P2X receptors are ATP-gated ion channels in the plasma membrane, but activation of the P2X 7 receptor also leads to rapid cytoskeletal re-arrangements such as membrane blebbing. We identified 11 proteins in human embryonic kidney cells that interact with the rat P2X 7 receptor, by affinity purification followed by mass spectroscopy and immunoblotting [laminin α3, integrin β2, β-actin, α-actinin, supervillin, MAGuK, three heat shock proteins, phosphatidylinositol 4-kinase and receptor protein tyrosine phosphatase-β (RPTPβ)]. 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 it also slowed membrane bleb formation. By mutagenesis, we identified Tyr 343 in the putative second transmembrane domain as the site of phosphorylation. Thus, we have identified a P2X 7 receptor signalling complex, some members of which may initiate cytoskeletal rearrangements following receptor activation. Others, such as RPTPβ, might exert feedback control of the channel itself through its dephosphorylation.