Reporter mutation studies show that nicotinic acetylcholine receptor (nAChR) α5 subunits and/or variants modulate function of α6*-nAChR

Abstract

To further the understanding of functional α6α5*- nicotinic acetylcholine receptors (nAChR; the asterisk (*) indicates known or possible presence of other subunits), we have heterologously expressed in oocytes different, mouse or human, nAChR subunit combinations. Coexpression with wild-type α5 subunits or chimeric α5/β3 subunits (in which the human α5 subunit N-terminal, extracellular domain is linked to the remaining domains of the human β3 subunit) almost completely abolishes the very small amount of function seen for α6β4*-nAChR and does not induce function of α6β2*-nAChR. Coexpression with human α5 V9,S subunits bearing a valine 290 to serine mutation in the 9′ position of the second transmembrane domain does not rescue the function of α6β4*-nAChR or induce function of α6β2*-nAChR. However, coexpression with mutant chimeric α5/β3 V9,S subunits has a gain-of-function effect (higher functional expression and agonist sensitivity and spontaneous opening inhibited by mecamylamine) on α6β4*- nAChR. Moreover, N143D + M145V mutations in the α6 subunit N-terminal domain enable α5/β3 V9,S subunits to have a gain-of-function effect on α6β2*-nAChR. nAChR containing chimeric α6/α3 subunits plus either β2 or β4 subunits have some function that is modulated in the presence of α5 or α5/β3 subunits. Coexpression with α5/β3 V9,S subunits has a gain-of-function effect more pronounced than that in the presence of α5 V9,S subunits. Gain-of-function effects are dependent, sometimes subtly, on the nature and apparently the extracellular, cytoplasmic, and/or transmembrane domain topology of partner subunits. These studies yield insight into assembly of functional α6α5*-nAChR and provide tools for development of α6*- nAChR-selective ligands that could be important in the treatment of nicotine dependence, and perhaps other neurological diseases. © 2011 by The American Society for Biochemistry and Molecular Biology, Inc.