Signaling by a non-dissociated complex of G protein βγ and α subunits stimulated by a receptor-independent activator of G protein signaling, AGS8

Accumulating evidence suggests that heterotrimeric G protein activation may not require G protein subunit dissociation. Results presented here provide evidence for a subunit dissociation-independent mechanism for G protein activation by a receptor-independent activator of G protein signaling, AGS8. AGS8 is a member of the AGS group III family of AGS proteins thought to activate G protein signaling primarily through interactions with G beta gamma subunits. Results are presented demonstrating that AGS8 binds to the effector and alpha subunit binding "hot spot" on G beta gamma yet does not interfere with G alpha subunit binding to G beta gamma or phospholipase C beta 2 activation. AGS8 stimulates activation of phospholipase C beta 2 by heterotrimeric G alpha beta gamma and forms a quaternary complex with G alpha(i1), G beta(1)gamma(2), and phospholipase C beta 2. AGS8 rescued phospholipase C beta binding and regulation by an inactive beta subunit with a mutation in the hot spot (beta(1)(W99A)gamma(2)) that normally prevents binding and activation of phospholipase C beta 2. This demonstrates that, in the presence of AGS8, the hot spot is not used for G beta gamma interactions with phospholipase C beta 2. Mutation of an alternate binding site for phospholipase C beta 2 in the amino- terminal coiled- coil region of G beta gamma prevented AGS8- dependent phospholipase C binding and activation. These data implicate a mechanism for AGS8, and potentially other G beta gamma binding proteins, for directing G beta gamma signaling through alternative effector activation sites on G beta gamma in the absence of subunit dissociation.