We have discovered a protein family termed RGSs that impair signal transduction through pathways that use seven trans-membrane receptors and heterotrimeric G proteins. Such receptors, when activated following the binding of a ligand such as a hormone or chemokine, trigger the G alpha subunit to exchange GTP for GDP; this causes the dissociation of G alpha and G beta-gamma subunits and downstream signaling. RGS proteins bind G alpha subunits and function as GTPase activating proteins (GAPs), thereby deactivating the G alpha subunit and facilitating their re-association with G beta-gamma. We have shown that RGS proteins modulate signaling through chemokine receptors and that they can inhibit chemotaxis. RGS1 expressing B lymphocytes fail to migrate in response to the chemokine SDF-1. Conversely, RGS1 deficient B cells obtained from mice in which the RGS1 gene has been disrupted by gene targeting have an enhanced chemotaxic response to SDF-1. In addition these mice have impaired mucosal immune respones. We have also shown that certain RGS proteins can directly inhibit the activation of adenylyl cyclase, thereby providing a mechanism by which these proteins can inhibit Gs induced cAMP production. These findings are relevant to the olfactory system. Odorants activate the Gs family member Golf, which leads to activation of adenylyl cyclase type III (AC III) and the production of cAMP. RGS2 potently inhibits AC III mediated cAMP production. Olfactory neurons express both RGS2 and RGS3 and the microinjection of an antibody to RGS2 into olfactory neurons profoundly enhances odorant induced signal transduction. This suggests that the level of RGS2 in olfacotory neurons profoundly affects their response to odorants. Another unusal property of RGS proteins is the ability of RGS3 to profoundly inhibit the activation of pathways triggered by Gbeta gamma subunits. This is independent of RGS3's GAP activity and likely depends upon the binding of RGS3 to free beta gamma subunits. We have shown that RGS14 is highly expressed in lymphocytes and its expression levels are modulated by signaling through the B-cell and T-cell antigen receptors. RGS14 also inhibits G13alpha signaling through a yet unknown mechanism. We have also identified a panel of proteins with which RGS14 interacts. Their roles in RGS14 function is being assessed. RGS14 has been transgenically introduced into the mouse genome to enhance its level of expression. Homozygous mice carrying the transgene have been identified. A detailed analysis of these mice is in progress. The murine RGS13, RGS3, and RGS5 genes have been isolated and gene targeting of the mouse RGS3 and RGS14 loci is in progress. An analysis of the RGS3 gene locus has identified a novel splice variant of RGS3 with an altered N-terminus. An antibody which specially recognizes this variant has been produced. We have carried out a series of biochemical studies of RGS5, which indicate that RGS5 is a potent Gqalpha GAP and functions in the regulation of cardiovascular function. Specific antibodies to RGS1, RGS2, RGS3, RGS5, RGS8, RGS10, RGS12, RGS13, RGS14, and RGS15 have been produced. Using these antibodies and in situ hybridization for specific RGS mRNAs, we are examining the developmental exrpession of the RGS family. To examine the consequences of persistent heterotrimeric G-protein signaling in lymphocyte development and function, transgenic mice expressing GTPase deficient G12 have been established. Suggesting a role for G12 in B cell development these mice have a partial block in the B-lymphocyte developmental pathway.
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