Inhibitory effects of chloride on the activation of caspase-1, IL-1beta secretion, and cytolysis by the P2X7 receptor.
The P2X7 receptor (P2X7R) is an ATP-gated cation channel that activates caspase-1 leading to the maturation and secretion of IL-1beta. Because previous studies indicated that extracellular Cl- exerts a negative allosteric effect on ATP-gating of P2X7R channels, we tested whether Cl- attenuates the P2X7R-->caspase-1-->IL-1beta signaling cascade in murine and human macrophages. In Bac1 murine macrophages, substitution of extracellular Cl- with gluconate produced a 10-fold increase in the rate and extent of ATP-induced IL-1beta processing and secretion, while reducing the EC50 for ATP by 5-fold. Replacement of Cl- with gluconate also increased the potency of ATP as an inducer of mature IL-1beta secretion in primary mouse bone marrow-derived macrophages and in THP-1 human monocytes/macrophages. Our observations were consistent with actions of Cl- at three levels: 1) a negative allosteric effect of Cl-, which limits the ability of ATP to gate the P2X7R-mediated cation fluxes that trigger caspase-1 activation; 2) an intracellular accumulation of Cl- via nonselective pores induced by P2X7R with consequential repression of caspase-1-mediated processing of IL-1beta; and 3) a facilitative effect of Cl- substitution on the cytolytic release of unprocessed pro-IL-1beta that occurs with sustained activation of P2X7R. This cytolysis was repressed by the cytoprotectant glycine, permitting dissociation of P2X7R-regulated secretion of mature IL-1beta from the lytic release of pro-IL-1beta. These results suggest that under physiological conditions P2X7R are maintained in a conformationally restrained state that limits channel gating and coupling of the receptor to signaling pathways that regulate caspase-1.
Verhoef
Philip A
PA
Department of Physiology and Biophysics, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106-4970, USA.
Kertesy
Sylvia B
SB
Lundberg
Kathleen
K
Kahlenberg
J Michelle
JM
Dubyak
George R
GR
D000818:Animals; D015153:Blotting, Western; D020170:Caspase 1 / Q000276:immunology / Q000378:metabolism*; D002478:Cells, Cultured; D002712:Chlorides / Q000378:metabolism*; D004789:Enzyme Activation / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D005942:Gluconates / Q000378:metabolism; D006801:Humans; D007375:Interleukin-1 / Q000276:immunology / Q000378:metabolism*; D008264:Macrophages / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D018048:Receptors, Purinergic P2 / Q000276:immunology / Q000378:metabolism*; D058486:Receptors, Purinergic P2X7; D015398:Signal Transduction / Q000276:immunology; D048051:p38 Mitogen-Activated Protein Kinases / Q000276:immunology / Q000378:metabolism
D002712:Chlorides; D005942:Gluconates; D007375:Interleukin-1; C546468:P2RX7 protein, human; C546469:P2rx7 protein, mouse; D018048:Receptors, Purinergic P2; D058486:Receptors, Purinergic P2X7; D048051:p38 Mitogen-Activated Protein Kinases; D020170:Caspase 1; C030691:gluconic acid