Molecular Basis of RGS Protein Function in the Striatum

描述（由申请人提供）：纹状体中的G蛋白信号通路介导一系列关键的神经元过程，这些过程控制行为、运动、疼痛感知和药物滥用和成瘾。这些通路的正常功能取决于G蛋白信号传导调节蛋白（regulatory of G protein signaling， RGS）对信号持续时间的严格控制。我们的长期目标是阐明控制神经元中RGS蛋白功能的机制，作为理解神经系统疾病过程及其治疗手段的必要前提。这些研究的主要重点是RGS9-2，这是一种纹状体特异性调节因子，通过多巴胺和阿片受体系统至关重要地控制信号效率，这些系统被滥用药物所利用。然而，RGS9-2功能的分子机制在很大程度上是未知的，需要进一步阐明，以更好地了解成瘾的神经化学基础。为此，我们最近发现纹状体中的RGS9-2与一种新的神经元蛋白复合物存在，我们将其命名为R7结合蛋白（R7BP）。初步数据表明，R7BP在神经元中是RGS9-2功能的关键调节剂。这一假设将通过解决以下具体目标来检验：探讨RGS9-2的稳定性和定位机制。拟开展的研究将确定R7BP绕过RGS9-2蛋白水解的机制，并验证R7BP也决定RGS9-2在神经元中的定位的假设。2. 了解R7BP在调控RGS9-2催化活性中的作用。我们将采用体外酶法和蛋白-蛋白相互作用试验相结合的方法来确定R7BP在调节RGS9-2刺激G蛋白GTPase活性的能力中的动力学机制。3. 进一步表征纹状体神经元G蛋白失活复合物的分子组成。我们将使用蛋白质组学方法来鉴定涉及RGS9-2/R7BP复合物的大分子集合的其他组分。这些研究将有助于理解纹状体中的信号调节，并对药物滥用和神经系统疾病引起的G蛋白信号中断的分子机制产生深入的见解。

DESCRIPTION (provided by applicant): G protein signaling pathways in the striatum mediate a range of critical neuronal processes that control behavior, locomotion, pain perception and underlie drug abuse and addiction. The normal functioning of these pathways is hinged on the tight control of signal duration mediated by the Regulators of G protein signaling (RGS) proteins. Our long term goal is to elucidate the mechanisms governing the function of RGS proteins in neurons as a necessary prerequisite to understanding neurological disease processes and therapeutic means of their treatment. The main focus of the proposed studies is on RGS9-2, a striatum specific regulator that crucially controls signaling efficiency through dopamine and opioid receptor systems which are exploited by dugs of abuse. However, the molecular mechanisms of RGS9-2 function are largely unknown and need to be elucidated for better understanding of the neurochemical basis of addiction. To this end, we have recently discovered that RGS9-2 in the striatum exists in a complex with a novel neuronal protein which we named R7 Binding Protein (R7BP). Preliminary data suggest that R7BP serves as a critical modulator of RGS9-2 function in neurons. This HYPOTHESIS will be tested by addressing the following SPECIFIC AIMS: 1. To determine the mechanisms mediating stability and localization of RGS9-2. Proposed studies will determine the mechanisms by which R7BP circumvents the proteolysis of RGS9-2 and test the hypothesis that R7BP also determines the localization of RGS9-2 in neurons. 2. To understand the role of R7BP in the regulation of RGS9-2 catalytic activity. We will use a combination of in vitro enzymatic approaches and protein-protein interaction assays to determine the kinetic mechanism of R7BP action in the regulation of RGS9-2's ability to stimulate the GTPase activity of G proteins. 3. To further characterize the molecular composition of the G protein inactivating complex in striatal neurons. We will use proteomics approaches to identify additional components of the macromolecular ensemble involving the RGS9-2/R7BP complex. These studies should provide an understanding of the regulation of signaling in the striatum and generate insights into the molecular mechanisms of G protein signal disruption caused by drug abuse and neurological diseases.