Palmitoylation is required for signaling functions and membrane attachment of G(q)α and G(s)α

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(n) 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 (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.