PHOSPHORYLATION OF 4 AMINO-ACID-RESIDUES IN THE CARBOXYL-TERMINUS OF THE RAT SOMATOSTATIN RECEPTOR SUBTYPE-3 IS CRUCIAL FOR ITS DESENSITIZATION AND INTERNALIZATION

Agonist-dependent internalization of the rat somatostatin receptor sub type 3 (SSTR3) requires four hydroxyl amino acids (Ser(341), Ser(346), Ser(351), and Thr(357)) in the receptor C terminus (Roth, A., Kreienk amp, H.-J., Nehring, R., Roostermann, D., Meyerhof, W. and Richter, D. (1997) DNA Cell Biol. 16, 111-119). Here we report on the molecular m echanism responsible for the endocytotic process by analyzing the agon ist-dependent phosphorylation of wild-type and mutant receptors expres sed in human embryonic kidney cells, Wild-type SSTR3 is phosphorylated in response to agonist treatment, Phosphorylation is markedly reduced in a S341A/S346A/S351A triple mutant and is also reduced, but to a le sser extent, in the T357A point mutant. Internalization of the wild-ty pe receptor is preceded by a functional desensitization of the recepto r; in contrast, the triple serine mutant does not desensitize after tr eatment with agonists as assayed by its ability to inhibit forskolin-s timulated adenylate cyclase activity, After internalization via a clat hrin-coated vesicle mediated endocytotic pathway, SSTR3 efficiently re cycles to the cell surface, suggesting that agonist mediated endocytos is is necessary for the functional resensitization of a phosphorylated and desensitized receptor.