Binding of G beta gamma subunits to cRaf1 downregulates G-protein-coupled receptor signalling

Receptors of the seven transmembrane domain family are coupled to heterotri meric G proteins [1]. Binding of ligand to these receptors induces dissocia tion of the heterotrimeric complex into free GTP-G alpha and G beta gamma s ubunits, which then interact with their respective effector molecules to st imulate specific cellular responses. In some cases, these cellular response s involve mitogenic signalling [2]. The mitogen-activated protein (MAP) kin ase cascade is initiated by the protein kinase cRaf1 and links growth facto r receptor signalling to cell growth and differentiation [3]. The main acti vator of cRaf1 is the small GTP-binding protein Pas [4], and the binding of cRaf1 to GTP-Ras translocates cRaf1 to the plasma membrane, where it is ac tivated [5]. It has been reported that cRaf1 associates directly with the b eta subunit of heterotrimeric G proteins in vitro, and with the beta gamma subunit complex in vivo [6], but the role of this association is not yet un derstood. Here, we show that cRaf1 associates with G beta 1 gamma 2, and th at this association in mammalian cells is significantly enhanced when activ e p21(Ras) is present or when cRaf1 is otherwise targeted to the membrane, Association with G beta 1 gamma 2 has no effect on the kinase activity of c Raf1, but cRaf1 can affect G beta gamma-mediated signalling events. Thus, m embrane localised cRaf1 inhibits G protein-coupled receptor (GPCR)-stimutat ed activation of phospholipase C beta (PLC beta) by sequestration of G beta gamma subunits, an effect also observed with endogenous levels of cRaf1. O ur data suggest that cRaf1 may be an important regulator of signalling by G beta gamma, particularly in those GPCR systems that stimulate the MAP kina se cascade through the activation of p21(Ras).