exchanges. Maintenance of such an intimate cellular partnership involves many crosstalks between the partners. To better characterize symbiotic relationships between a cnidarian host and its dinoflagellate symbionts, we conducted a large-scale EST study on a symbiotic sea anemone, Anemonia viridis, in which the two tissue layers (epiderm and gastroderm) can be easily separated. RESULTS: A single cDNA library was constructed from symbiotic tissue of sea anemones A. viridis in various environmental conditions (both normal and stressed). We generated 39,939 high quality ESTs, which were assembled into 14,504 unique sequences (UniSeqs). Sequences were analysed and sorted according to their putative origin (animal, algal or bacterial). We identified many new repeated elements in the 3'UTR of most animal genes, suggesting that these elements potentially have a biological role, especially with respect to gene expression regulation. We identified genes of animal origin that have no homolog in the non-symbiotic starlet sea anemone Nematostella vectensis genome, but in other symbiotic cnidarians, and may therefore be involved in the symbiosis relationship in A. viridis. Comparison of protein domain occurrence in A. viridis with that in N. vectensis demonstrated an increase in abundance of some molecular functions, such as protein binding or antioxidant activity, suggesting that these functions are essential for the symbiotic state and may be specific adaptations. CONCLUSION: This large dataset of sequences provides a valuable resource for future studies on symbiotic interactions in Cnidaria. The comparison with the closest available genome, the sea anemone N. vectensis, as well as with EST datasets from other symbiotic cnidarians provided a set of candidate genes involved in symbiosis-related molecular crosstalks. Altogether, these results provide new molecular insights that could be used as a starting-point for further functional genomics studies.

Genome sequence of the metazoan plant-parasitic nematode Meloidogyne incognita.

Plant-parasitic nematodes are major agricultural pests worldwide and novel approaches to control them are sorely needed. We report the draft genome sequence of the root-knot nematode Meloidogyne incognita, a biotrophic parasite of many crops, including tomato, cotton and coffee. Most of the assembled sequence of this asexually reproducing nematode, totaling 86 Mb, exists in pairs of homologous but divergent segments. This suggests that ancient allelic regions in M. incognita are evolving toward effective haploidy, permitting new mechanisms of adaptation. The number and diversity of plant cell wall-degrading enzymes in M. incognita is unprecedented in any animal for which a genome sequence is available, and may derive from multiple horizontal gene transfers from bacterial sources. Our results provide insights into the adaptations required by metazoans to successfully parasitize immunocompetent plants, and open the way for discovering new antiparasitic strategies.

Design of a data model for developing laboratory information management and analysis systems for protein production.

Data management has emerged as one of the central issues in the high-throughput processes of taking a protein target sequence through to a protein sample. To simplify this task, and following extensive consultation with the international structural genomics community, we describe here a model of the data related to protein production. The model is suitable for both large and small facilities for use in tracking samples, experiments, and results through the many procedures involved. The model is described in Unified Modeling Language (UML). In addition, we present relational database schemas derived from the UML. These relational schemas are already in use in a number of data management projects.

An evolving hierarchical family classification for glycosyltransferases.

Glycosyltransferases are a ubiquitous group of enzymes that catalyse the transfer of a sugar moiety from an activated sugar donor onto saccharide or non-saccharide acceptors. Although many glycosyltransferases catalyse chemically similar reactions, presumably through transition states with substantial oxocarbenium ion character, they display remarkable diversity in their donor, acceptor and product specificity and thereby generate a potentially infinite number of glycoconjugates, oligo- and polysaccharides. We have performed a comprehensive survey of glycosyltransferase-related sequences (over 7200 to date) and present here a classification of these enzymes akin to that proposed previously for glycoside hydrolases, into a hierarchical system of families, clans, and folds. This evolving classification rationalises structural and mechanistic investigation, harnesses information from a wide variety of related enzymes to inform cell biology and overcomes recurrent problems in the functional prediction of glycosyltransferase-related open-reading frames.