Dash_2014_Neuropharmacol_79_715

A cytosine to thymidine (C --> T) missense mutation in the signal peptide (SP) sequence (rs2472553) of the nicotinic acetylcholine receptor (nAChR) alpha2 subunit produces a threonine-to-isoleucine substitution (T22I) often associated with nicotine dependence (ND). We assessed effects on function of alpha2*-nAChR ('*'indicates presence of additional subunits) of this mutation, which could alter SP cleavage, RNA/protein secondary structure, and/or efficiency of transcription, translation, subunit assembly, receptor trafficking or cell surface expression. Two-electrode voltage clamp analyses indicate peak current responses to ACh or nicotine are decreased 2.8-5.8-fold for putative low sensitivity (LS; 10:1 ratio of alpha:beta subunit cRNAs injected) alpha2beta2- or alpha2beta4-nAChR and increased for putative high sensitivity (HS; 1:10 alpha:beta subunit ratio) alpha2beta2- (5.7-15-fold) or alpha2beta4- (1.9-2.2-fold) nAChR as a result of the mutation. Agonist potencies are decreased 1.6-4-fold for putative LS or HS alpha2(T22I)beta2-nAChR or for either alpha2*-nAChR subtype formed in the presence of equal amounts of subunit cRNA, slightly decreased for LS alpha2(T22I)beta4-nAChR, but increased 1.4-2.4-fold for HS alpha2(T22I)beta4-nAChR relative to receptors containing wild-type alpha2 subunits. These effects suggest that the alpha2 subunit SP mutation generally favors formation of LS receptor isoforms. We hypothesize that lower sensitivity of human alpha2*-nAChR to nicotine could contribute to increased susceptibility to ND. To our knowledge this is the first report of a SP mutation having a functional effect in a member of cys-loop family of ligand-gated ion channels.