DESENSITIZATION AND PHOSPHORYLATION OF THE GLUCAGON-LIKE PEPTIDE-1 (GLP-1) RECEPTOR BY GLP-1 AND 4-PHORBOL 12-MYRISTATE 13-ACETATE

Glucagon-like peptide-1 (GLP-1) stimulates glucose-induced insulin sec retion by binding to a specific G protein-coupled receptor linked to a ctivation of the adenylyl cyclase pathway. Here, using insulinoma cell lines, we studied homologous and heterologous desensitization of GLP- 1-induced cAMP production. Preexposure of the cells to GLP-1 induced a decrease in GLP-1-mediated cAMP production, as assessed by a 3- to 5- fold rightward shift of the dose-response curve and an approximately 2 0 percent decrease in the maximal production of cAMP. Activation of pr otein kinase C by the phorbol ester phorbol 12-myristate 13-acetate (P MA) also induced desensitization of the GLP-1-mediated response, leadi ng to a 6- to 9-fold shift in the EC(50) and a 30% decrease in the max imal production of cAMP. Both forms of desensitization were additive, and the protein kinase C inhibitor RO-318220 inhibited PMA-induced des ensitization, but not agonist-induced desensitization. GLP-1- and PMA- dependent desensitization correlated with receptor phosphorylation, an d the levels of phosphorylation induced by the two agents were additiv e. Furthermore, PMA-induced, but not GLP-1-induced, phosphorylation wa s totally inhibited by RO-318220. Internalization of the GLP-1 recepto r did not participate in the desensitization induced by PMA, as a muta nt GLP-1 receptor lacking the last 20 amino acids of the cytoplasmic t ail was found to be totally resistant to the internalization process, but was still desensitized after PMA preexposure. PMA and GLP-1 were n ot able to induce the phosphorylation of a receptor deletion mutant la cking the last 33 amino acids of the cytoplasmic tail, indicating that the phosphorylation sites were located within the deleted region. The cAMP production mediated by this deletion mutant was not desensitized by PMA and was only poorly desensitized by GLP-1. Together, our resul ts indicate that the production of cAMP and, hence, the stimulation of insulin secretion induced by GLP-1 can be negatively modulated by hom ologous and heterologous desensitization, mechanisms that involve rece ptor phosphorylation.