MUTATIONS OF MUSCARINIC CHOLINERGIC RECEPTOR GENES

这个项目的总体目标是确定分子 激活、信号转导和调节的机制 原型G蛋白偶联受体(GPCR)，即人毒扁豆碱 胆碱能HM1受体。胆碱能缺陷是老年人的一个标志 痴呆症，如阿尔茨海默氏症，以及对 胆碱能神经传递在治疗的发展中是必需的 战略。这个项目使用了一种基因方法来确定 通过广泛的突变分析HM1受体的功能域， WHIT专注于对HM1仍知之甚少的三个关键过程 具体而言，以及一般的GPCRs。第一个包括分子 G蛋白激活受体和信号转导的机制 和第二信使通道。第二，作为激活的结果， 受体经历快速的细胞运输，即隔离， 内部化和再循环。第三个也是较慢的过程涉及 功能受体的破坏(下调)。快速受体 在几个被测试的组织中没有检测到HM1的脱敏作用，以及 因此，这里没有对其进行研究。这个实验室已经建造了 大量的HM1突变体首次允许NE解剖这些 并确定几个必需的受体结构域的位置。 其中包括第二个细胞内环的中心部分(I2)。 出人意料地发现，它在G蛋白激活中起着主要作用 和内部化，一个位于i3循环中间的S/T富集域 监管内部化和随后的下调，以及 I3环的结构域，它介导了不同的第二条途径 降低监管。因为尚未为任何 GPCRs，它们的完整特征将显著增加我们的 了解gpr的调控。相似的受体结构域存在于大多数 GPCRs，以及这些新领域的一般意义因此 通过选择的附加受体的突变分析来确定d 这个大家庭中的基因。因为受监管的i3环域 包括几个丝氨酸和苏氨酸残基，参与 蛋白激酶在内化过程中的磷酸化作用 将会受到考验。这些研究将扩展到分析 HM2、HM3、β-2肾上腺素受体和促甲状腺激素的相似结构域 释放激素(TRH)受体以测试 GPCRs的功能域。这项研究的结果将澄清 HM1的分子机制及其功能意义 激活和贩卖手机。负责任的受体结构域 将作为一种工具来分离目标蛋白 受体的功能。了解HM1受体调控知识将有所裨益 M受体激动剂治疗阿尔茨海默病。

The overall objective of this project is to determine the molecular mechanism of activation, signal transduction, and regulation of a prototype G protein coupled receptor (GPCR), i.e., the human muscarinic cholinergic Hm1 receptor. Cholinergic deficits are a hallmark of senile dementias such as Alzheimer, and a fundamental understanding of cholinergic neurotransmission is needed in development of therapeutic strategies. This project employees a genetic approach to determine the functional domains of the Hm1 receptor by extensive mutational analysis, whit focus on three key processes that remain poorly understood for Hm1 specifically and for GPCRs in general. The first includes the molecular mechanisms of receptor activation and signal transduction via G proteins and second messenger pathways. Second, as a result of activation, the receptors undergo rapid cellular trafficking, i.e., sequestration, internalization, and recycling. The third and slower process involves the destruction of functional receptor (down-regulation_. Rapid receptor desensitization is not detectable for Hm1 in several tissue tested, and it is therefore not studied here. This laboratory has constructed numerous Hm1 mutants that permit ne for the first time to dissect these pathways and define the location of several requisite receptor domains. These include the central portion of the second intracellular loop (i2) which was unexpectedly found to play a major role in G protein activation and internalization, an S/T rich domain in the middle of the i3 loop which regulation internalization and subsequent downregulation, and a domain of the i3 loop which mediates a distinct second pathway of downregulation. As such domains have not been identified for any of the GPCRs, their complete characterization will add significantly to our understanding of GPCR regulation. Similar receptor domains exist in most GPCRs, and the general significance of these novel domains will therefore be determine d by mutational analysis of selected additional receptor genes within this large family. Because the regulatory i3 loop domain includes several serine and threonine residues, the involvement of protein kinase mediated phosphorylation in the internalization process will be tested. These studies will be extended to the analysis of similar domains in Hm2, Hm3, beta-2 adrenoceptor, and the thyrotropin releasing hormone (TRH) receptor to test the general validity of the functional domains for GPCRS. The results from this study will clarify the molecular mechanisms and the functional significance of Hm1 activation and cellular trafficking. The responsible receptor domains will then serve as a tool to isolate the target protein mediating receptor functions. Knowledge of Hm1 receptor regulation will benefit therapy of Alzheimer disease with muscarinic agonist.