Palmitoylation is required for signaling functions and membrane attachment of G_qα and G_sα

We have identified the palmitoylated cysteine residues of α_q and α_s, α subunits of two heterotrimeric G proteins. Mutational substitutions of serines for cysteines 9 and 10 in α_q and cysteine 3 in α_s profoundly alter behavior of the subunits expressed in HEK293 cells. Neither mutant α subunit incorporates palmitate; both mutant proteins are found in the soluble rather than the particulate fraction; mutant α_q or α_s cannot couple a co-expressed receptor to stimulation of phospholipase C or adenylylcyclase, respectively; cysteine substitution prevents a mutationally activated α_q (R183C) from stimulating phospholipase C directly, and reduces but does not abolish the ability of a similarly activated α_s (R201C) to stimulate cAMP synthesis. Substitution of a myristoylation sequence for the palmitoylation sites leads to labeling of α_q and α_s by myristate, rather than by palmitate. Myristoylation restores the abilities of both nonpalmitoylated α_q and α_s to attach to membranes and, in the case of α_q, restores its ability to stimulate phospholipase C, whether triggered by the R183C mutation or by receptor activation. These findings identify palmitoylation as a critical determinant of membrane attachment for α_q and α_s and show that this modification is required for normal signaling by these proteins.