Phosphorylation of EEA1 by p38 MAP kinase regulates μ opioid receptor endocytosis

Morphine analgesic properties and side effects such as tolerance are mediated by the l opioid receptor (MOR) whose endocytosis is considered of primary importance for opioid pharmacological effects. Here, we show that p38 mitogen-activated protein kinase ( MAPK) activation is required for MOR endocytosis and sufficient to trigger its constitutive internalization in the absence of agonist. Further studies established a functional link between p38 MAPK and the small GTPase Rab5, a key regulator of endocytosis. Expression of an activated mutant of Rab5 stimulated endocytosis of MOR ligand-independently in wild-type but not in p38 alpha(-/-) cells. We found that p38 alpha can phosphorylate the Rab5 effectors EEA1 and Rabenosyn-5 on Thr-1392 and Ser-215, respectively, and these phosphorylation events regulate the recruitment of EEA1 and Rabenosyn-5 to membranes. Moreover, phosphomimetic mutation of Thr-1392 in EEA1 can bypass the requirement for p38a in MOR endocytosis. Our results highlight a novel mechanism whereby p38 MAPK regulates receptor endocytosis under physiological conditions via phosphorylation of Rab5 effectors.