Full-Length P2X7 Structures Reveal How Palmitoylation Prevents Channel Desensitization

Abstract

P2X receptors are trimeric, non-selective cation channels activated by extracellular ATP. The P2X 7 receptor subtype is a pharmacological target because of involvement in apoptotic, inflammatory, and tumor progression pathways. It is the most structurally and functionally distinct P2X subtype, containing a unique cytoplasmic domain critical for the receptor to initiate apoptosis and not undergo desensitization. However, lack of structural information about the cytoplasmic domain has hindered understanding of the molecular mechanisms underlying these processes. We report cryoelectron microscopy structures of full-length rat P2X 7 receptor in apo and ATP-bound states. These structures reveal how one cytoplasmic element, the C-cys anchor, prevents desensitization by anchoring the pore-lining helix to the membrane with palmitoyl groups. They show a second cytoplasmic element with a unique fold, the cytoplasmic ballast, which unexpectedly contains a zinc ion complex and a guanosine nucleotide binding site. Our structures provide first insights into the architecture and function of a P2X receptor cytoplasmic domain.

P2 Receptors as Therapeutic Targets in the Salivary Gland: From Physiology to Dysfunction

“…The P2X7R is a 595 amino acid protein that includes two transmembrane domains, intracellular carboxy and amino termini and a bulky hydrophilic extracellular loop with a cysteine rich region that forms disulfide bridges (McCarthy et al, 2019). It shares 40-50% amino acid homology with the other P2X receptors, but is structurally distinct in that its C-terminal tail extends for an additional 100-200 amino acids (North, 2002;Adinolfi et al, 2005;Sluyter, 2017).…”

Evolutionary Origin of the P2X7 C-ter Region: Capture of an Ancient Ballast Domain by a P2X4-Like Gene in Ancient Jawed Vertebrates

“…Protein structure homology-modeling was performed using the SWISS-MODEL program, accessible via the ExPASy web server (https://swissmodel.expasy.org/), using as template the structure of the rat P2X7 receptor obtained by single-particle cryoelectron microscopy [PDB ID: 6u9v (33)]. The relevant domains of the models were extracted and compared using Pymol (available at https://pymol.org/2/).…”

“…They bind to the same hydrophobic pocket away from the ATP binding site acting as allosteric non-competitive inhibitors (32). A major breakthrough was recently achieved by McCarthy et al (33) who published the first complete structure of the rat P2X7 receptor obtained by single-particle cryoelectron microscopy. The structure of the carboxy-terminal portion of the P2X7 receptor, which is unique to this P2X, defines a novel fold called ≪ ballast ≫ which contains a dinuclear Zn ion complex and a pocket containing a guanosine nucleotide.…”

“…In this work, we focused on the carboxy-terminal, intracytoplasmic sequence of P2X7 receptor and its conserved motifs including Zn-coordinating set of Cysteines and the GDP binding domain identified in McCarthy et al (33). We looked for proteins comprising related domains within vertebrates and beyond.…”

Shaping of Innate Immune Response by Fatty Acid Metabolite Palmitate

“…In immune cells, palmitoylation regulates the surface receptor trafficking and coordination involved in the relevant signaling pathways [44]. For example, cytosolic domain palmitoylation of P2X7, a purinergic receptor for extracellular ATP signal transduction, prevents receptor desensitization and reveals the unique properties of P2X7 signals [45]. P2X7 receptors are highly expressed membrane proteins in innate immune cells, particularly in mast cells, monocytes, macrophages, and microglia.…”