Modulation of G(s) activity by phorbol myristate acetate in rat hepatocytes

Activation of protein kinase C promotes heterologous desensitization of hepatic adenylate cyclase. The basis for this desensitization was explored by use of a strategy with several independent approaches. Although not influencing the amount of forskolin-stimulated adenylate cyclase activity (catalyst), treatment with phorbol 12-myristate 13-acetate (PMA) decreased adenylate cyclase activation in response to either sodium fluoride or guanylyl imidodiphosphate [Gpp(NH)p]. Adenosine 3',5'-cyclic monophosphate (cAMP) accumulation in cholera toxin-treated hepatocytes and both the basal and GTP-stimulated adenylate cyclase activity of membranes from toxin-treated cells displayed a marked reduction in response to PMA. The ability of cholate extracts of hepatocyte membranes to reconstitute β-adrenergic-stimulated adenylate cyclase activity of membrane of S49 mouse lymphoma cyc^- cells was reduced by treatment with PMA. Cholera toxin-catalyzed labeling of G(s)α-subunits was likewise diminished by phorbol ester treatment. Immunoblots of membranes from control or PMA-treated hepatocytes showed no difference in the amount of G(s)α. Immunoprecipitation studies failed to detect phosphorylation of this G protein α-subunit. The data demonstrate that PMA induces an alteration in the functional status of G(s) without altering the amount of this transmembrane signaling element. The alteration in G(s) function may play a significant role in heterologous desensitization.