Receptor-independent activators of heterotrimeric G-protein signaling pathways

Heterotrimeric G-protein signaling systems are activated via cell surface r eceptors possessing the seven-membrane span motif, Several observations sug gest the existence of other modes of stimulus input to heterotrimeric G-pro teins, As part of an overall effort to identify such proteins we developed a functional screen based upon the pheromone response pathway in Saccharomy ces cerevisiae, We identified two mammalian proteins, AGS2 and AGS3 (activa tors of G-protein Signaling), that activated the pheromone response pathway at the level of heterotrimeric G-proteins in the absence of a typical rece ptor. beta-galactosidase reporter assays in yeast strains expressing differ ent 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 select ively activated G-protein heterotrimers. AGS3 was only active in the G(i)al pha(2) and G(i)alpha(3) genetic backgrounds, whereas AGS2 was active in eac h of the genetic backgrounds except Gpa1, In protein interaction studies, A GS2 selectively associated with G beta gamma, whereas AGS3 bound G alpha an d exhibited a preference for G alpha GDP versus G alpha GTP gamma S, Subseq uent studies indicated that the mechanisms of G-protein activation by AGS2 and AGS3 were distinct from that of a typical G-protein-coupled receptor. A GS proteins provide unexpected mechanisms for input to heterotrimeric G-pro tein signaling pathways. AGS2 and AGS3 may also serve as novel binding part ners for G alpha and G beta gamma that allow the subunits to subserve funct ions that do not require initial heterotrimer formation.