Serine 363 is Required for NOPR Desensitization, Internalization, and Arrestin Signaling

Abstract

Nociceptin/Orphanin FQ peptide receptor (NOPR) plays a key role in pain modulation, opiate tolerance, and responsivity to stress and anxiety. C‐terminal phosphorylation at serine (S), threonine (T), and tyrosine (Y) residues are required for μ and κ opioid receptor internalization, desensitization, arrestin recruitment, and mitogen‐activated protein kinase phosphorylation (pMAPK), so we generated mutations in the NOPR c‐terminus. We showed that NOPR‐YFP internalized shortly after nociceptin treatment while NOPR‐S363A blocked internalization. We determined time course and concentration‐dependence of NOPR‐YFP‐mediated pMAPK. NOPR‐YFP‐induced pERK peaked at 10min following treatment and pJNK peaked at 30min. NOPR‐S363A showed reduced pJNK levels. We determined that NOPR‐YFP internalization is blocked by arrestin3 and GRK3 shRNAs and that NOPR‐S363A internalization and pJNK can be rescued via a dominant positive arrestin3. These data implicate GRK/arrestin in NOPR MAPK signaling and highlight the potential for the development of functionally selective NOPR‐ligands. This work was partially supported by NIH grant R00DA034929 (to MRB), T32DA007261 (to SDC) and HHMI SURF Program (to NZ).