Isoform-specific mechanisms of α3β4*-nicotinic acetylcholine receptor modulation by the prototoxin lynx1

This study investigates-for the first time to our knowledge-the existence and mechanisms of functional interactions between the endogenous mammalian prototoxin, lynx1, and α3- and β4-subunit-containing human nicotinic acetylcholine receptors (α3β4*-nAChRs). Concatenated gene constructs were used to express precisely defined α3β4*-nAChR isoforms (α3β4)₂β4-, (α3β4)₂α3-, (α3β4)₂α5(398D)-, and (α3β4)₂α5(398N)-nAChR in Xenopus oocytes. In the presence or absence of lynx1, α3β4*-nAChR agonist responses were recorded by using 2-electrode voltage clamp and single-channel electrophysiology, whereas radioimmunolabeling measured cell-surface expression. Lynx1 reduced (α3β4)₂β4-nAChR function principally by lowering cell-surface expression, whereas single-channel effects were primarily responsible for reducing (α3β4)₂α3-nAChR function [decreased unitary conductance (≥50%), altered burst proportions (3-fold reduction in the proportion of long bursts), and enhanced closed dwell times (3- to 6-fold increase)]. Alterations in both cell-surface expression and single-channel properties accounted for the reduction in (α3β4)₂α5-nAChR function that was mediated by lynx1. No effects were observed when α3β4*-nAChRs were coexpressed with mutated lynx1 (control). Lynx1 is expressed in the habenulopeduncular tract, where α3β4*-α5*-nAChR subtypes are critical contributors to the balance between nicotine aversion and reward. This gives our findings a high likelihood of physiologic significance. The exquisite isoform selectivity of lynx1 interactions provides new insights into the mechanisms and allosteric sites [α(-)-interface containing] by which prototoxins can modulate nAChR function.-George, A. A., Bloy, A., Miwa, J. M., Lindstrom, J. M., Lukas, R. J., Whiteaker, P. Isoform-specific mechanisms of α3β4*-nicotinic acetylcholine receptor modulation by the prototoxin lynx1.

GPI-linked proteins; TEVC; concatenated receptor subunits; neurotoxins; single-channel electrophysiology.