Abstract

Accessory subunits in heteromeric nicotinic receptors (AChRs) do not take part in forming ACh binding sites. α5 and β3 subunits can function only as accessory subunits. We show that both α5 and β3 efficiently assemble in human α4β2^* AChRs expressed in permanently transfected human embryonic kidney (HEK) cell lines. Only (α4β2)₂α5, not (α4β2)₂β3 AChRs, have been detected in brain. The α4β2α5 line expressed 40% more AChRs than the parent α4β2 line and was equally sensitive to up-regulation by nicotine. The α4β2β3 line expressed 25-fold more AChRs than the parental line and could not be further up-regulated by nicotine. Relative sensitivity to activation by ACh depends on the accessory subunit, β2 conferring the greatest sensitivity, α5 less, and β3 and α4 much less. Accessory subunits form binding sites for positive allosteric modulators, as illustrated by the observation that α5 conferred high sensitivity to galanthamine. In the presence of α5 or β3, stable, partially degraded, dead end intermediates accumulated within the cells. These may have the form α5α4β2α5. The efficiency with which α5 and β3 assemble with α4 and β2 and the necessity of avoiding formation of potentially toxic intermediates may explain why α5 and β3 seem to be transcribed at low levels in brain. Autosomal dominant nocturnal frontal lobe epilepsy can be caused by the α4 mutation S247F. This mutant did not produce functional AChRs unless cells were cotransfected with α5, β3, or α6 to replace α4 as accessory subunit.