Heteromeric Neuronal Nicotinic Acetylcholine Receptors with Mutant β Subunits Acquire Sensitivity to α7-Selective Positive Allosteric Modulators

Homomeric α 7 nicotinic acetylcholine receptors (nAChR) have an intrinsically low probability of opening that can be overcome by α 7-selective positive allosteric modulators (PAMs), which bind at a site involving the second transmembrane domain (TM2). Mutation of a methionine that is unique to α 7 at the 15′ position of TM2 to leucine, the residue in most other nAChR subunits, largely eliminates the activity of such PAMs. We tested the effect of the reverse mutation (L15′M) in heteromeric nAChR receptors containing α 4 and β 2, which are the nAChR subunits that are most abundant in the brain. Receptors containing these mutations were found to be strongly potentiated by the α 7 PAM 3 a ,4,5,9 b -tetrahydro-4-(1-naphthalenyl)-3 H -cyclopentan[ c ]quinoline-8-sulfonamide (TQS) but insensitive to the alternative PAM 1-(5-chloro-2,4-dimethoxyphenyl)-3-(5-methylisoxazol-3-yl)-urea. The presence of the mutation in the β 2 subunit was necessary and sufficient for TQS sensitivity. The primary effect of the mutation in the α 4 subunit was to reduce responses to acetylcholine applied alone. Sensitivity to TQS required only a single mutant β subunit, regardless of the position of the mutant β subunit within the pentameric complex. Similar results were obtained when β 2L15′M was coexpressed with α 2 or α 3 and when the L15′M mutation was placed in β 4 and coexpressed with α 2, α 3, or α 4. Functional receptors were not observed when β 1L15′M subunits were coexpressed with other muscle nAChR subunits. The unique structure-activity relationship of PAMs and the α 4 β 2L15′M receptor compared with α 7 and the availability of high-resolution α 4 β 2 structures may provide new insights into the fundamental mechanisms of nAChR allosteric potentiation.