Differential Regulation of Endosomal GPCR/β-Arrestin Complexes and Trafficking by MAPK

Background: Regulation between GPCRs and -arrestins in endosomes has never been reported. Results: A novel ERK regulatory site in -arrestin-2 controls the binding to GPCRs in endosomes, and receptor trafficking and signaling. Conclusion: Differential MAPK-dependent regulation of endosomal complexes exists among -arrestin subtypes and species. Significance: Such divergent mode of regulation may help understanding the physiological role of the endosomal GPCR/-arrestins signaling axis.-Arrestins are signaling adaptors that bind to agonist-occupied G protein-coupled receptors (GPCRs) and target them for endocytosis; however, the mechanisms regulating receptor/-arrestin complexes and trafficking in endosomes, remain ill defined. Here we show, in live cells, differential dynamic regulation of endosomal bradykinin B2 receptor (B2R) complexes with either -arrestin-1 or -2. We find a novel role for MAPK in the B2R/-arrestin-2 complex formation, receptor trafficking and signaling mediated by an ERK1/2 regulatory motif in the hinge domain of the rat -arrestin-2 ((PETP)-P-178), but not rat -arrestin-1 ((PERP)-P-177). While the ERK1/2 regulatory motif is conserved between rat and mouse -arrestin-2, it is surprisingly not conserved in human -arrestin-2 ((PEKP)-P-178). However, mutation of lysine 178 to threonine is sufficient to confer MAPK sensitivity to the human -arrestin-2. Furthermore, substitution for a phosphomimetic residue in both the rat and the human -arrestin-2 (T/K178D) significantly stabilizes B2R/-arrestin complexes in endosomes, delays receptor recycling to the plasma membrane and maintains intracellular MAPK signaling. Similarly, the endosomal trafficking of 2-adrenergic, angiotensin II type 1 and vasopressin V2 receptors was altered by the -arrestin-2 T178D mutant. Our findings unveil a novel subtype specific mode of MAPK-dependent regulation of -arrestins in intracellular trafficking and signaling of GPCRs, and suggest differential endosomal receptor/-arrestin-2 signaling roles among species.