Proteomic and functional evidence for a P2X(7) receptor signalling complex

P2X receptors are ATP-gated ion channels in the plasma membrane, but activa tion of the P2X(7) receptor also leads to rapid cytoskeletal re-arrangement s such as membrane blebbing. We identified 11 proteins in human embryonic k idney cells that interact with the rat P2X(7) receptor, by affinity purific ation followed by mass spectroscopy and immunoblotting [laminin alpha3, int egrin beta2, beta -actin, alpha -actinin, supervillin, MAGuK, three heat sh ock proteins, phosphatidylinositol 4-kinase and receptor protein tyrosine p hosphatase-beta (RPTP beta)]. Activation of the P2X(7) receptor resulted in its dephosphorylation. Whole-cell recordings from cells expressing P2X(7) receptors showed that this markedly reduced subsequent ionic currents and i t also slowed membrane bleb formation. By mutagenesis, we identified Tyr(34 3) in the putative second transmembrane domain as the site of phosphorylati on. Thus, we have identified a P2X(7) receptor signalling complex, some mem bers of which may initiate cytoskeletal rearrangements following receptor a ctivation. Others, such as RPTP beta, might exert feedback control of the c hannel itself through its dephosphorylation.