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Identification of phosphorylation sites in the COOH-terminal tail of the μ-opioid receptor

Ying-Ju Chen1, Sue Oldfield, Adrian J Butcher

  • 1School of Physiology and Pharmacology, University of Bristol, Bristol, UK.

Journal of Neurochemistry
|October 31, 2012
PubMed

Insights

Micro opioid receptor (MOPr) C-terminal tail phosphorylation sites were identified using mass spectrometry. Agonists stimulate additional phosphorylation, influencing arrestin binding and MOPr regulation by multiple kinases.

Area of Science:

  • Pharmacology
  • Molecular Biology
  • Cell Signaling

Background:

  • Phosphorylation is crucial for micro opioid receptor (MOPr) signaling and regulation.
  • Understanding MOPr phosphorylation provides insights into its cellular functions.

Purpose of the Study:

  • To identify basal and agonist-stimulated phosphorylation sites on the MOPr C-terminal tail.
  • To investigate the kinases responsible for MOPr phosphorylation.
  • To determine the effect of phosphorylation on arrestin binding.

Main Methods:

  • Mass spectrometry was used to analyze MOPr phosphorylation in HEK-293 cells.
  • Glutathione S transferase (GST) fusion proteins and point mutations of the MOPr C-terminal tail were employed.
  • In vitro kinase assays were performed with purified GRK2, PKC, and CaMKII.

Main Results:

  • Basal MOPr phosphorylation occurred at Ser(363) and Thr(370).
  • Morphine or DAMGO stimulation led to additional phosphorylation at Ser(356), Thr(357), and Ser(375).
  • GRK2, PKC, and CaMKII specifically phosphorylated Ser(375), Ser(363), and Thr(370), respectively.
  • MOPr C-terminal tail phosphorylation enhanced arrestin-2 and -3 binding.

Conclusions:

  • This study maps key phosphorylation sites on the MOPr C-terminal tail.
  • Multiple protein kinases regulate MOPr phosphorylation.
  • Phosphorylation modulates MOPr interaction with arrestins, impacting receptor signaling.

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