Molecular Cloning and Characterization of Novel Glutamate-Gated Chloride Channel Subunits from Schistosoma mansoni

PLoS Pathogens

Joseph A. Dent¹  Claudia Wever¹  Robin N. Beech²  Timothy G. Geary²  Vanessa Dufour² 

[1] Centre for Host-Parasite Interactions, Department of Biology, McGill University, Montréal, Québec, Canada;Centre for Host-Parasite Interactions, Institute of Parasitology, McGill University – MacDonald Campus, Sainte-Anne-de-Bellevue, Québec, Canada

关键词: Schistosoma mansoni;    Xenopus oocytes;    Flatworms;    Phylogenetic analysis;    Gamma-aminobutyric acid;    Schistosoma;    Glutamate;    Chlorides;   

DOI  :  10.1371/journal.ppat.1003586

学科分类：生物科学（综合）

来源: Public Library of Science

【 摘 要 】

Cys-loop ligand-gated ion channels (LGICs) mediate fast ionotropic neurotransmission. They are proven drug targets in nematodes and arthropods, but are poorly characterized in flatworms. In this study, we characterized the anion-selective, non-acetylcholine-gated Cys-loop LGICs from Schistosoma mansoni. Full-length cDNAs were obtained for SmGluCl-1 (Smp_096480), SmGluCl-2 (Smp_015630) and SmGluCl-3 (Smp_104890). A partial cDNA was retrieved for SmGluCl-4 (Smp_099500/Smp_176730). Phylogenetic analyses suggest that SmGluCl-1, SmGluCl-2, SmGluCl-3 and SmGluCl-4 belong to a novel clade of flatworm glutamate-gated chloride channels (GluCl) that includes putative genes from trematodes and cestodes. The flatworm GluCl clade was distinct from the nematode-arthropod and mollusc GluCl clades, and from all GABA receptors. We found no evidence of GABA receptors in S. mansoni. SmGluCl-1, SmGluCl-2 and SmGluCl-3 subunits were characterized by two-electrode voltage clamp (TEVC) in Xenopus oocytes, and shown to encode Cl−-permeable channels gated by glutamate. SmGluCl-2 and SmGluCl-3 produced functional homomers, while SmGluCl-1 formed heteromers with SmGluCl-2. Concentration-response relationships revealed that the sensitivity of SmGluCl receptors to L-glutamate is among the highest reported for GluCl receptors, with EC50 values of 7–26 µM. Chloride selectivity was confirmed by current-voltage (I/V) relationships. SmGluCl receptors are insensitive to 1 µM ivermectin (IVM), indicating that they do not belong to the highly IVM-sensitive GluClα subtype group. SmGluCl receptors are also insensitive to 10 µM meclonazepam, a schistosomicidal benzodiazepine. These results provide the first molecular evidence showing the contribution of GluCl receptors to L-glutamate signaling in S. mansoni, an unprecedented finding in parasitic flatworms. Further work is needed to elucidate the roles of GluCl receptors in schistosomes and to explore their potential as drug targets.

【 授权许可】

CC BY