Caenorhabditis elegans functional genomics: omic resonance.

The nematode Caenorhabditis elegans is widely used as a model organism for studying many fundamental aspects of development and cell biology, including processes underlying human disease. The genome of C. elegans encodes over 19,000 protein-coding genes and hundreds of non-coding RNAs. The availability of whole genome sequence has facilitated the development of high throughput techniques for elucidating the function of individual genes and gene products. Furthermore, attempts can now be made to integrate these substantial functional genomics data collections and to understand at a global level how the flow of genomic information that is at the core of the central dogma leads to the development of a multicellular organism.     

Identification of Heterodera glycines (soybean cyst nematode [SCN]) cDNA sequences with high identity to those of Caenorhabditis elegans having lethal mutant or RNAi phenotypes

The soybean cyst nematode (SCN; Heterodera glycines) is a devastating obligate parasite of Glycine max (soybean) causing one billion dollars in losses to the US economy per year and over ten billion dollars in losses worldwide. While much is understood about the pathology of H. glycines, its genome sequence is not well characterized or fully sequenced. We sought to create bioinformatic tools to mine the H. glycines nucleotide database. One way is to use a comparative genomics approach by anchoring our analysis with an organism, like the free-living nematode Caenorhabditis elegans. Unlike H. glycines, the C. elegans genome is fully sequenced and is well characterized with a number of lethal genes identified through experimental methods. We compared an EST database of H. glycines with the C. elegans genome. Our goal was identifying genes that may be essential for H. glycines survival and would serve as an automated pipeline for RNAi studies to both study and control H. glycines. Our analysis yielded a total of nearly 8334 conserved genes between H. glycines and C. elegans. Of these, 1508 have lethal phenotypes/phenocopies in C. elegans. RNAi of a conserved ribosomal gene from H. glycines (Hg-rps-23) yielded dead and dying worms as shown by positive Sytox fluorescence. Endogenous Hg-rps-23 exhibited typical RNA silencing as shown by RT-PCR. However, an unrelated gene Hg-unc-87 did not exhibit RNA silencing in the Hg-rps-23 dsRNA-treated worms, demonstrating the specificity of the silencing.     

Caenorhabditis elegans and the study of gene function in parasites.

The free-living nematode Caenorhabditis elegans is a tractable experimental model system for the study of both vertebrate and invertebrate biology. Its most significant advantages are its simplicity, both in anatomy and in genomic organization, and the elaborate methods that have been developed to attribute function to previously uncharacterized genes. Importantly, > 40% of parasitic nematode genes exhibit high levels of homology to genes within the C. elegans genome. Studying such genes using the C. elegans model should yield new insights into key molecules and their possible implications in parasite survival, leading to the discovery of new drug targets and vaccine candidates.     

High-throughput RNAi in Caenorhabditis elegans: genome-wide screens and functional genomics

The phenomenon of RNA-mediated interference (RNAi) was first discovered in the nematode Caenorhabditis elegans, in which introduction of double-stranded RNA causes specific inactivation of genes with corresponding sequences. Technical advances in RNAi methodology and the availability of the complete genome sequence have enabled the high-throughput, genome-wide RNAi analysis of this organism. Several groups have used large-scale RNAi to systematically examine every C. elegans gene for knock-down phenotypes, providing basal information to be mined in more detailed studies. Now, in addition to functional genomic RNAi analyses, high-throughput RNAi is also routinely used for rapid, genome-wide screens for genes involved in specific biological processes. The integration of high-throughput RNAi experiments with other large-scale data, such as DNA microarrays and protein-protein interaction maps, enhances the speed and reliability of such screens. The accumulation of RNAi phenotype data dramatically accelerates our understanding of this organism at the genetic level.     

Efficient and cell specific knock-down of gene function in targeted C. elegans neurons

The nematode C. elegans has become an important model for understanding how genes influence behavior. However, in this organism the available approaches for identifying the neuron(s) where the function of a gene is required for a given behavioral trait are time consuming and restricted to non essential genes for which mutants are available. We describe a simple reverse genetics approach for reducing, in chosen C. elegans neurons, the function of genes. The method is based on the expression, under cell specific promoters, of sense and antisense RNA corresponding to a gene of interest. By targeting the genes osm-10, osm-6 and the Green Fluorescent Protein gene, gfp, we show that this approach leads to efficient, heritable and cell autonomous knock-downs of gene function, even in neurons usually refractory to classic RNA interference (RNAi). By targeting the essential and ubiquitously expressed gene, gpb-1, which encodes a G protein beta subunit, we identify for the first time two distinct sets of neurons in which the function of gpb-1 is required to regulate two distinct behaviors: egg-laying and avoidance of repellents. The cell specific knock-downs obtained with this approach provide information that is complementary to that provided by the cell specific rescue of loss-of-function mutations and represents a useful new tool for dissecting the role that genes play in selected neurons.     

Informatics issues in large-scale sequence analysis: elucidating the protein kinases of C. elegans

With the availability of the nearly complete genomic sequence of C. elegans, the first multicellular organism to be sequenced, molecular biology has definitely entered the postgenomic era. Annotation of the genomic sequence, which refers to identifying the genes and other biologically relevant sections of the genome, is an important and nontrivial next step. A first-pass annotation will be necessarily incomplete but will drive further biological experiments, which in turn will help to annotate the genome better. Given the scale of the genome sequence analysis, it is clear that the annotation should be automated as much as possible without sacrificing the quality of analysis. In this work, we outline our approach to identifying the protein kinases of C. elegans from the genomic sequence. We describe new tools we have developed for analysis, management and visualization of genomic data. By developing modular and scalable solutions, this study has provided a framework for future analysis of the Drosophila and human genomes.     

The use of Caenorhabditis elegans in parasitic nematode research

There is increasing interest in the use of the free-living nematode Caenorhabditis elegans as a tool for parasitic nematode research and there are now a number of compelling examples of its successful application. C. elegans has the potential to become a standard tool for molecular helminthology researchers, just as yeast is routinely used by molecular biologists to study vertebrate biology. However, in order to exploit C. elegans in a meaningful manner, we need a detailed understanding of the extent to which different aspects of C. elegans biology have been conserved with particular groups of parasitic nematodes. This review first considers the current state of knowledge regarding the conservation of genome organisation across the nematode phylum and then discusses some recent evolutionary development studies in free-living nematodes. The aim is to provide some important concepts that are relevant to the extrapolation of information from C. elegans to parasitic nematodes and also to the interpretation of experiments that use C. elegans as a surrogate expression system. In general, examples have been specifically chosen because they highlight the importance of careful experimentation and interpretation of data. Consequently, the focus is on the differences that have been found between nematode species rather than the similarities. Finally, there is a detailed discussion of the current status of C. elegans as a heterologous expression system to study parasite gene function and regulation using successful examples from the literature.     

Operon structure and trans-splicing in the nematode Pristionchus pacificus

In the nematode Caenorhabditis elegans, up to 15% of the genes are organized in operons. Polycistronic precursor RNAs are processed by trans-splicing at the 5' ends of genes by adding a specific trans-spliced leader. Ten different spliced leaders are known in C. elegans that differ in sequence and abundance. The SL1 leader is most abundant and is spliced to the 5' ends of monocistronic genes and to upstream genes in operons. Trans-splicing is common among nematodes and was observed in the genera Panagrellus, Ascaris, Haemonchus, Anisakis, and Brugia. However, little is known about operons in nonrhabditid nematodes. Dolichorhabditis CEW1, another rhabditid nematode that is now called Oscheius CEW1, contains operons and SL2 trans-splicing. We have studied the presence of operons and trans-splicing in Pristionchus pacificus, a species of the Diplogastridae that has recently been developed as a satellite organism in evolutionary developmental biology. We provide evidence that P. pacificus contains operons and that downstream genes are trans-spliced to SL2. Surprisingly, the one operon analyzed so far in P. pacificus is not conserved in C. elegans, suggesting unexpected genomic plasticity.     

线虫(Caenorhabditis elegans)基因组的研究进展

线虫（Ｃａｅｎｏｒｈａｂｄｉｔｉｓ ｅｌｅｇａｎｓ）是重要的模式生物,其基因组序列分析工作于１９９８年底基本完成,已有１９０００多个基因被鉴定。本文概述线虫基因组研究中遗传图谱、物理图谱、序列测定和基因识别等方面的研究成果,以及线虫基因组计划将对生命科学研究产生的影响。     

Genes coding for 5S ribosomal RNA of the nematode Caenorhabditis elegans

We have identified a 1-kb genomic sequence that represents the major class of 5S rRNA genes in the nematode Caenorhabditis elegans. This 1-kb sequence is tandemly repeated 110 times in the haploid genome forming a single homogeneous gene family. Other nematode genomic sequences, distinct from the major 1-kb repeat class but homologous to it, may represent dispersed 5S rRNA genes or the ends of a gene cluster. One such fragment shows a restriction fragment length difference between two C. elegans strains. This should allow the genetic analysis of 5S rRNA-coding DNA (5S X rDNA) and its flanking regions in C. elegans.     

Updating the str and srj (stl) families of chemoreceptors in Caenorhabditis nematodes reveals frequent gene movement within and between chromosomes

The seven transmembrane receptor (str) and srj (renamed from stl) families of chemoreceptors have been updated and the genes formally named following completion of the Caenorhabditis elegans genome sequencing project. Analysis of gene locations revealed that 84% of the 320 genes and pseudogenes in these two families reside on the large chromosome V. Movements to other chromosomes, especially chromosome IV, have nevertheless been relatively common, but only one has led to further gene family diversification. Comparisons with homologs in C. briggsae indicated that 22.5% of these genes have been newly formed by gene duplication since the species split, while also showing that four have been lost by large deletions. These patterns of gene evolution are similar to those revealed by analysis of the equally large srh family of chemoreceptors, and are likely to reflect general features of nematode genome dynamics. Thus large random deletions presumably balance the rapid proliferation of genes and their degeneration into pseudogenes, while gene movement within and between chromosomes keeps these nematode genomes in flux.     

C. elegans clk-2, a gene that limits life span, encodes a telomere length regulator similar to yeast telomere binding protein Tel2p.

An important quest in modern biology is to identify genes involved in aging. Model organisms such as the nematode Caenorhabditis elegans are particularly useful in this regard. The C. elegans genome has been sequenced [1], and single gene mutations that extend adult life span have been identified [2]. Among these longevity-controlling loci are four apparently unrelated genes that belong to the clk family. In mammals, telomere length and structure can influence cellular, and possibly organismal, aging. Here, we show that clk-2 encodes a regulator of telomere length in C. elegans.     

Mining predicted essential genes of Brugia malayi for nematode drug targets

We report results from the first genome-wide application of a rational drug target selection methodology to a metazoan pathogen genome, the completed draft sequence of Brugia malayi, a parasitic nematode responsible for human lymphatic filariasis. More than 1.5 billion people worldwide are at risk of contracting lymphatic filariasis and onchocerciasis, a related filarial disease. Drug treatments for filariasis have not changed significantly in over 20 years, and with the risk of resistance rising, there is an urgent need for the development of new anti-filarial drug therapies. The recent publication of the draft genomic sequence for B. malayi enables a genome-wide search for new drug targets. However, there is no functional genomics data in B. malayi to guide the selection of potential drug targets. To circumvent this problem, we have utilized the free-living model nematode Caenorhabditis elegans as a surrogate for B. malayi. Sequence comparisons between the two genomes allow us to map C. elegans orthologs to B. malayi genes. Using these orthology mappings and by incorporating the extensive genomic and functional genomic data, including genome-wide RNAi screens, that already exist for C. elegans, we identify potentially essential genes in B. malayi. Further incorporation of human host genome sequence data and a custom algorithm for prioritization enables us to collect and rank nearly 600 drug target candidates. Previously identified potential drug targets cluster near the top of our prioritized list, lending credibility to our methodology. Over-represented Gene Ontology terms, predicted InterPro domains, and RNAi phenotypes of C. elegans orthologs associated with the potential target pool are identified. By virtue of the selection procedure, the potential B. malayi drug targets highlight components of key processes in nematode biology such as central metabolism, molting and regulation of gene expression.     

C. elegans as a model for human inherited degenerative diseases

The nematode C. elegans is an established model for developmental biology. Since the early 90's, this simple model organism has been increasingly used for studying human disease pathogenesis. C. elegans models based either on the mutagenesis of human disease genes conserved in this nematode or transgenesis with disease genes not conserved in C. elegans show several features that are observed in mammalian models. These observations suggest that the genetic dissection and pharmacological manipulation of disease-like phenotypes in C. elegans will shed light on the cellular mechanisms that are altered in human diseases, and the compounds that may be used as drugs. This review illustrates these aspects by commenting on two inherited degenerative diseases, Duchenne's muscular dystrophy and Huntington's neurodegenerative disease.     

Mining nematode genome data for novel drug targets

Expressed sequence tag projects have currently produced over 400 000 partial gene sequences from more than 30 nematode species and the full genomic sequences of selected nematodes are being determined. In addition, functional analyses in the model nematode Caenorhabditis elegans have addressed the role of almost all genes predicted by the genome sequence. This recent explosion in the amount of available nematode DNA sequences, coupled with new gene function data, provides an unprecedented opportunity to identify pre-validated drug targets through efficient mining of nematode genomic databases. This article describes the various information sources available and strategies that can expedite this process.     

Nematodes of the genus Pristionchus are closely associated with scarab beetles and the Colorado potato beetle in Western Europe

Evolutionary developmental biology examines how changes in developmental programmes give rise to developmental and, ultimately, morphological novelty. To this end, comparisons of related but distinct organisms have to be performed. The diplogastrid nematode Pristionchus pacificus has been developed as a satellite system for a detailed comparison of various developmental processes to the model organism Caenorhabditis elegans, a rhabditid nematode. In addition to developmental and genetic studies, a genomic platform has been established to analyse the biology of this organism. However, only little is known about where and how Pristionchus pacificus and its relatives live in the wild. Here we show that nematodes of the genus Pristionchus live in close association with scarabaeoid beetles and the Colorado potato beetle. In total, we generated 371 isogenic female lines from 4242 beetles collected at 25 sampling sites all over Europe. Isogenic female lines were subjected to sequence analysis and mating experiments for species determination. The 371 isolates fell into six species. Two hermaphroditic species account for about 60% of the collected nematodes. We found Pristionchus maupasi almost exclusively on cockchafers and Pristionchus entomophagus predominantly on dung beetles. Colorado potato beetles carried the gonochoristic species Pristionchus uniformis, which was only rarely observed on scarabaeoid beetles. We describe the initial evidence for the association of Pristionchus nematodes with beetles and provide a phylogeny based on sequence analysis of the small subunit ribosomal RNA gene.