Understanding mechanisms of regulation of G protein-mediated transmembrane signaling is critical to comprehension and prediction of the effects of hormones, neurotransmitters, and related drugs on cellular function. A large a novel family of negative regulators of G protein function has been discovered recently [1] termed RGS proteins (Regulators of G protein Signaling), these proteins act as powerful GTPase-activating proteins (GAPs) for several heterodimeric G proteins, increasing their rate of GTPase hydrolysis more than 40-fold [2]. RGS proteins act catalytically and are thus likely to play a significant role in attenuation or down regulation of receptor-effector systems where G proteins act as intermediary transducers. I propose to examine the role and regulation of RGS proteins in vivo using a two-tiered approach. The first tier employs RNase protection assays to comprehensively catalog the identity and conditions required for RGS expression. The information obtained from this approach will subsequently be used for the second tier of study; namely, to examine the regulation of RGS protein activity in vivo. RGS proteins identified in the first tier will be tested for expression levels, sub- cellular localization, and post-translational status (e.g. phosphorylation, lipidation and association with regulatory proteins) and correlated with in vitro GAP assay activity. and in vivo effects on G/s, G/i, and G/q coupled receptor systems. RGS expression and activity will be compared in the presence and absence of appropriate hormones and neurotransmitters..
