Sequestration and phosphorylation of the prostaglandin E2EP4 receptor: dependence on the C-terminal tail

Citation
D. Slipetz et al., Sequestration and phosphorylation of the prostaglandin E2EP4 receptor: dependence on the C-terminal tail, BIOCH PHARM, 62(8), 2001, pp. 997-1012
Citations number
44
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Pharmacology & Toxicology
Journal title
BIOCHEMICAL PHARMACOLOGY
ISSN journal
0006-2952 → ACNP
Volume
62
Issue
8
Year of publication
2001
Pages
997 - 1012
Database
ISI
SICI code
0006-2952(20011015)62:8<997:SAPOTP>2.0.ZU;2-X
Abstract
The prostaglandin E-2 (PGE(2)) EP4 subtype is one of four prostanoid recept ors that use PGE(2) as the preferred ligand. We have investigated the agoni st-mediated regulation of EP4 using a multifaceted approach. Short-term (30 min) agonist challenge of recombinant EP4 expressed in human embryonic kid ney 293 cells (EP4-HEK293 cells) with PGE(2) (1 muM) resulted in the desens itization of intracellular cyclic AMP (cAMP) accumulation and a reduction i n cell surface [H-3]PGE(2) specific binding sites. These events correlated with sequestration of EP4, as visualized by immunofluorescence confocal mic roscopy and phosphorylation, as shown by [P-32]orthophosphate labeling of t he receptor. Stimulation of protein kinase A activity in EP4-HEK293 cells ( 10 muM forskolin or 1 mM 8-bromo-cAMP) did not induce EP4 desensitization, sequestration, or phosphorylation. In contrast, stimulation of protein kina se C activity (100 nM phorbol 12-myristate 13-acetate) attenuated PGE(2)-in duced adenylyl cyclase activity and increased EP4 phosphorylation, but did not induce sequestration or a reduction in [H-3]PGE(2) specific binding sit es. EP4 receptors containing a third intracellular loop deletion [EP4 (del. 215-263)] or a carboxyl-terminal tail truncation [EP4 (del. 355)] of EP4 w ere used to demonstrate that the C-terminal tail governs sequestration as w ell as phosphorylation of the receptor. (C) 2001 Elsevier Science Inc. All rights reserved.