A cluster of Ser/Thr residues at the C-terminus of mu-opioid receptor is required for G protein-coupled receptor kinase 2-mediated desensitization

To investigate the functional role of G protein-coupled receptor kinases (GRK) in homologous desensitization of the mu-opioid receptor, human embryonic kidney (HEK) 293 cells, which express a significant level of GRK2, were stably transfected with the cDNA encoding the rat mu-opioid receptor. Wild-type mu-opioid receptors developed homologous desensitization after 30 min pretreatment with DAMGO ([D-Ala(2),N-methyl-Phe(4),Gly-ol(5)]enkephalin), a specific mu-opioid receptor agonist. The ability of mu-opioid receptors to develop homologous desensitization was greatly impaired following the transfection of a cDNA fragment encoding the GRK2(495-689) polypeptide, which is believed to block G(beta gamma)-mediated transduction events including the membrane translocation and activation of GRK2. The mu(C Delta 45) receptor, a deletion mutant that lacks 45 C-terminal amino acids, failed to exhibit homologous desensitization after 30 min pretreatment of DAMGO. The mu(C Delta 41) receptor, which differs from the mu(C Delta 45) receptor by having four more Ser/Thr residues (Thr(354)Ser(355)Ser(356)Th(357)), developed GRK2-mediated desensitization. These results suggest that homologous desensitization of rat mu-opioid receptors results from the activation of GRK2 and that a cluster of Ser/Thr residues (Thr(354)Ser(355)Ser(356)Thr(357)) at the intracellular carboxyl tail plays an important role in GRK2-mediated mu-opioid receptor desensitization. (C) 2000 Elsevier Science Ltd. All rights reserved.