Intermolecular Activation of GPCR

GPCR 的分子间激活

• 批准号: 7230383
• 负责人: Tae H. Ji
• 金额: $ 0.85万
• 依托单位: UNIVERSITY OF KENTUCKY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-08-01 至 2009-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7230383
• 关键词: G protein; SDS polyacrylamide gel electrophoresis; affinity labeling; binding proteins; binding sites; biological signal transduction; cell surface receptors; cyclic AMP; enzyme activity; follicle stimulating hormone; hormone binding protein; hormone receptor; inositol phosphates; intermolecular interaction; luteinizing hormone; mass spectrometry; organism; protein purification; protein sequence; receptor coupling; receptor expression; thyrotropin; tissue /cell culture; transfection

DESCRIPTION (provided by applicant): G protein coupled receptors (GPCRs) are the major hormone receptors, constituting approximately 3 percent of the human genome. It has been a dogma that, when a hormone1 binds to a GPCR, the liganded receptor activates itself to generate hormone signals (cis-activation). In contrast to this dogmatic cis-activation, the evidence is emerging that a liganded receptor molecule can intermolecularly activate nonliganded receptor molecules (trans-activation). There are only several reports on trans-activation of GPCRs: two on the thrombin receptors and our reports on the LH receptor (LHR). LHR consists of two equal halves, the extracellular N-terminal exo-domain and the membrane associated heptahelical endo-domain. Hormone binds to the exo-domain, whereas the endo-domain generates two distinct hormone signals: one for adenylyl cyclase to produce cAMP (cAMP signal) and the other for phospholipase cbeta to produce diacylglycerol and inositol phosphate (IP signal). Trans-activation is difficult to test, because it has to be differentiated from cis-activation and hormone binding needs to be distinguished from signal generation. LHR meets these requirements and offers a unique model to test trans-activation. We have established a large library of two mutant groups, one deficient in hormone binding and the other deficient in signal generation. Co-expression of a binding deficient mutant and a signal deficient mutant in a cell rescues signal generation. Our preliminary data show that the trans-activation generates either the cAMP signal or IP signal, but not both. Aim 1 will test the hypotheses that trans-activation occurs not only to the mutant receptor pairs but also to the wild type receptor and that the signal selection is specific. Aim 2 will define the mechanistics of the trans-activation and signal selection. These new paradigms will have broad implications on the mechanisms of GPCR signal generation and development of new receptor therapeutics, particularly to control a specific signal without invoking other signals. Receptors generate multiple signals, which is a source of undesirable/toxic side effects of hormone therapeutics.

描述（由申请人提供）：G 蛋白偶联受体 (GPCR) 是主要的激素受体，约占人类基因组的 3%。人们普遍认为，当激素 1 与 GPCR 结合时，配体受体会自我激活以产生激素信号（顺式激活）。与这种教条式顺式激活相反，越来越多的证据表明配体受体分子可以分子间激活非配体受体分子（反式激活）。关于 GPCR 反式激活的报道仅有几篇：两篇关于凝血酶受体的报道，以及我们关于 LH 受体 (LHR) 的报道。 LHR 由两个相等的半部分组成，即细胞外 N 端外结构域和膜相关的七螺旋内结构域。激素与外结构域结合，而内结构域产生两种不同的激素信号：一种用于腺苷酸环化酶产生 cAMP（cAMP 信号），另一种用于磷脂酶 cbeta 产生二酰基甘油和磷酸肌醇（IP 信号）。 反式激活很难测试，因为它必须与顺式激活区分开来，并且激素结合需要与信号产生区分开。 LHR 满足这些要求，并提供了一个独特的模型来测试反式激活。我们建立了一个包含两个突变体群体的大型文库，一个缺乏激素结合，另一个缺乏信号生成。细胞中结合缺陷突变体和信号缺陷突变体的共表达挽救了信号的产生。我们的初步数据表明反式激活会产生 cAMP 信号或 IP 信号，但不会同时产生两者。目标 1 将测试以下假设：反式激活不仅发生在突变型受体对上，而且也发生在野生型受体上，并且信号选择是特异性的。目标 2 将定义反式激活和信号选择的机制。这些新范例将对 GPCR 信号生成机制和新受体疗法的开发产生广泛影响，特别是在不调用其他信号的情况下控制特定信号。受体产生多种信号，这是激素治疗不良/毒副作用的根源。