Neuronal Nicotinic Acetylcholine Receptors on Bovine Chromaffin Cells: Cloning, Expression, and Genomic Organization of Receptor Subunits
Neuronal Nicotinic Acetylcholine Receptors on Bovine Chromaffin Cells: Cloning, Expression, and Genomic Organization of Receptor Subunits
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DOI：
10.1046/j.1471-4159.1997.68020488.x
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发表时间：
1997-02
影响因子：
4.7
通讯作者：
A. Campos-Caro;F. I. Smillie;E. D. D. Toro-E.-D.-D.-Toro-2271684175;J. Rovira;F. Vicente-Agulló;J. Chapuli;J. Juiz;S. Sala;F. Sala;J. J. Ballesta-J.;Manuel Criado
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文献类型：
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作者：
A. Campos-Caro;F. I. Smillie;E. D. D. Toro-E.-D.-D.-Toro-2271684175;J. Rovira;F. Vicente-Agulló;J. Chapuli;J. Juiz;S. Sala;F. Sala;J. J. Ballesta-J.;Manuel Criado
摘要：来自牛肾上腺髓质嗜铬细胞的神经元烟碱乙酰胆碱受体在触发儿茶酚胺分泌中起主要作用。在本研究中，对其组成亚基进行了表征。除了我们之前克隆的 α3 亚基外，通过对肾上腺髓质 mRNA 进行逆转录 PCR 分析，还检测到 α5 和 β4 亚基的存在，但未检测到 β2 亚基的存在。原位杂交表明α3、α5和β4亚基在所有嗜铬细胞中共表达。将亚基 cRNA 共注射到非洲爪蟾卵母细胞中后，确定了 α5 和 β4 亚基的一级结构，并获得了功能性受体。与其他含 β4 的烟碱受体相比，牛 β4 亚基形成的烟碱受体对激动剂金雀花碱不敏感。最后，我们对 α3 和 α5 亚基的基因间区域进行了表征，它们与 β4 亚基一起形成了大鼠和鸡的基因簇。 RNase 检测和重叠 cDNA 的存在表明，在牛基因组中，α3 和 α5 基因在其 3' 端重叠。这一事实可能是由于多聚腺苷酸化信号较弱导致转录终止效率低下。
Abstract: Neuronal nicotinic acetylcholine receptors from bovine adrenomedullary chromaffin cells play a primary role in triggering catecholamine secretion. In the present study, their constituent subunits were characterized. In addition to the α3 subunit, which we have previously cloned, the presence of α5 and β4 but not of β2 subunits was detected by reverse transcription‐PCR analysis of mRNA from adrenal medulla. In situ hybridization indicated that α3, α5, and β4 subunits are coexpressed in all chromaffin cells. The primary structure of α5 and β4 subunits was determined and functional receptors were obtained upon coinjection of subunit cRNAs into Xenopus oocytes. In contrast to other β4‐containing nicotinic receptors, the ones formed by the bovine β4 subunit are insensitive to the agonist cytisine. Finally, we characterized the intergenic region of α3 and α5 subunits, which together with the β4 subunit, form a gene cluster in rats and chickens. RNase assays and the existence of overlapping cDNAs indicate that, in the bovine genome, the α3 and α5 genes overlap at their 3′ ends. This fact is probably due to inefficient transcription termination, as a result of weak polyadenylation signals.