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Investigating Arrestin‐3 Recruitment to Nociceptin/Orphanin FQ Opioid Receptor (NOPR)
Author(s) -
Hansen Marissa M.,
Spangler Skylar M.,
Bruchas Michael R.
Publication year - 2018
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2018.32.1_supplement.689.5
Subject(s) - nociceptin receptor , arrestin , g protein coupled receptor , phosphorylation , internalization , chemistry , microbiology and biotechnology , opioid receptor , agonist , receptor , opioid , biology , signal transduction , pharmacology , opioid peptide , biochemistry
Nociceptin/orphanin FQ opioid receptor (NOPR) has therapeutic potential for treating drug addiction as demonstrated by the FDA‐approved drug Buprenorphine, a NOPR partial agonist, and recent studies show NOPR agonism blocks reward seeking behavior in rodents across multiple drug classes. Despite this, little is known about the signaling dynamics by which NOPR exerts these effects. It has been previously determined that the endogenous ligand nociceptin induces recruitment of arrestin‐3, a scaffolding protein which mediates GPCR desensitization and internalization, through NOPR c‐terminal tail phosphorylation. This study proposes that NOPR c‐terminal tail phosphorylation encodes arrestin functionality, as the specific phosphorylation patterns that recruit arrestin‐3 and 2 remain unknown. The biased signaling of NOPR in arrestin‐3 recruitment was explored using NOPR phospho‐mutants and biased ligands (reported in Chang et al., 2015) 3‐endo‐8‐[bis(2‐methylphenyl)methyl]‐3‐phenyl‐8‐azabicyclo[3.2.1]octan‐3‐ol (SCH 221,510) and 11‐[1‐(1‐methylcyclooctyl)‐4‐piperidinyl]‐2‐(3R)‐3‐piperidinyl‐1H‐benzimidazole trihydrochloride (MCOPPB) in order to dissect the molecular mechanisms of NOPR‐arrestin recruitment. Bioluminescence resonance energy transfer (BRET) assays were conducted to quantify real‐time binding between NOPR and arrestin to construct concentration response curves as a means of identifying signaling bias. This study shows that arrestin‐3 recruitment is not dependent on phosphorylation of c‐terminal tail sites as indicated by NOPR‐arrestin interaction with the NOPR c‐terminal phospho‐null mutant, however is absent in the 10A mutant which includes the 3rd intracellular loop. Further decoding of phosphorylation sites on the third intracellular loop could reveal the mechanism by which NOPR phosphorylation initiates downstream signaling via arrestin and thus the pathways by which the receptor controls neuronal function in dopamine neurons. Support or Funding Information This work was funded and supported by the National Institutes of Health Blueprint Program for Enhancing Neuroscience Diversity through Undergraduate Research Education Experiences (R25) and Washington University in St. Louis. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .