Cloning and structural analysis of partial acetylcholine receptor subunit genes from the parasitic nematode Teladorsagia circumcincta

Nematode nicotinic acetylcholine receptors (nAChRs) are the sites of action for the anthelmintic drug levamisole. Recent findings indicate that the mo lecular mechanism of levamisole resistance may involve changes in the numbe r and/or functions of target nAChRs. Accordingly, we have used an RT-PCR ap proach to isolate and characterise partial cDNA clones (tca-1 and tca-2) en coding putative nAChR subunits from the economically important trichostrong yloid, Teladorsagia circumcincta. The predicted tca-1 gene product is a 248 aa fragment (TCA-1) which contains structural motifs typical of ligand-bin ding (alpha-) subunits, and which shows very high sequence similarities (98 .8 and 97.2% amino acid identities) to the alpha -subunits encoded by tar-1 and hca-1 from Trichostrongylus colubriformis and Haemonchus contortus, re spectively. Sequence analyses of partial tca-1 cDNAs from one levamisole-re sistant and two susceptible populations of ir: circumcincta revealed polymo rphism at the predicted amino acid level, but there was no apparent associa tion of any particular tca-1 allele with resistance, tca-2 encodes a 67 aa fragment (TCA-2) containing the TM4 transmembrane domain and carboxyl termi nus of a putative nAChR structural (non-alpha) subunit. The deduced amino a cid sequence of TCA-2 shows highest similarity (75% amino acid identity) to ACR-2, a structural subunit involved in forming levamisole-gated ion chann els in Caenorhabditis elegans, but low similarity (43% identity) to the cor responding regions of TAR-1 and HCA-1. tca-2 is the first nAChR subunit gen e of this type to be isolated from parasitic nematodes, and it provides a b asis for further characterisation of structural subunits in trichostrongylo ids. (C) 2001 Elsevier Science B.V. All rights reserved.