P2X7 Receptor Cell Surface Expression and Cytolytic Pore Formation Are Regulated by a Distal C-terminal Region

Abstract

The importance of the cytosolic C-terminal region of the P2X7 receptor (P2X7R) is unquestioned, yet little is known about the functional domains of this region and how they may contribute to the numerous properties ascribed to this receptor. A structure-function analysis of truncated and single-residue-mutated P2X7 receptors was performed in HEK-293 cells and Xenopus oocytes. Cells expressing receptors truncated at residue 581 (of 595) have negligible ethidium ion uptake, whereas those expressing the P2X7R truncated at position 582 give wild type ethidium ion uptake suggesting that pore formation requires over 95% of the C-terminal tail. Channel function was evident even in receptors that were truncated at position 380 indicating that only a small portion of the cytosolic region is required for channel activity. Surprisingly, truncations in the region between residues 551 and 581 resulted in non-functional receptors with no detectable cell surface expression in HEK-293 cells. A more detailed analysis revealed that mutations of single residues within this region could also abolish receptor function and cell surface expression, suggesting that this region may participate in regulating the surface expression of the pore-forming P2X7R.

Patch–clamp coordinated spectroscopy shows P2X 2 receptor permeability dynamics require cytosolic domain rearrangements but not Panx-1 channels

“…First, P2X 7 receptor mediated NMDG ϩ permeability increases and YOPRO1 uptake have been observed in oocytes that lack native Panx-1 (2,31,32). In contrast, a recent study concluded that these properties could only be measured when Panx-1 was coexpressed (18).…”

Conserved ectodomain cysteines are essential for rat P2X7 receptor trafficking

“…The conserved YXXXK motif found in Cterminal of P2XRs other than P2X7R is necessary for their expression at the cell surface [24]. In human P2X7R, a motif located in the distal region of its C-terminus tail is necessary for proper receptor trafficking; progressive deletions of this tail between residues 551 and 581 are nonfunctional and are not found in the plasma membrane [10,41]. The same motif is also involved in pore dilation [45,48].…”

“…The receptor exhibits low sensitivity to ATP, does not desensitize, and gradually develops permeability to organic cations, causing a sustained current growth accompanied by cell blebbing and death [1,12,32,45,50]. The C-terminus of P2X7R is longest among P2XRs and was suggested to account for many of these receptor-specific characteristics [2,41,45]. In contrast, the relevance of the ectodomain structure in P2X7R function has not been studied extensively.…”

The role of P2X7 receptors in tissue fibrosis: a brief review

“…Only one α-helix is predicted in the TM1 domain, and a major propensity for β-sheet conformation is expected in the TM2 region. The extracellular loop, with 10 conserved cysteine residues forming disulfide bridges and glycosylation sites represents the ATP binding site [22]. The stoichiometry of P2X 7 R involves a trimeric pore that consists of homomultimers [23].…”