Homologous desensitization of adenylate cyclase is associated with phosphorylation of the beta-adrenergic receptor.

We recently demonstrated that heterologous desensitization of adenylate cyclase in turkey erythrocytes is highly correlated with phosphorylation of the beta-adrenergic receptor. In contrast, little is known of the biochemical mechanisms underlying the homologous form of beta-adrenergic receptor desensitization, which is agonist-specific and not cAMP-mediated. Accordingly, the present studies were undertaken to examine if phosphorylation of the beta-adrenergic receptor is also associated with this form of desensitization in a well studied model system, the frog erythrocyte. Preincubation of these cells with the beta-adrenergic agonist isoproterenol leads to a 45% decline in isoproterenol-stimulated adenylate cyclase activity without significant changes in basal, prostaglandin E1-, NaF-, guanyl-5'-yl-imidodiphosphate-, forskolin-, or MnCl2-stimulated enzyme activities. There is also a 48% decline in 125Iiodocyanopindolol membrane binding sites. Conversely, preincubation of the cells with prostaglandin E1 attenuates only the prostaglandin E1-stimulated enzyme activity and does not affect 125Iiodocyanopindolol binding. Phosphorylation of the beta-adrenergic receptor was assessed by preincubating the cells with 32Pi and desensitizing them, and subsequently purifying the receptors by affinity chromatography. Under basal conditions there is about 0.62 mol of phosphate/mol of receptor whereas after desensitization with isoproterenol this increases to 1.9 mol/mol. This isoproterenol-induced receptor phosphorylation exhibits stereospecificity and is blocked by the beta-adrenergic antagonist propranolol. In addition, preincubation with prostaglandin E1 does not promote beta-adrenergic receptor phosphorylation. These data suggest that receptor phosphorylation is involved in homologous as well as heterologous forms of desensitization and may provide a unifying mechanism for desensitization of adenylate cyclase-coupled hormone receptors.