Functional analysis of mutation and partial duplication of α7 nicotinic receptor subunit gene : implication in schizophrenia

1.α7烟碱受体亚单位（CHRNA 7）基因被认为是精神分裂症的易感基因。在这项研究中，我们发现了一个新的突变CHRNA 7基因在日本精神分裂症患者。由于突变导致受体胞质环中的氨基酸取代（G423 S），我们研究了在非洲爪蟾卵母细胞中表达的野生型（WT）和α7-G423 S突变体受体之间的功能差异。在存在蛋白激酶C（PKC）激活剂的情况下，α7激动剂的刺激诱导α7-G423 S突变体受体的磷酸化和抑制，但不诱导WT受体。这些结果表明G423 S突变通过蛋白激酶C依赖的机制促进受体脱敏. CHRNA 7基因是部分重复的，并与新基因FAM 7A形成杂合体（CHRFAM 7A）。本研究发现CHRFAM 7A基因cDNA在非洲爪蟾卵母细胞和人SH-SY 5 Y细胞中翻译成蛋白（dup-α7），并与α7受体亚基发生免疫共沉淀。此外，电生理和生化研究表明，dup-α7蛋白的表达引起α7受体活性的降低。这些结果提示dup-α7蛋白可能通过一种新的机制调节α7受体的活性.我们研究了几种两栖类和海鞘类生物碱对神经元烟碱受体的作用，发现生物碱（+）-205B是α7烟碱受体的特异性选择性阻断剂.我们发现，慢性尼古丁刺激可通过t烟碱受体引起下丘脑GT 1 -7细胞促性腺激素释放激素表达增强，并上调血管平滑肌细胞α7烟碱受体，提示CHRNA 7基因G423 S突变和dup-α7蛋白过量干扰α7烟碱受体功能，是中枢胆碱能障碍的易感因素。例如精神分裂症。

1. The α7 nicotinic receptor subunit (CHRNA7) gene has been considered as a susceptible gene for schizophrenia. In this study, we found a novel mutation in CHRNA7 gene in a Japanese schizophrenic patient. Since the mutation results in amino acid substitution (G423S) in the receptor cytoplasmic loop, we investigated functional differences between wild-type (WT) and the α7-G423S mutant receptor expressed in Xenopus oocytes. In the presence of a protein kinase C (PKC) activator, stimulation with α7 agonist induces phosphorylation and inhibition of the α7-G423S mutant receptor, but not WT receptor. These results demonstrate that the G423S mutation promotes the receptor desensitization by a protein kinase C-dependent mechanism.2. The CHRNA7 gene is partially duplicated and forms a hybrid (CHRFAM7A) with a novel gene FAM7A. In this study, we found that the cDNA of CHRFAM7A gene was translated into protein (dup-α7) in Xenopus oocytes and human SH-SY5Y cells, and that the protein was coimmunoprecipitated with α7 receptor subunit. Moreover, electrophysiological and biochemical studies demonstrated that the expression of dup-α7 protein caused the reduction of α7 receptor activity. These findings suggest the existence of a novel mechanism for regulating α7 receptor activity via dup-α7 protein.3. We examined effects of several alkaloids from amphibians and ascidians on neuronal nicotinic receptors, and found that the alkaloid (+)-205B is a patent and selective blocker of α7 nicotinic receptor.4. We found that chronic nicotine stimulation caused enhanced expression of gonadotropin releasing hormone in hypothalamic GT1-7 cells via t nicotinic receptor, and upregulation of α7 nicotinic receptor in vascular smooth muscle cells.These results suggest that the G423S mutation in CHRNA7 and the dup-α7 protein excess perturb the function of α7 nicotinic receptor and serve as predisposing factors for central cholinergic disorders, such as schizophrenia.