title: Lysosomal alkalinization, lipid oxidation, and reduced phagosome clearance triggered by activation of the P2X7 receptor.
abstract: Lysosomal enzymes function optimally at low pH; as accumulation of waste material contributes to cell aging and disease, dysregulation of lysosomal pH may represent an early step in several pathologies. Here, we demonstrate that stimulation of the P2X7 receptor (P2X7R) for ATP alkalinizes lysosomes in cultured human retinal pigmented epithelial (RPE) cells and impairs lysosomal function. P2X7R stimulation did not kill RPE cells but alkalinized lysosomes by 0.3 U. Receptor stimulation also elevated cytoplasmic Ca(2+); Ca(2+) influx was necessary but not sufficient for lysosomal alkalinization. P2X7R stimulation decreased access to the active site of cathepsin D. Interestingly, lysosomal alkalinization was accompanied by a rise in lipid oxidation that was prevented by P2X7R antagonism. Likewise, the autofluorescence of phagocytosed photoreceptor outer segments increased by lysosomal alkalinization was restored 73% by a P2X7R antagonist. Together, this suggests that endogenous autostimulation of the P2X7R may oxidize lipids and impede clearance. The P2X7R was expressed on apical and basolateral membranes of mouse RPE; mRNA expression of P2X7R and extracellular ATP marker NTPDase1 was raised in RPE tissue from the ABCA4(-/-) mouse model of Stargardt's retinal degeneration. In summary, P2X7R stimulation raises lysosomal pH and impedes lysosomal function, suggesting a possible role for overstimulation in diseases of accumulation.
authors: 1Department of Anatomy and Cell Biology, University of Pennsylvania, 440 Levy Bldg., 240 S. 40th St., Philadelphia, PA 19104, USA. chm@exchange.upenn.edu.
authors: Sonia
authors: S
authors: Guha
authors: Gabriel C
authors: GC
authors: Baltazar
authors: Erin E
authors: EE
authors: Coffey
authors: Leigh-Anne
authors: LA
authors: Tu
authors: Jason C
authors: JC
authors: Lim
authors: Jonathan M
authors: JM
authors: Beckel
authors: Shaun
authors: S
authors: Patel
authors: Thor
authors: T
authors: Eysteinsson
authors: Wennan
authors: W
authors: Lu
authors: Ann
authors: A
authors: O'Brien-Jenkins
authors: Alan M
authors: AM
authors: Laties
authors: Claire H
authors: CH
authors: Mitchell
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000235:genetics; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism; D002417:Cattle; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D003593:Cytoplasm / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D050356:Lipid Metabolism; D008247:Lysosomes / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D010084:Oxidation-Reduction; D010588:Phagosomes / Q000378:metabolism*; D058486:Receptors, Purinergic P2X7 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012162:Retinal Degeneration / Q000235:genetics / Q000378:metabolism; D055213:Retinal Pigment Epithelium / Q000166:cytology / Q000378:metabolism; D014158:Transcription, Genetic
chemical_list: D018528:ATP-Binding Cassette Transporters; C512841:Abca4 protein, mouse; D058486:Receptors, Purinergic P2X7; D000255:Adenosine Triphosphate; D002118:Calcium
keywords: autophagy; lipofuscin; purinergic signaling; retinal degeneration
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