ATP stimulates GRK-3 phosphorylation and beta-arrestin-2-dependent internalization of P2X7 receptor
- PMID: 15728711
- PMCID: PMC2598767
- DOI: 10.1152/ajpcell.00315.2004
ATP stimulates GRK-3 phosphorylation and beta-arrestin-2-dependent internalization of P2X7 receptor
Abstract
The objective of this study was to understand the mechanisms involved in P2X(7) receptor activation. Treatments with ATP or with the P2X(7) receptor-specific ligand 2',3'-O-(4-benzoylbenzoyl)adenosine 5'-triphosphate (BzATP) induced pore formation, but the effect was slower in CaSki cells expressing endogenous P2X(7) receptor than in human embryonic kidney (HEK)-293 cells expressing exogenous P2X(7) receptor (HEK-293-hP2X(7)-R). In both types of cells Western blots revealed expression of three forms of the receptor: the functional 85-kDa form present mainly in the membrane and 65- and 18-kDa forms expressed in both the plasma membrane and the cytosol. Treatments with ATP transiently decreased the 85-kDa form and increased the 18-kDa form in the membrane, suggesting internalization, degradation, and recycling of the receptor. In CaSki cells ATP stimulated phosphorylation of the 85-kDa form on tyrosine and serine residues. Phosphorylation on threonine residues increased with added ATP, and it increased ATP requirements for phosphorylation on tyrosine and serine residues, suggesting a dominant-negative effect. In both CaSki and in HEK-293-hP2X(7)-R cells ATP also increased binding of the 85-kDa form to G protein-coupled receptor kinase (GRK)-3, beta-arrestin-2, and dynamin, and it stimulated beta-arrestin-2 redistribution into submembranous regions of the cell. These results suggest a novel mechanism for P2X(7) receptor action, whereby activation involves a GRK-3-, beta-arrestin-2-, and dynamin-dependent internalization of the receptor into clathrin domains, followed in part by receptor degradation as well as receptor recycling into the plasma membrane.
Figures
References
-
- Attramandal H, Arriza JL, Aoki C, Dawson TM, Codina J, Kwatra MM, Snyder SH, Caron MG, Lefkowitz RJ. β-Arrestin 2, a novel member of the arrestin/β-arrestin gene family. J Biol Chem. 1992;267:17882–17890. - PubMed
-
- Barak LS, Ferguson SS, Zhang J, Caron MG. A β-arrestin/green fluorescent protein biosensor for detecting G protein-coupled receptor activation. J Biol Chem. 1997;272:27497–27500. - PubMed
-
- Bardini M, Lee HY, Burnstock G. Distribution of P2X receptor subtypes in the rat female reproductive tract at late pro-oestrus/early oestrus. Cell Tissue Res. 2000;299:105–113. - PubMed
-
- Budagian V, Bulanova E, Brovko L, Orinska Z, Fayad R, Paus R, Bulfone-Paus S. Signaling through P2X7 receptor in human T cells involves p56lck, MAP kinases, and transcription factors AP-1 and NF-κB. J Biol Chem. 2003;278:1549–1560. - PubMed
Publication types
MeSH terms
Substances
Grants and funding
LinkOut - more resources
Full Text Sources
Other Literature Sources
Medical
Miscellaneous
