The Beta-2-adrenergic receptor/beta parrestin complex recruits the clathrin adaptor AP-2 during endocytosis

摘要

BETA-arrestins mediate the desensitization of the beta-2-adrcnergic receptor (beta-2AR) and many other G proteincoupled receptors (GPCRs). Additionally, parrestins initiate the endocytosis of these receptors via clathrin coated-pits and interact directly with clathrin. Consequently, it has been proposed that beta-parrestins serve as clathrin adaptors for the GPCR family by linking these receptors to clathrin lattices. AP-2, the heterotctrameric clathrin adaptor protein, has been demonstrated to mediate the internalization of many types of plasma membrane proteins other than GPCRs. AP-2 interacts with the clathrin heavy chain and cytoplasmic domains of receptors such as those for epidermal growth factor and transferrin. In the present study we demonstrate the formation of an agonist-induced multimeric complex containing a GPCR, parrestin 2, and the beta-2-adaptin subunit of AP-2. beta-2-Adaptin binds parrestin 2 in a yeast two-hybrid assay and coimmunoprecipitates with parrestins and beta-2AR in an agonist- dependent manner in HEK-293 cells. Moreover, beta-2-adaptin translocates from the cytosol to the plasma membrane in response to the beta-2AR agonist isoproterenol and colocalizes with beta-2AR in clathrin-coated Pits. Finally, expression of parrestin 2 minigene constructs containing the beta-2-adaptin interacting region inhibits beta-2AR endocytosis. These findings point to a role for AP-2 in GPCR endocytosis, and they suggest that AP-2 functions as a clathrin adaptor for the cndocytosis of diverse classes of membrane receptors.