A signal peptide missense mutation associated with nicotine dependence alters α2*-nicotinic acetylcholine receptor function.
A cytosine to thymidine (C → T) missense mutation in the signal peptide (SP) sequence (rs2472553) of the nicotinic acetylcholine receptor (nAChR) α2 subunit produces a threonine-to-isoleucine substitution (T22I) often associated with nicotine dependence (ND). We assessed effects on function of α2*-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 α:β subunit cRNAs injected) α2β2- or α2β4-nAChR and increased for putative high sensitivity (HS; 1:10 α:β subunit ratio) α2β2- (5.7-15-fold) or α2β4- (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 α2(T22I)β2-nAChR or for either α2*-nAChR subtype formed in the presence of equal amounts of subunit cRNA, slightly decreased for LS α2(T22I)β4-nAChR, but increased 1.4-2.4-fold for HS α2(T22I)β4-nAChR relative to receptors containing wild-type α2 subunits. These effects suggest that the α2 subunit SP mutation generally favors formation of LS receptor isoforms. We hypothesize that lower sensitivity of human α2*-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.
Department of Psychiatry and Neurobehavioral Sciences, School of Medicine, University of Virginia, Charlottesville, VA 22911, USA.
Bhagirathi
B
Dash
Division of Neurobiology, Barrow Neurological Institute, Phoenix, AZ 85013, USA.
Ronald J
RJ
Lukas
Department of Psychiatry and Neurobehavioral Sciences, School of Medicine, University of Virginia, Charlottesville, VA 22911, USA. Electronic address: ml2km@virginia.edu.
Ming D
MD
Li
D000109:Acetylcholine / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D020022:Genetic Predisposition to Disease; D006801:Humans; D008464:Mecamylamine / Q000494:pharmacology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D020125:Mutation, Missense; D009419:Nerve Tissue Proteins / Q000378:metabolism; D009538:Nicotine / Q000494:pharmacology; D018722:Nicotinic Agonists / Q000494:pharmacology; D018733:Nicotinic Antagonists / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D020641:Polymorphism, Single Nucleotide; D011978:Receptors, Nicotinic / Q000235:genetics* / Q000378:metabolism*; D013045:Species Specificity; D014029:Tobacco Use Disorder / Q000235:genetics*; D014982:Xenopus laevis
D000109:Acetylcholine; D008464:Mecamylamine; D009419:Nerve Tissue Proteins; D009538:Nicotine; D018722:Nicotinic Agonists; D018733:Nicotinic Antagonists; D011978:Receptors, Nicotinic; C436489:CHRNA2 protein, human; C447159:CHRNB4 protein, human
Electrophysiology; Missense mutation; Nicotinic acetylcholine receptor; Receptor structure-function; Signal peptide; Single nucleotide polymorphism