Abstract
The invertebrate glutamate-gated chloride channels (GluCls) are receptor molecules and targets for the avermectin–milbemycin (AM) group of anthelmintics. Mutations in GluCls are associated with ivermectin resistance in the soil dwelling nematode Caenorhabditis elegans and the parasitic nematode Cooperia oncophora. In this study, full-length cDNAs encoding α and β subunits of GluCl were cloned and sequenced in Cylicocyclus nassatus, a common and important cyathostomin nematode parasite of horses. Both genes possess the sequence characteristics typical of GluCls, and phylogenetic analysis confirms that these genes are evolutionarily closely related to GluCls of other nematodes and flies. Complete coding sequences of C. nassatus GluCl-α and GluCl-β were subcloned into pTL1 mammalian expression vector, and proteins were expressed in COS-7 cells. Ivermectin-binding characteristics were determined by incubating COS-7 cell membranes expressing C. nassatus GluCl-α and GluCl-β proteins with [ 3H]ivermectin. In competitive binding experiments, fitting the data to a one site competition model, C. nassatus GluCl-α was found to bind [ 3H]ivermectin with a high amount of displaceable binding (IC 50 = 208 pM). Compared to the mock-transfected COS-7 cells, the means of [ 3H]ivermectin binding were significantly different for C. nassatus GluCl-α and the Haemonchus contortus GluCl (HcGluCla) ( p = 0.018 and 0.023, respectively) but not for C. nassatus GluCl-β ( p = 0.370). This is the first report of orthologs of GluCl genes and in vitro expression of an ivermectin-binding protein in a cyathostomin species. These data suggest the likelihood of a similar mechanism of action of AM drugs in these parasites, and suggest that mechanisms of resistance may also be similar.
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.