Dataset Information

Hc-daf-2 encodes an insulin-like receptor kinase in the barber's pole worm, Haemonchus contortus, and restores partial dauer regulation.

ABSTRACT: Infective L3s (iL3s) of parasitic nematodes share common behavioural, morphological and developmental characteristics with the developmentally arrested (dauer) larvae of the free-living nematode Caenorhabditis elegans. It is proposed that similar molecular mechanisms regulate entry into or exit from the dauer stage in C. elegans, and the transition from free-living to parasitic forms of parasitic nematodes. In C. elegans, one of the key factors regulating the dauer transition is the insulin-like receptor (designated Ce-DAF-2) encoded by the gene Ce-daf-2. However, nothing is known about DAF-2 homologues in most parasitic nematodes. Here, using a PCR-based approach, we identified and characterised a gene (Hc-daf-2) and its inferred product (Hc-DAF-2) in Haemonchus contortus (a socioeconomically important parasitic nematode of ruminants). The sequence of Hc-DAF-2 displays significant sequence homology to insulin receptors (IR) in both vertebrates and invertebrates, and contains conserved structural domains. A sequence encoding an important proteolytic motif (RKRR) identified in the predicted peptide sequence of Hc-DAF-2 is consistent with that of the human IR, suggesting that it is involved in the formation of the IR complex. The Hc-daf-2 gene was transcribed in all life stages of H. contortus, with a significant up-regulation in the iL3 compared with other stages. To compare patterns of expression between Hc-daf-2 and Ce-daf-2, reporter constructs fusing the Ce-daf-2 or Hc-daf-2 promoter to sequence encoding GFP were microinjected into the N2 strain of C. elegans, and transgenic lines were established and examined. Both genes showed similar patterns of expression in amphidial (head) neurons, which relate to sensation and signal transduction. Further study by heterologous genetic complementation in a daf-2-deficient strain of C. elegans (CB1370) showed partial rescue of function by Hc-daf-2. Taken together, these findings provide a first insight into the roles of Hc-daf-2/Hc-DAF-2 in the biology and development of H. contortus, particularly in the transition to parasitism.

SUBMITTER: Li F

PROVIDER: S-EPMC4516220 | biostudies-literature | 2014 Jun

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Hc-daf-2 encodes an insulin-like receptor kinase in the barber's pole worm, Haemonchus contortus, and restores partial dauer regulation.

Li Facai F Lok James B JB Gasser Robin B RB Korhonen Pasi K PK Sandeman Mark R MR Shi Deshi D Zhou Rui R Li Xiangrui X Zhou Yanqin Y Zhao Junlong J Hu Min M

International journal for parasitology 20140412 7

Infective L3s (iL3s) of parasitic nematodes share common behavioural, morphological and developmental characteristics with the developmentally arrested (dauer) larvae of the free-living nematode Caenorhabditis elegans. It is proposed that similar molecular mechanisms regulate entry into or exit from the dauer stage in C. elegans, and the transition from free-living to parasitic forms of parasitic nematodes. In C. elegans, one of the key factors regulating the dauer transition is the insulin-like ...[more]

PMID: 24727120

Similar Datasets

Project description:BackgroundSignalling pathways have been extensively investigated in the free-living nematode Caenorhabditis elegans, but very little is known about these pathways in parasitic nematodes. Here, we constructed a model for the dauer-associated signalling pathways in an economically highly significant parasitic worm, Haemonchus contortus.MethodsGuided by data and information available for C. elegans, we used extensive genomic and transcriptomic datasets to infer gene homologues in the dauer-associated pathways, explore developmental transcriptomic, proteomic and phosphoproteomic profiles in H. contortus and study selected molecular structures.ResultsThe canonical cyclic guanosine monophosphate (cGMP), transforming growth factor-β (TGF-β), insulin-like growth factor 1 (IGF-1) and steroid hormone signalling pathways of H. contortus were inferred to represent a total of 61 gene homologues. Compared with C. elegans, H. contortus has a reduced set of genes encoding insulin-like peptides, implying evolutionary and biological divergences between the parasitic and free-living nematodes. Similar transcription profiles were found for all gene homologues between the infective stage of H. contortus and dauer stage of C. elegans. High transcriptional levels for genes encoding G protein-coupled receptors (GPCRs), TGF-β, insulin-like ligands (e.g. ins-1, ins-17 and ins-18) and transcriptional factors (e.g. daf-16) in the infective L3 stage of H. contortus were suggestive of critical functional roles in this stage. Conspicuous protein expression patterns and extensive phosphorylation of some components of these pathways suggested marked post-translational modifications also in the L3 stage. The high structural similarity in the DAF-12 ligand binding domain among nematodes indicated functional conservation in steroid (i.e. dafachronic acid) signalling linked to worm development.ConclusionsTaken together, this pathway model provides a basis to explore hypotheses regarding biological processes and regulatory mechanisms (via particular microRNAs, phosphorylation events and/or lipids) associated with the development of H. contortus and related nematodes as well as parasite-host cross talk, which could aid the discovery of new therapeutic targets.

Project description:BackgroundThe strongylid nematode Haemonchus contortus is a parasite of major concern for modern livestock husbandry because hostile environmental conditions may induce diapause in the early fourth-stage larvae.MethodsA new gene Hc-daf-22 was identified which is the homologue of Ce-daf-22 and human SCPx. Genome walking and RACE were performed to obtain the whole cDNA and genomic sequence of this gene. Using qRT-PCR with all developmental stages as templates to explore the transcription level and micro-injection was applied to confirm the promoter activity of the 5'-flanking region. Overexpression, rescue and RNA interference experiments were performed in N2, daf-22 mutant (ok 693) strains of C. elegans to study the gene function of Hc-daf-22.ResultsThe full length gene of Hc-daf-22 (6,939 bp) contained 16 exons separated by 15 introns, and encoded a cDNA of 1,602 bp (533 amino acids, estimated at about 59.3 kDa) with a peak in L3 and L4 in transcriptional level. The Hc-DAF-22 protein was consisted of a 3-oxoacyl-CoA thiolase domain and a SCP2 domain and evolutionarily conserved. The 1,548 bp fragment upstream of the 5'-flanking region was confirmed to have promoter activity compared with 5'-flanking region of Ce-daf-22. The rescue experiment by micro-injection of daf-22 (ok693) mutant strain showed significant increase in body size and brood size in the rescued worms with significantly reduced or completely absent fat granules confirmed by Oil red O staining, indicating that Hc-daf-22 could partially rescue the function of Ce-daf-22. Furthermore, RNAi with Hc-daf-22 could partially silence the endogenous Ce-daf-22 in N2 worms and mimic the phenotype of daf-22 (ok693) mutants.ConclusionThe gene Hc-daf-22 was isolated and its function identified using C. elegans as a model organism. Our results indicate that Hc-daf-22 shared similar characteristics and function with Ce-daf-22 and may play an important role in peroxisomal β-oxidation and the development in H. contortus.

Project description:Despite their phylogenetic diversity, parasitic nematodes share attributes of longevity and developmental arrest (=hypobiosis) with free-living nematodes at key points in their life cycles, particularly in larval stages responsible for establishing infection in the host. Insulin-like signalling plays crucial roles in the regulation of life span and arrest (=dauer formation) in the free-living nematode, Caenorhabditis elegans. Insulin-like signalling in C. elegans negatively regulates the fork head boxO (FoxO) transcription factor encoded by daf-16, which is linked to initiating a dauer-specific pattern of gene expression. Orthologues of daf-16 have been identified in several species of parasitic nematode. Although function has been demonstrated for an orthologue from the parasitic nematode Strongyloides stercoralis (Rhabditida), the functional capabilities of homologues/orthologues in bursate nematodes (Strongylida) are unknown. In the present study, we used a genomic approach to determine the structures of two complete daf-16 orthologues (designated Hc-daf-16.1 and Hc-daf-16.2) and their transcripts in the parasitic nematode Haemonchus contortus, and assessed their function(s) using C. elegans as a genetic surrogate. Unlike the multiple isoforms of Ce-DAF-16 and Ss-DAF-16, which are encoded by a single gene and produced by alternative splicing, mRNAs encoding the proteins Hc-DAF-16.1 and Hc-DAF-16.2 are transcribed from separate and distinct loci. Both orthologues are transcribed in all developmental stages and both sexes of H. contortus, and the inferred proteins (603 and 556 amino acids) each contain a characteristic, highly conserved fork head domain. In spite of distinct differences in genomic organisation compared with orthologues in C. elegans and S. stercoralis, genetic complementation studies demonstrated here that Hc-daf-16.2, but not Hc-daf-16.1, could restore daf-16 function to a C. elegans strain carrying a null mutation at this locus. These findings are consistent with previous results for S. stercoralis and demonstrate functional conservation of the daf-16b orthologue between key parasitic nematodes from two different taxonomic orders and C. elegans. We conclude from these experiments that the fork head transcription factor DAF-16 and, by inference, other insulin-like signalling elements, are conserved in H. contortus, a parasitic nematode of paramount economic importance. We demonstrate that functionality is sufficiently conserved in Hc-DAF-16.2 that it can replace Ce-DAF-16 in promoting dauer arrest in C. elegans.

Dataset Information

Hc-daf-2 encodes an insulin-like receptor kinase in the barber's pole worm, Haemonchus contortus, and restores partial dauer regulation.

Publications

Hc-daf-2 encodes an insulin-like receptor kinase in the barber's pole worm, Haemonchus contortus, and restores partial dauer regulation.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets