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Nicotine-modulated subunit stoichiometry affects stability and trafficking of ?3?4 nicotinic receptor.

ABSTRACT: Heteromeric nAChRs are pentameric cation channels, composed of combinations of two or three ? and three or two ? subunits, which play key physiological roles in the central and peripheral nervous systems. The prototypical agonist nicotine acts intracellularly to upregulate many nAChR subtypes, a phenomenon that is thought to contribute to the nicotine dependence of cigarette smokers. The ?3?4 subtype has recently been genetically linked to nicotine dependence and lung cancer; however, the mode of action of nicotine on this receptor subtype has been incompletely investigated. Here, using transfected mammalian cells as model system, we characterized the response of the human ?3?4 receptor subtype to nicotine and the mechanism of action of the drug. Nicotine, when present at 1 mm concentration, elicited a ?5-fold increase of cell surface ?3?4 and showed a more modest upregulatory effect also at concentrations as low as 10 ?M. Upregulation was obtained if nicotine was present during, but not after, pentamer assembly and was caused by increased stability and trafficking of receptors assembled in the presence of the drug. Experimental determinations as well as computational studies of subunit stoichiometry showed that nicotine favors assembly of pentamers with (?3)2(?4)3 stoichiometry; these are less prone than (?3)3(?4)2 receptors to proteasomal degradation and, because of the presence in the ? subunit of an endoplasmic reticulum export motif, more efficiently transported to the plasma membrane. Our findings uncover a novel mechanism of nicotine-induced ?3?4 nAChR upregulation that may be relevant also for other nAChR subtypes.

SUBMITTER: Mazzo F

PROVIDER: S-EPMC6618676 | biostudies-literature | 2013 Jul

REPOSITORIES: biostudies-literature

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Nicotine-modulated subunit stoichiometry affects stability and trafficking of α3β4 nicotinic receptor.

Mazzo Francesca F Pistillo Francesco F Grazioso Giovanni G Clementi Francesco F Borgese Nica N Gotti Cecilia C Colombo Sara Francesca SF

The Journal of neuroscience : the official journal of the Society for Neuroscience 20130701 30

Heteromeric nAChRs are pentameric cation channels, composed of combinations of two or three α and three or two β subunits, which play key physiological roles in the central and peripheral nervous systems. The prototypical agonist nicotine acts intracellularly to upregulate many nAChR subtypes, a phenomenon that is thought to contribute to the nicotine dependence of cigarette smokers. The α3β4 subtype has recently been genetically linked to nicotine dependence and lung cancer; however, the mode o ...[more]

PMID: 23884938

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Project description:The CHRNA5-CHRNA3-CHRNB4 gene cluster, encoding the α5, α3, and β4 nicotinic acetylcholine receptor (nAChR) subunits, has been linked to nicotine dependence. The habenulo-interpeduncular (Hb-IPN) tract is particularly enriched in α3β4 nAChRs. We recently showed that modulation of these receptors in the medial habenula (MHb) in mice altered nicotine consumption. Given that β4 is rate-limiting for receptor activity and that single nucleotide polymorphisms (SNPs) in CHRNB4 have been linked to altered risk of nicotine dependence in humans, we were interested in determining the contribution of allelic variants of β4 to nicotine receptor activity in the MHb. We screened for missense SNPs that had allele frequencies >0.0005 and introduced the corresponding substitutions in Chrnb4. Fourteen variants were analyzed by co-expression with α3. We found that β4A90I and β4T374I variants, previously shown to associate with reduced risk of smoking, and an additional variant β4D447Y, significantly increased nicotine-evoked current amplitudes, while β4R348C, the mutation most frequently encountered in sporadic amyotrophic lateral sclerosis (sALS), showed reduced nicotine currents. We employed lentiviruses to express β4 or β4 variants in the MHb. Immunoprecipitation studies confirmed that β4 lentiviral-mediated expression leads to specific upregulation of α3β4 but not β2 nAChRs in the Mhb. Mice injected with the β4-containing virus showed pronounced aversion to nicotine as previously observed in transgenic Tabac mice overexpressing Chrnb4 at endogenous sites including the MHb. Habenular expression of the β4 gain-of-function allele T374I also resulted in strong aversion, while transduction with the β4 loss-of function allele R348C failed to induce nicotine aversion. Altogether, these data confirm the critical role of habenular β4 in nicotine consumption, and identify specific SNPs in CHRNB4 that modify nicotine-elicited currents and alter nicotine consumption in mice.

Dataset Information

Nicotine-modulated subunit stoichiometry affects stability and trafficking of ?3?4 nicotinic receptor.

Publications

Nicotine-modulated subunit stoichiometry affects stability and trafficking of α3β4 nicotinic receptor.

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