Receptor-independent activators of heterotrimeric G-protein signaling pathways

Heterotrimeric G-protein signaling systems are activated via cell surface receptors possessing the seven-membrane span motif, Several observations suggest the existence of other modes of stimulus input to heterotrimeric G-proteins, As part of an overall effort to identify such proteins we developed a functional screen based upon the pheromone response pathway in Saccharomyces cerevisiae, We identified two mammalian proteins, AGS2 and AGS3 (activators of G-protein Signaling), that activated the pheromone response pathway at the level of heterotrimeric G-proteins in the absence of a typical receptor. beta-galactosidase reporter assays in yeast strains expressing different G alpha subunits (Gpa1, G(s)alpha, G(i)alpha(2(Gpal(1-41))), G(i)alpha(3(Gpa1(1-41))), G alpha(16(Gpa1(1-41)))) indicated that AGS proteins selectively activated G-protein heterotrimers. AGS3 was only active in the G(i)alpha(2) and G(i)alpha(3) genetic backgrounds, whereas AGS2 was active in each of the genetic backgrounds except Gpa1, In protein interaction studies, AGS2 selectively associated with G beta gamma, whereas AGS3 bound G alpha and exhibited a preference for G alpha GDP versus G alpha GTP gamma S, Subsequent studies indicated that the mechanisms of G-protein activation by AGS2 and AGS3 were distinct from that of a typical G-protein-coupled receptor. AGS proteins provide unexpected mechanisms for input to heterotrimeric G-protein signaling pathways. AGS2 and AGS3 may also serve as novel binding partners for G alpha and G beta gamma that allow the subunits to subserve functions that do not require initial heterotrimer formation.