A divergent INS protein in Caenorhabditis elegans structurally resembles human insulin and activates the human insulin receptor

Caenorhabditis elegans contains a family of putative insulin-like genes proposed to regulate dauer arrest and senescence. These sequences often lack characteristic sequence features of human insulin essential for its folding, structure, and function. Here, we describe the structure and receptor-binding properties of INS-6, a single-chain polypeptide expressed in specific neurons. Despite multiple nonconservative changes in sequence, INS-6 recapitulates an insulin-like fold. Although lacking classical receptor-binding determinants, INS-6 binds to and activates the human insulin receptor. Its activity is greater than that of an analogous single-chain human insulin analog.     

The protein L-isoaspartyl-O-methyltransferase functions in the Caenorhabditis elegans stress response

The efficient use of nutrients is important in development and aging. In this study, we asked if the protein repair methyltransferase has a related or additional role in energy metabolism and stress response in the nematode Caenorhabditis elegans. Worms lacking the pcm-1 gene encoding this enzyme exhibit reduced longevity as SDS-isolated dauer larvae and as arrested L1 larvae under starvation stress, while overexpression leads to increased adult longevity. These findings led us to question whether pcm-1 deficient C. elegans may have inappropriate metabolic responses to stress. We assayed dauer and dauer-like larvae for starvation survival and observed a two-fold reduction of median survival time for pcm-1 mutants compared to N2 wild-type worms. Under these conditions, pcm-1 deficient dauer larvae had reduced fat stores, suggesting that PCM-1 may have a role in the initiation of the correct metabolic responses to stress starvation. We show expression of the pcm-1 gene in neurons, body wall and reproductive tissues. Upon heat shock and dauer formation-inducing conditions, we observe additional pcm-1 expression in body wall muscle nuclei and actomyosin filaments and in hypodermal cells. These results suggest that this enzyme may be important in stress response pathways, including proper decision making for energy storage.     

Genomic analysis of Caenorhabditis elegans reveals ancient families of retroviral-like elements.

Retrotransposons are the most abundant and widespread class of eukaryotic transposable elements. The recent genome sequencing of Caenorhabditis elegans has provided an unprecedented opportunity to analyze the evolutionary relationships among the entire complement of retrotransposons within a multicellular eukaryotic organism. In this article we report the results of an analysis of retroviral-like long terminal repeat retrotransposons in C. elegans that indicate that this class of elements may be even more abundant and divergent than previously expected. The unexpected presence, in C. elegans, of an element displaying a number of characteristics previously thought to be unique to vertebrate retroviruses suggests an ancient lineage for this important class of infectious agents.     

Systematic identification of functional orthologs based on protein network comparison

Annotating protein function across species is an important task that is often complicated by the presence of large paralogous gene families. Here, we report a novel strategy for identifying functionally related proteins that supplements sequence-based comparisons with information on conserved protein-protein interactions. First, the protein interaction networks of two species are aligned by assigning proteins to sequence homology clusters using the Inparanoid algorithm. Next, probabilistic inference is performed on the aligned networks to identify pairs of proteins, one from each species, that are likely to retain the same function based on conservation of their interacting partners. Applying this method to Drosophila melanogaster and Saccharomyces cerevisiae, we analyze 121 cases for which functional orthology assignment is ambiguous when sequence similarity is used alone. In 61 of these cases, the network supports a different protein pair than that favored by sequence comparisons. These results suggest that network analysis can be used to provide a key source of information for refining sequence-based homology searches.     

Suppression of polyglutamine-induced protein aggregation in Caenorhabditis elegans by torsin proteins

Torsion dystonia is an autosomal dominant movement disorder characterized by involuntary, repetitive muscle contractions and twisted postures. The most severe early-onset form of dystonia has been linked to mutations in the human DYT1 (TOR1A) gene encoding a protein termed torsinA. While causative genetic alterations have been identified, the function of torsin proteins and the molecular mechanism underlying dystonia remain unknown. Phylogenetic analysis of the torsin protein family indicates these proteins share distant sequence similarity with the large and diverse family of AAA+ proteins. We have established the nematode, Caenorhabditis elegans, as a model system for examining torsin activity. Using an in vivo assay for polyglutamine repeat-induced protein aggregation in living animals, we have determined that ectopic overexpression of both human and C. elegans torsin proteins results in a dramatic reduction of polyglutamine-dependent protein aggregation in a manner similar to that previously reported for molecular chaperones. The suppressive effects of torsin overexpression persisted as animals aged, whereas a mutant nematode torsin protein was incapable of ameliorating aggregate formation. Antibody staining of transgenic animals indicated that both the C. elegans torsin-related protein TOR-2 and ubiquitin were localized to sites of protein aggregation. These data represent the first functional evidence of a role for torsins in effectively managing protein folding and suggest that possible breakdown in a neuroprotective mechanism that is, in part, mediated by torsins may be responsible for the neuronal dysfunction associated with dystonia.     

Fluorodeoxyuridine affects the identification of metabolic responses to daf-2 status in Caenorhabditis elegans

Fluorodeoxyuridine (FUdR) is often used to help maintain synchronous populations of Caenorhabditis elegans adults, for instance as would typically be the case in studying age-related effects. However, given that FUdR inhibits DNA synthesis and therefore reproduction, it will clearly have significant wide-ranging biological effects. It is often assumed that these can be compensated for using appropriate controls. We show here that this is not the case for a metabolomic analysis of a long-lived daf-2 mutant strain: not only were the effects of FUdR much greater than the effects of the mutation, there were clear interactions between FUdR and genotype, such that identification of daf-2-dependent metabolites would have been compromised on FUdR plates. This indicates that FUdR should only be used with caution for C. elegans ageing experiments, and should not be assumed to be independent of other factors being studied.     

Molecular pathways that influence human tau-induced pathology in Caenorhabditis elegans

Mutations in the gene encoding tau cause frontotemporal dementia with parkinsonism--chromosome 17 type (FTDP-17). In FTDP-17, Alzheimer's disease, and other tauopathies, aggregated hyper-phosphorylated tau forms the neurofibrillary tangles characteristic of these disorders. We previously reported a Caenorhabditis elegans model for tauopathies using human normal and FTDP-17 mutant tau as transgenes. Neuronal transgene expression caused insoluble phosphorylated tau accumulation, neurodegeneration and uncoordinated (Unc) movement. Here we describe a genome-wide RNA-mediated interference (RNAi) screen for genes that modify the tau-induced Unc phenotype. We tested RNAi sequences for 16,757 genes and found 75 that enhanced the transgene-induced Unc phenotype. Forty-six of these genes have sequence similarity to known human genes and fall into a number of broad classes including kinases, chaperones, proteases and phosphatases. The remaining 29 modifiers have sequence similarity only with other nematode genes. To determine if the enhancers are specific for the tau-induced Unc behavior, we exposed several non-tau Unc mutants to tau RNAi enhancer clones. Fifteen enhancers modified phenotypes in multiple Unc mutants, whereas 60 modified only the Unc phenotype in the tau transgenic lines. We also introduced the tau transgene into the background of genetic loss-of-function mutations for a subset of the enhancer genes. Tau transgenic animals homozygous for loss of these enhancer genes exhibited increased impaired motility relative to the tau transgene line alone. This work uncovers novel candidate genes that prevent tau toxicity, as well as genes previously implicated in tau-mediated neurodegeneration.     

The Drosophila septate junctions beyond barrier function: Review of the literature,prediction of human orthologs of the SJ‐related proteins and identification of protein domain families

The involvement of Septate Junctions (SJs) in critical cellular functions that extend beyond their role as diffusion barriers in the epithelia and the nervous system has made the fruit fly an ideal model for the study of human diseases associated with impaired Tight Junction (TJ) function. In this study, we summarized current knowledge of the Drosophila melanogaster SJ‐related proteins, focusing on their unconventional functions. Additionally, we sought to identify human orthologs of the corresponding genes as well as protein domain families. The systematic literature search was performed in PubMed and Scopus databases using relevant key terms. Orthologs were predicted using the DIOPT tool and aligned protein regions were determined from the Pfam database. 3‐D models of the smooth SJ proteins were built on the Phyre2 and DMPFold protein structure prediction servers. A total of 30 proteins were identified as relatives to the SJ cellular structure. Key roles of these proteins, mainly in the regulation of morphogenetic events and cellular signalling, were highlighted. The investigation of protein domain families revealed that the SJ‐related proteins contain conserved domains that are required not only for cell‐cell interactions and cell polarity but also for cellular signalling and immunity. DIOPT analysis of orthologs identified novel human genes as putative functional homologs of the fruit fly SJ genes. A gap in our knowledge was identified regarding the domains that occur in the proteins encoded by eight SJ‐associated genes. Future investigation of these domains is needed to provide functional information.     

Different genes govern Yersinia pestis pathogenicity in Caenorhabditis elegans and human lice

To assess the role of virulence factors identified in Caenorhabditis elegans in the transmission of plague by lice, we infected 100 lice by feeding them on rabbits and made them bacteremic; the rabbits had been intravenously inoculated with 10(9) CFU of six different mutant Yersinia pestis strains of lower pathogenicity for C. elegans, obtained from the KIM5 strain. This strain lacks genes used for biofilm formation. High mortality rates were observed in all lice, which excreted viable bacteria in their feces. Mutants killed rabbits when infected intravenously, but mutants were not transmitted to rabbits by infected lice. We conclude that the genes governing pathogenicity in C. elegans and louse are not identical.     

Neuropeptidergic signaling in the nematode Caenorhabditis elegans

The nematode Caenorhabditis elegans joins the menagerie of behavioral model systems next to the fruit fly Drosophila melanogaster, the marine snail Aplysia californica and the mouse. In contrast to Aplysia, which contains 20,000 neurons having cell bodies of hundreds of microns in diameter, C. elegans harbors only 302 tiny neurons from which the cell lineage is completely described, as is the case for all the other somatic cells. As such, this nervous system appears at first sight incommensurable with those of higher organisms, although genome-wide comparison of predicted C. elegans genes with their counterparts in vertebrates revealed many parallels. Together with its short lifespan and ease of cultivation, suitability for high-throughput genetic screenings and genome-wide RNA interference approaches, access to an advanced genetic toolkit and cell-ablation techniques, it seems that this tiny transparent organism of only 1mm in length has nothing to hide. Recently, highly exciting developments have occurred within the field of neuropeptidergic signaling in C. elegans, not only because of the availability of a sequenced genome since 1998, but especially because of state of the art post genomic technologies, that allow for molecular characterization of the signaling molecules. Here, we will focus on endogenous, bioactive (neuro)peptides and mainly discuss biosynthesis, peptide sequence information, localization and G-protein coupled receptors of the three major peptide families in C. elegans.     

A Caenorhabditis elegans cohesion protein with functions in meiotic chromosome pairing and disjunction

We have studied four Caenorhabditis elegans homologs of the Rad21/Scc1/Rec8 sister-chromatid cohesion protein family. Based on the RNAi phenotype and protein localization, it is concluded that one of them, W02A2.6p, is the likely worm ortholog of yeast Rec8p. The depletion of C. elegans W02A2.6p (called REC-8) by RNAi, induced univalent formation and splitting of chromosomes into sister chromatids at diakinesis. Chromosome synapsis at pachytene was defective, but primary homology recognition seemed unaffected, as a closer-than-random association of homologous fluorescence in situ hybridization (FISH) signals at leptotene/zygotene was observed. Depletion of REC-8 also induced chromosome fragmentation at diakinesis. We interpret these fragments as products of unrepaired meiotic double-stranded DNA breaks (DSBs), because fragmentation was suppressed in a spo-11 background. Thus, REC-8 seems to be required for successful repair of DSBs. The occurrence of DSBs in REC-8-depleted meiocytes suggests that DSB formation does not depend on homologous synapsis. Anti-REC-8 immunostaining decorated synaptonemal complexes (SCs) at pachytene and chromosomal axes in bivalents and univalents at diakinesis. Between metaphase I and metaphase II, REC-8 is partially lost from the chromosomes. The partial loss of REC-8 from chromosomes between metaphase I and metaphase II suggests that worm REC-8 might function similarly to yeast Rec8p. The loss of yeast Rec8p from chromosome arms at meiosis I and centromeres at meiosis II coordinates the disjunction of homologs and sister chromatids at the two meiotic divisions.     

A role for Caenorhabditis elegans in understanding the function and interactions of human disease genes

A growing number of medical research teams have begun to explore the experimental advantages of using a genetic animal model, the nematode worm Caenorhabditis elegans, with a view to enhancing our understanding of genes underlying human congenital disorders. In this study, we have compared sequences of positionally cloned human disease genes with the C.elegans database of predicted genes. Drawing on examples from spinal muscular atrophy, polycystic kidney disease, muscular dystrophy and Alzheimer's disease, we illustrate how data from C.elegans can yield new insights into the function and interactions of human disease genes.