Habecker B A, Tietje K M, van Koppen C J, Creason S A, Goldman P S, Migeon J C, Parenteau L A, Nathanson N M
Department of Pharmacology, University of Washington, Seattle 98195.
Life Sci. 1993;52(5-6):429-32. doi: 10.1016/0024-3205(93)90298-h.
The regulation of expression and function of the muscarinic acetylcholine receptor has been studied using several different systems. The role of glycosylation of the m2 receptor was examined by removal of glycosylation sites using site-directed mutagenesis followed by expression in stably transfected cells. The results demonstrated that glycosylation was not required for the synthesis and appearance of the receptors on the cell surface or for the coupling of the receptors to inhibition of adenylyl cyclase activity. Site-directed mutagenesis also was used to demonstrate that the single cysteine in the carboxy terminal domain of the m2 receptor was not required for receptor function, thus rendering unlikely a model suggesting a requirement for palmitoylation of this cysteine in receptor function. The muscarinic receptors expressed in embryonic chick heart were identified by molecular cloning. Two genes were initially identified which are expressed in chick heart and correspond to the chick m2 and m4 receptors. Experiments using the polymerase chain reaction to identify low abundance mRNAs indicate that at least one addition receptor gene is expressed in chick heart. In cell culture, activation of the muscarinic receptors decreases the levels of mRNA encoding the cm2 and cm4 receptors. This probably results from decreased gene transcription due to both mAChR-mediated inhibition of adenylyl cyclase and mAChR-mediated stimulation of phospholipase C. The elucidation of the factors which regulate the expression and function of muscarinic acetylcholine receptors (mAChR) is of obvious importance in understanding the mechanisms underlying cholinergic transmission. In this chapter, we will describe studies on the expression and function of wild type and mutant muscarinic receptors, the molecular characterization of mAChR expressed in chick heart, and the regulation of mAChR gene expression in response to muscarinic receptor activation.
已使用多种不同系统对毒蕈碱型乙酰胆碱受体的表达和功能调节进行了研究。通过定点诱变去除糖基化位点，随后在稳定转染的细胞中进行表达，从而研究了m2受体糖基化的作用。结果表明，受体在细胞表面的合成和出现以及受体与腺苷酸环化酶活性抑制的偶联并不需要糖基化。定点诱变还用于证明m2受体羧基末端结构域中的单个半胱氨酸对于受体功能并非必需，因此一种认为该半胱氨酸在受体功能中需要棕榈酰化的模型不太可能成立。通过分子克隆鉴定了在胚胎鸡心脏中表达的毒蕈碱受体。最初鉴定出两个在鸡心脏中表达且对应于鸡m2和m4受体的基因。使用聚合酶链反应鉴定低丰度mRNA的实验表明，鸡心脏中至少还表达一种额外的受体基因。在细胞培养中，毒蕈碱受体的激活会降低编码cm2和cm4受体的mRNA水平。这可能是由于mAChR介导的腺苷酸环化酶抑制和mAChR介导的磷脂酶C刺激导致基因转录减少所致。阐明调节毒蕈碱型乙酰胆碱受体（mAChR）表达和功能的因素对于理解胆碱能传递的潜在机制显然具有重要意义。在本章中，我们将描述关于野生型和突变型毒蕈碱受体的表达和功能、在鸡心脏中表达的mAChR的分子特征以及毒蕈碱受体激活后mAChR基因表达调节的研究。
