Identification of phosphorylation sites in the COOH-terminal tail of the µ-opioid receptor.
J Neurochem
; 124(2): 189-99, 2013 Jan.
Article
em En
| MEDLINE
| ID: mdl-23106126
ABSTRACT
Phosphorylation is considered a key event in the signalling and regulation of the µ opioid receptor (MOPr). Here, we used mass spectroscopy to determine the phosphorylation status of the C-terminal tail of the rat MOPr expressed in human embryonic kidney 293 (HEK-293) cells. Under basal conditions, MOPr is phosphorylated on Ser(363) and Thr(370), while in the presence of morphine or [D-Ala2, NMe-Phe4, Gly-ol5]-enkephalin (DAMGO), the COOH terminus is phosphorylated at three additional residues, Ser(356) , Thr(357) and Ser(375). Using N-terminal glutathione S transferase (GST) fusion proteins of the cytoplasmic, C-terminal tail of MOPr and point mutations of the same, we show that, in vitro, purified G protein-coupled receptor kinase 2 (GRK2) phosphorylates Ser(375), protein kinase C (PKC) phosphorylates Ser(363), while CaMKII phosphorylates Thr(370). Phosphorylation of the GST fusion protein of the C-terminal tail of MOPr enhanced its ability to bind arrestin-2 and -3. Hence, our study identifies both the basal and agonist-stimulated phospho-acceptor sites in the C-terminal tail of MOPr, and suggests that the receptor is subject to phosphorylation and hence regulation by multiple protein kinases.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Receptores Opioides mu
Tipo de estudo:
Diagnostic_studies
Limite:
Animals
/
Humans
Idioma:
En
Ano de publicação:
2013
Tipo de documento:
Article