μ-Opioid Receptor Desensitization

Abstract

It is generally accepted that the internalization and desensitization of -opioid receptor (MOR) involves receptor phosphorylation and ␤-arrestin recruitment. However, a mutant MOR, which is truncated after the amino acid residue Ser 363 (MOR363D), was found to undergo phosphorylation-independent internalization and desensitization. As expected, MOR363D, missing the putative agonist-induced phosphorylation sites, did not exhibit detectable agonist-induced phosphorylation. MOR363D underwent slower internalization as reflected in the attenuation of membrane translocation of ␤-arrestin 2 when compared with wild type MOR, but the level of receptor being internalized was similar to that of wild type MOR after 4 h of etorphine treatment. Furthermore, MOR363D was observed to desensitize faster than that of wild type MOR upon agonist activation. Surface biotinylation assay demonstrated that the wild type receptors recycled back to membrane after agonist-induced internalization, which contributed to the receptor resensitization and thus partially reversed the receptor desensitization. On the contrary, MOR363D did not recycle after internalization. Hence, MOR desensitization is controlled by the receptor internalization and the recycling of internalized receptor to cell surface in an active state. Taken together, our data indicated that receptor phosphorylation is not absolutely required in the internalization, but receptor phosphorylation and subsequent ␤-arrestin recruitment play important roles in the resensitization of internalized receptors.The regulation of G protein-coupled receptors (GPCRs) 1 signaling highlights the complex relationship between multiple mechanisms acting at different levels of signal propagation. Following the agonist activation of the receptor, G proteincoupled receptor kinases (GRKs) are recruited to the membrane and phosphorylate the agonist-activated GPCRs. GRKmediated receptor phosphorylation enhances the interaction between GPCRs and cytoplasmic proteins, arrestins, which uncouple the GPCRs from G proteins and also target the GPCRs to clathrin-coated pits for internalization (endocytosis). The internalized receptors can subsequently be dephosphorylated and recycled back to the plasma membrane, which contributes to resensitization. Alternatively, the internalized receptors can be targeted to lysosome for degradation (1-4). Receptor desensitization, the progressive loss of receptor function under continued exposure to an agonist, occurs in many GPCRs. Desensitization can be a consequence of multiple processes of receptor uncoupling, internalization, degradation, and recycling. Therefore, phosphorylation of agonist-activated receptors and subsequent arrestin recruitment are important processes in the modulation of GPCRs responsiveness.However, multiple studies challenge the critical role phosphorylation played in desensitization and internalization of many GPCRs. Desensitization and internalization of human parathyroid hormone receptor (5), AT 1A angiotensin receptor (6), and metabotropic gluta...

Morphine-induced μ-opioid receptor rapid desensitization is independent of receptor phosphorylation and β-arrestins

“…GRKs overexpression was the most commonly used method to study the effect of phosphorylation on receptor desensitization [33,40]. In order to overcome the difficulties in correlating the phosphorylation states of the receptor and desensitization, previous report has investigated the effect of receptor phosphorylation on desensitization by eliminating the putative phosphorylation sites in OPRM1 [41]. However, involvement of receptor phosphorylation in rapid desensitization was still unclear, since the relatively long agonist pretreated time and high agonist concentration resulted in OPRM1 desensitization rate reflecting both the uncoupling of OPRM1 from the G-protein and the internalization of OPRM1.…”

Membrane Glycoprotein M6a Interacts with the μ-Opioid Receptor and Facilitates Receptor Endocytosis and Recycling

“…However, experiments with C-terminal chimeras or truncations of the -and ␦-opioid receptors revealed that the C-tail cannot be the only region involved in receptor regulation (35)(36)(37)(38). It is further questionable whether a MOPr-derived endocytic recycling sequence (MRS) recently identified in the C terminus of -opioid receptors (39) is in fact essential for inducing receptor recycling because the C-terminal MOPr splice variants (MOPr-B and MOPr-D), which lack the MRS motif, undergo faster agonist-induced endocytosis and recycling than MOPr (40).…”

Chemical Genetic Engineering of G Protein-coupled Receptor Kinase 2

“…An important mechanism that regulates this process is termed receptor desensitization, which results in a reduction in the rate of G protein activation by the agonist-bound receptor complex (5,6). Desensitization is thought to be mediated through phosphorylation by G protein-coupled receptor kinases (GRKs) and second messenger-dependent protein kinases such as cAMP-dependent kinase (PKA), protein kinase C (PKC), and calmodulin-dependent kinase (7)(8)(9)(10)(11). Only GRKs selectively phosphorylate agonist-activated receptors, whereas second messenger-dependent kinases can phosphorylate receptors in the presence or absence of agonist.…”