Reporter Mutation Studies Show That Nicotinic Acetylcholine Receptor (nAChR) α5 Subunits and/or Variants Modulate Function of α6*-nAChR*

To further the understanding of functional alpha 6 alpha 5*-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 alpha 5 subunits or chimeric alpha 5/beta 3 subunits (in which the human alpha 5 subunit N-terminal, extracellular domain is linked to the remaining domains of the human beta 3 subunit) almost completely abolishes the very small amount of function seen for alpha 6 beta 4*-nAChR and does not induce function of alpha 6 beta 2*-nAChR. Coexpression with human alpha 5(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 alpha 6 beta 4*-nAChR or induce function of alpha 6 beta 2*-nAChR. However, coexpression with mutant chimeric alpha 5/beta 3(V9)'(S) subunits has a gain-of-function effect (higher functional expression and agonist sensitivity and spontaneous opening inhibited by mecamylamine) on alpha 6 beta 4*-nAChR. Moreover, N143D + M145V mutations in the alpha 6 subunit N-terminal domain enable alpha 5/beta 3(V9)'(S) subunits to have a gain-of-function effect on alpha 6 beta 2*-nAChR. nAChR containing chimeric alpha 6/alpha 3 subunits plus either beta 2 or beta 4 subunits have some function that is modulated in the presence of alpha 5 or alpha 5/beta 3 subunits. Coexpression with alpha 5/beta 3(V9)'(S) subunits has a gain-of-function effect more pronounced than that in the presence of alpha 5(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 alpha 6 alpha 5*-nAChR and provide tools for development of alpha 6*-nAChR-selective ligands that could be important in the treatment of nicotine dependence, and perhaps other neurological diseases.