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Tracking Drug-induced Changes in Receptor Post-internalization Trafficking by Colocalizational Analysis
Published on: July 3, 2015
Agonist-Dependent and -Independent κ Opioid Receptor Phosphorylation: Distinct Phosphorylation Patterns and Different
Yi-Ting Chiu1, Chongguang Chen1, Daohai Yu1
1Center for Substance Abuse Research and Department of Pharmacology (Y.-T.C., C.C., L.-Y.L.-C.) and Department of Clinical Sciences (D.Y.), Lewis Katz School of Medicine, Temple University, Philadelphia, Pennsylvania; and Institute of Pharmacology and Toxicology, Jena University Hospital, Friedrich-Schiller-University, Jena, Germany (S.S.).
Abstract:
We reported previously that the selective agonist U50,488H promoted phosphorylation of the mouse κ opioid receptor (KOPR) at residues S356, T357, T363, and S369. Here, we found that agonist (U50,488H)-dependent KOPR phosphorylation at all the residues was mediated by Gi/o proteins and multiple protein kinases [GRK2, GRK3, GRK5, GRK6 and protein kinase C (PKC)]. In addition, PKC activation by phorbol ester induced agonist-independent KOPR phosphorylation. Compared with U50,488H, PKC activation promoted much higher S356/T357 phosphorylation, much lower T363 phosphorylation, and similar levels of S369 phosphorylation. After U50,488H treatment, GRKs, but not PKC, were involved in agonist-induced KOPR internalization. In contrast, PKC activation caused a lower level of agonist-independent KOPR internalization, compared with U50,488H. U50,488H-induced activation of extracellular signal-regulated kinase 1/2 (ERK1/2) was G protein-, but not β-arrestin-, dependent. After U50,488H treatment, GRK-mediated, but not PKC-mediated, KOPR phosphorylation followed by β-arrestin recruitment desensitized U50,488H-induced ERK1/2 response. Therefore, agonist-dependent (GRK- and PKC-mediated) and agonist-independent (PKC-promoted) KOPR phosphorylations show distinct phosphorylation patterns, leading to diverse cellular outcomes.
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