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. ? ? ?
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