TY - JOUR
T1 - K+ regulates Ca2+ to drive inflammasome signaling
T2 - Dynamic visualization of ion flux in live cells
AU - Yaron, J. R.
AU - Gangaraju, S.
AU - Rao, M. Y.
AU - Kong, X.
AU - Zhang, L.
AU - Su, F.
AU - Tian, Y.
AU - Glenn, H. L.
AU - Meldrum, Deirdre
N1 - Funding Information:
Support for this work was provided by the Microscale Life Sciences Center, an NIH NHGRI Center of Excellence in Genomic Science (5P50HG002360 to DRM), and the NIH Common Fund LINCS (Library of Integrated Network-Based Cellular Signatures) program (U01CA164250 to DRM).
Funding Information:
Acknowledgements. Support for this work was provided by the Microscale Life Sciences Center, an NIH NHGRI Center of Excellence in Genomic Science (5P50 HG002360 to DRM), and the NIH Common Fund LINCS (Library of Integrated Network-Based Cellular Signatures) program (U01CA164250 to DRM).
Publisher Copyright:
© 2015 Macmillan Publishers Limited All rights reserved.
PY - 2015/10/1
Y1 - 2015/10/1
N2 - P2X7 purinergic receptor engagement with extracellular ATP induces transmembrane potassium and calcium flux resulting in assembly of the NLRP3 inflammasome in LPS-primed macrophages. The role of potassium and calcium in inflammasome regulation is not well understood, largely due to limitations in existing methods for interrogating potassium in real time. The use of KS6, a novel sensor for selective and sensitive dynamic visualization of intracellular potassium flux in live cells, multiplexed with the intracellular calcium sensor Fluo-4, revealed a coordinated relationship between potassium and calcium. Interestingly, the mitochondrial potassium pool was mobilized in a P2X signaling, and ATP dose-dependent manner, suggesting a role for mitochondrial sensing of cytosolic ion perturbation. Through treatment with extracellular potassium we found that potassium efflux was necessary to permit sustained calcium entry, but not transient calcium flux from intracellular stores. Further, intracellular calcium chelation with BAPTA-AM indicated that P2X induced potassium depletion was independent of calcium mobilization. This evidence suggests that both potassium efflux and calcium influx are necessary for mitochondrial reactive oxygen generation upstream of NLRP3 inflammasome assembly and pyroptotic cell death. We propose a model wherein potassium efflux is necessary for calcium influx, resulting in mitochondrial reactive oxygen generation to trigger the NLRP3 inflammasome.
AB - P2X7 purinergic receptor engagement with extracellular ATP induces transmembrane potassium and calcium flux resulting in assembly of the NLRP3 inflammasome in LPS-primed macrophages. The role of potassium and calcium in inflammasome regulation is not well understood, largely due to limitations in existing methods for interrogating potassium in real time. The use of KS6, a novel sensor for selective and sensitive dynamic visualization of intracellular potassium flux in live cells, multiplexed with the intracellular calcium sensor Fluo-4, revealed a coordinated relationship between potassium and calcium. Interestingly, the mitochondrial potassium pool was mobilized in a P2X signaling, and ATP dose-dependent manner, suggesting a role for mitochondrial sensing of cytosolic ion perturbation. Through treatment with extracellular potassium we found that potassium efflux was necessary to permit sustained calcium entry, but not transient calcium flux from intracellular stores. Further, intracellular calcium chelation with BAPTA-AM indicated that P2X induced potassium depletion was independent of calcium mobilization. This evidence suggests that both potassium efflux and calcium influx are necessary for mitochondrial reactive oxygen generation upstream of NLRP3 inflammasome assembly and pyroptotic cell death. We propose a model wherein potassium efflux is necessary for calcium influx, resulting in mitochondrial reactive oxygen generation to trigger the NLRP3 inflammasome.
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U2 - 10.1038/cddis.2015.277
DO - 10.1038/cddis.2015.277
M3 - Article
C2 - 26512962
AN - SCOPUS:84975292510
SN - 2041-4889
VL - 6
JO - Cell Death and Disease
JF - Cell Death and Disease
IS - 10
M1 - e1954
ER -