Abstract
The α7 nicotinic acetylcholine receptor subunit (CHRNA7) gene harbors a high degree of polymorphism. In this study, we found a novel variant (1267 G to A) in exon 10 of the CHRNA7 gene in a Japanese population. This variant results in glycine-to-serine substitution at position 423 (G423S) located in the large cytoplasmic loop of the protein. To clarify the possibility that the G423S mutation alters the pharmacological properties of α7 receptors, acetylcholine (ACh)-elicited current through α7-G423S mutant receptors expressed in Xenopus laevis oocytes was measured using the two-electrode voltage-clamp technique. We found that the current elicited by ACh (1 mM, 5 s) through α7-G423S receptors, but not through α7 receptors, was significantly decreased by treatment with a protein kinase C activator, phorbol-12-myristate-13-acetate (PMA, 10–30 nM). In addition, PMA (10 nM) selectively promoted a progressive decrease in α7-G423S current induced by repetitive application of ACh pulses (1 mM, 0.1 s, 0.17–0.33 Hz) compared with α7 current. PMA also enhanced the inactivation of α7-G423S mutant receptors induced by a prolonged application of choline (30 μM) without affecting α7 receptor responses. Western blot analysis showed that the treatment with PMA (30 nM) increased the serine phosphorylation level of the α7-G423S mutant receptors but not that of the wild-type receptors. These findings demonstrate that the G423S mutation promotes receptor desensitization by a protein kinase C-dependent mechanism. Thus, we provide the first evidence that a variant in the human CHRNA7 gene alters the function of α7 nicotinic receptors.
Footnotes
-
This work was supported in part by a Grant-in-Aid for Scientific Research from Japan Society for the Promotion of Science (to H.T.) and by the Smoking Research Foundation (Japan) (to I.K.).
-
H.T. and S.K. contributed equally to this work.
-
Article, publication date, and citation information can be found at http://molpharm.aspetjournals.org.
-
doi:10.1124/mol.106.030866.
-
ABBREVIATIONS: SNP, single nucleotide polymorphism; ACh, acetylcholine; DMSO, dimethyl sulfoxide; EGFP, enhanced green fluorescence protein; MLA, methyllycaconitine; NMDA, N-methyl-d-aspartate; ORF, open reading frame; PMA, phorbol-12-myristate-13-acetate; PVDF, polyvinylidene fluoride; PCR, polymerase chain reaction; SSCP, single-strand conformation polymorphism; bp, base pair(s).
- Received September 13, 2006.
- Accepted November 28, 2006.
- The American Society for Pharmacology and Experimental Therapeutics
MolPharm articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|