The Ras-dependent activation of mitogen-activated protein (MAP) kinase pathways by many receptors coupled to heterotrimeric guanine nucleotide binding proteins (G proteins) requires the activation of Src family tyrosine kinases. Stimulation of beta2 adrenergic receptors resulted in the assembly of a protein complex containing activated c-Src and the receptor. Src recruitment was mediated by beta-arrestin, which functions as an adapter protein, binding both c-Src and the agonist-occupied receptor. beta-Arrestin 1 mutants, impaired either in c-Src binding or in the ability to target receptors to clathrin-coated pits, acted as dominant negative inhibitors of beta2 adrenergic receptor-mediated activation of the MAP kinases Erk1 and Erk2. These data suggest that beta-arrestin binding, which terminates receptor-G protein coupling, also initiates a second wave of signal transduction in which the "desensitized" receptor functions as a critical structural component of a mitogenic signaling complex.

beta-Arrestins are proteins that bind phosphorylated heterotrimeric GTP-binding protein (G protein)-coupled receptors (GPCRs) and contribute to the desensitization of GPCRs by uncoupling the signal transduction process. Resensitization of GPCR responsiveness involves agonist-mediated receptor sequestration. Overexpression of beta-arrestins in human embryonic kidney cells rescued the sequestration of beta 2-adrenergic receptor (beta 2AR) mutants defective in their ability to sequester, an effect enhanced by simultaneous overexpression of beta-adrenergic receptor kinase 1. Wild-type beta 2AR sequestration was inhibited by the overexpression of two beta-arrestin mutants. These findings suggest that beta-arrestins play an integral role in GPCR internalization and thus serve a dual role in the regulation of GPCR function.

The G protein-coupled -opioid receptor (OR) mediates the physiological effects of endogenous opioid peptides as well as the structurally distinct opioid alkaloids morphine and etorphine. An intriguing feature of OR signaling is the differential receptor trafficking and desensitization properties following activation by distinct agonists, which have been proposed as possible mechanisms related to opioid tolerance. Here we report that the ability of distinct opioid agonists to differentially regulate OR internalization and desensitization is related to their ability to promote G protein-coupled receptor kinase (GRK)-dependent phosphorylation of the OR. Although both etorphine and morphine effectively activate the OR, only etorphine elicits robust OR phosphorylation followed by plasma membrane translocation of ␤-arrestin and dynamin-dependent receptor internalization. In contrast, corresponding to its inability to cause OR internalization, morphine is unable to either elicit OR phosphorylation or stimulate ␤-arrestin translocation. However, upon the overexpression of GRK2, morphine gains the capacity to induce OR phosphorylation, accompanied by the rescue of ␤-arrestin translocation and receptor sequestration. Moreover, overexpression of GRK2 also leads to an attenuation of morphine-mediated inhibition of adenylyl cyclase. These findings point to the existence of marked differences in the ability of different opioid agonists to promote OR phosphorylation by GRK. These differences may provide the molecular basis underlying the different analgesic properties of opioid agonists and contribute to the distinct ability of various opioids to induce drug tolerance.