Chapter Eight - Regulation of P2X Purinergic Receptor Signaling by Cholesterol
Introduction
P2X receptors have a very widespread distribution throughout the human body and purinergic signaling mediated by these receptors takes place in all tissues (Khakh & North, 2006). All cells release ATP by a combination of mechanisms including vesicular release, ion channel-mediated release, and cell lysis (Burnstock & Verkhratsky, 2012). Under normal physiological conditions, the levels of extracellular ATP are very low because of the highly active, membrane-bound ectonucleotidases that rapidly break down ATP to ADP, AMP, and adenosine. However, levels of extracellular ATP are much higher following injury or in diseases that are associated with inflammation. These include cancer, neurodegenerative, and metabolic diseases, in addition to the well-characterized chronic inflammatory diseases such as arthritis and atherosclerosis. High ATP concentrations are recognized as a danger signal, the effects of which are mediated by the uniquely low-affinity P2X7 receptor (Khakh & North, 2006). Conditions that lead to enhanced P2X7 receptor signaling are also conditions that result in perturbations of membrane lipids and remodeling of rafts (Fessler & Parks, 2011).
Lipid rafts are specialized, liquid-ordered microdomains in the plasma membrane enriched in cholesterol, sphingolipids, and saturated phospholipids. They can be identified biochemically by their low buoyant density on a sucrose density gradient, their resistance to solubilization in detergent, and the presence of protein markers such as caveolin (Lingwood and Simons, 2010, Pike, 2004). The precise nature of rafts can differ with respect to their size and lipid/protein composition. For example, caveolae are a special type of lipid raft that form small invaginations of the plasma membrane and contain caveolin, an integral membrane protein. Some other proteins, such as glycosylphosphatidylinositol-anchored proteins, are targeted to rafts, but others are excluded, resulting in compartmentalization and organization of signaling events. Dynamic remodeling of rafts is important in signaling and the control of cholesterol trafficking through rafts by environmental factors and signaling cascades can be an important determinant of the cellular response (Fessler & Parks, 2011).
Section snippets
P2X Receptors Form a Structurally Distinct Class of Ligand-Gated Ion Channels
P2X receptors assemble as homo- or heterotrimeric complexes. The seven identified subtypes share the same general structure, possessing two transmembrane regions connected via a large extracellular loop and intracellular N- and C-termini. The seven P2X subunits are 40%–50% identical in their amino acid sequence, but they range in size because of a very divergent C-terminal domain which is short in P2X1, P2X3, P2X4, and P2X6 (~ 30–50 residues) longer in P2X2 and P2X5 and considerably longer in
P2X Receptors Are Localized to Lipid Rafts
Several members of the P2X family are detected in the buoyant lipid raft fractions including P2X1, P2X2, P2X3, P2X4, and P2X7 (Allsopp et al., 2010, Barth et al., 2008, Barth et al., 2007, Gnanasekaran et al., 2011, Gonnord et al., 2009, Vacca et al., 2004, Vial and Evans, 2005, Vial et al., 2006). The other two subtypes, P2X5 and P2X6, have very limited or no function as homomeric receptors, and their association with rafts has not been reported. The presence of P2X receptors within lipid
P2X1 Receptors
The P2X1 receptor is expressed in smooth muscle where it mediates contraction of arteries, resistant arterioles, vas deferens, and bladder (Vial & Evans, 2000). Similar to the recombinant receptor expressed in HEK293 cells, the native receptor in smooth muscle is predominantly within lipid rafts (Vial and Evans, 2005, Vial et al., 2006). P2X1 receptor currents are profoundly sensitive to disruption of lipid rafts, with the current mediated by the recombinant receptor expressed in HEK293 cells
P2X7 Receptors and the Mechanisms by Which They Associate With Rafts
P2X7 receptors are highly expressed in immune cells, including mast cells, monocytes, macrophages, microglia, and T-lymphocytes, but are also found in other cells including epithelia, vascular endothelial cells, fibroblasts, osteoclasts, and osteoblasts (Surprenant & North, 2009). The nature of lipid rafts differs in these cells, with caveolae being well described in fibroblasts and epithelial cells, but less well characterized in macrophages (Fessler & Parks, 2011). Thus the regulation of P2X7
The Effects of Cholesterol on P2X7 Receptor Function and Signaling
There are several ways in which the recruitment of the P2X7 receptor to lipid rafts and the cholesterol content of the rafts could affect receptor signaling: first, by affecting the plasma membrane expression of the receptor, second, by affecting receptor function such as the gating of the channel pore, and third, by concentrating signaling molecules that act either as downstream effectors of the receptor or as regulators of the receptor.
Little is known about what controls the trafficking of
The Determinants of P2X7 Receptor Cholesterol Sensitivity
It is still unclear whether cholesterol exerts its effect on P2X receptor activity directly, by binding to a cholesterol-sensing motif (CSM) within the receptor, or indirectly, via an associated protein and/or by changing the biophysical properties of the membrane. Unlike the cholesterol regulation of P2X1 receptors, the potentiation of P2X7 receptor activity that occurs in response to cholesterol depletion is not affected by the cytoskeletal-stabilizing agent jasplakinolide, which argues
Future Work
In this review, we have focused on the inhibitory regulation of P2X7 receptors by cholesterol but have also briefly touched on other lipids, such as the products of the PLA2 pathway, which positively regulate the receptor. The underlying molecular mechanisms of this regulation are not yet established, but with the publishing of P2X receptor structures in the close, open, and desensitized states, the field is well placed to elucidate the detailed nature of how lipids directly or indirectly
Acknowledgments
I thank Dr. Andrew Thompson for assistance with Fig. 1 and for discussions. This work have been supported by the BM1406 COST Action.
References (58)
- et al.
Lipid raft association and cholesterol sensitivity of P2X1-4 receptors for ATP: Chimeras and point mutants identify intracellular amino-terminal residues involved in lipid regulation of P2X1 receptors
The Journal of Biological Chemistry
(2010) - et al.
Characterization of the molecular interaction between caveolin-1 and the P2X receptors 4 and 7 in E10 mouse lung alveolar epithelial cells
The International Journal of Biochemistry & Cell Biology
(2008) - et al.
Analysis of assembly and trafficking of native P2X4 and P2X7 receptor complexes in rodent immune cells
The Journal of Biological Chemistry
(2009) - et al.
The mechanism of docosahexaenoic acid-induced phospholipase D activation in human lymphocytes involves exclusion of the enzyme from lipid rafts
The Journal of Biological Chemistry
(2002) - et al.
Coupling of two pools of P2X7 receptors to distinct intracellular signaling pathways in rat submandibular gland
Journal of Lipid Research
(2006) - et al.
Interaction of membrane/lipid rafts with the cytoskeleton: Impact on signaling and function: Membrane/lipid rafts, mediators of cytoskeletal arrangement and cell signaling
Biochimica et Biophysica Acta
(2014) - et al.
Identification of human P2X1 receptor-interacting proteins reveals a role of the cytoskeleton in receptor regulation
The Journal of Biological Chemistry
(2011) - et al.
Ceramide selectively displaces cholesterol from ordered lipid domains (rafts): Implications for lipid raft structure and function
The Journal of Biological Chemistry
(2004) - et al.
Plasma membrane cholesterol as a regulator of human and rodent P2X7 receptor activation and sensitization
The Journal of Biological Chemistry
(2014) - et al.
C-terminal calmodulin-binding motif differentially controls human and rat P2X7 receptor current facilitation
The Journal of Biological Chemistry
(2010)
Identification of novel cholesterol-binding regions in Kir2 channels
The Journal of Biological Chemistry
A Thr357 to Ser polymorphism in homozygous and compound heterozygous subjects causes absent or reduced P2X7 function and impairs ATP-induced mycobacterial killing by macrophages
The Journal of Biological Chemistry
Disruption of lipid rafts inhibits P2X1 receptor-mediated currents and arterial vasoconstriction
The Journal of Biological Chemistry
Differential sensitivity of human platelet P2X1 and P2Y1 receptors to disruption of lipid rafts
Biochemical and Biophysical Research Communications
Trophic activity of a naturally occurring truncated isoform of the P2X7 receptor
The FASEB Journal
Potentiation of native and recombinant P2X7-mediated calcium signaling by arachidonic acid in cultured cortical astrocytes and human embryonic kidney 293 cells
Molecular Pharmacology
Phospholipases C and A2 control lysosome-mediated IL-1 beta secretion: Implications for inflammatory processes
Proceedings of the National Academy of Sciences of the United States of America
Caveolin-1 influences P2X7 receptor expression and localization in mouse lung alveolar epithelial cells
The FEBS Journal
Phosphoinositides regulate P2X4 ATP-gated channels through direct interactions
The Journal of Neuroscience
Acid sphingomyelinase activity triggers microparticle release from glial cells
The EMBO Journal
What does S-palmitoylation do to membrane proteins?
The FEBS Journal
Mechansims of ATP release and inactivation
Leukotriene B4 modulates P2X7 receptor-mediated Leishmania amazonensis elimination in murine macrophages
Journal of Immunology
Endosome-mediated retrograde axonal transport of P2X3 receptor signals in primary sensory neurons
Cell Research
CaMKIIalpha and caveolin-1 cooperate to drive ATP-induced membrane delivery of the P2X3 receptor
Journal of Molecular Cell Biology
The role of palmitoylation and transmembrane domain in sorting of transmembrane adaptor proteins
Journal of Cell Science
Sterols and membrane dynamics
Journal of Chemical Biology
ATP-mediated killing of intracellular mycobacteria by macrophages is a P2X(7)-dependent process inducing bacterial death by phagosome-lysosome fusion
Journal of Immunology
How cholesterol interacts with membrane proteins: An exploration of cholesterol-binding sites including CRAC, CARC, and tilted domains
Frontiers in Physiology
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2022, NeuropharmacologyCitation Excerpt :Lipid rafts and membrane/lipid rafts scaffolding proteins play a beneficial role also in ALS, by protecting spinal motor neurons, preserving neuromuscular function, and extending longevity in a familial mouse model of ALS (Wang et al., 2022). While the ionotropic P2X3 (Vacca et al., 2004) and the metabotropic P2Y12 (Bodin et al., 2003) were the very first purinergic receptors that were identified within lipid rafts, all P2/P1 subtypes and ectonucleotidases are now reported to associate into these specialized submembrane microdomains (Lasley, 2011; D’ Ambrosi and Volonté, 2013; Murrell-Lagnado, 2017). Additional experimental work by FRET, BRET, EM analysis has moreover demonstrated that not only P2X and P2Y subtypes, but also A1 and A2 receptors, associate in these microdomains into hetero-oligomeric complexes (Jiang et al., 2003; Nakata et al., 2005; Ecke et al., 2008; Compan et al., 2012; Schonenbach et al., 2016).
Lithocholic acid inhibits P2X2 and potentiates P2X4 receptor channel gating
2020, Journal of Steroid Biochemistry and Molecular BiologyCitation Excerpt :Neurosteroids have been reported to positively modulate [16–18] or negatively modulate [17] P2X2 and P2X4 receptor function. P2X7 function is modulated by cholesterol and other membrane lipids through direct interactions with the transmembrane domain [21–23]. Little is known about the modulation of P2X receptors by bile acids, other natural circulating products of cholesterol, which are considered to modulate other cell surface receptors and ion channels [24,25].
Direct and indirect cholesterol effects on membrane proteins with special focus on potassium channels
2020, Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids