Original Article

Citation: Cell Death and Disease (2015) 6, e1954; doi:10.1038/cddis.2015.277
Published online 29 October 2015

K+ regulates Ca2+ to drive inflammasome signaling: dynamic visualization of ion flux in live cells
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J R Yaron1,2, S Gangaraju1, M Y Rao1, X Kong1, L Zhang1, F Su1, Y Tian1, H L Glenn1 and D R Meldrum1

  1. 1Center for Biosignatures Discovery Automation, The Biodesign Institute, Arizona State University, Tempe, 85287 AZ, USA
  2. 2Biological Design Graduate Program, School of Biological and Health Systems Engineering, Ira A. Fulton Schools of Engineering, Arizona State University, Tempe, 85287 AZ, USA

Correspondence: DR Meldrum, Center for Biosignatures Discovery Automation, The Biodesign Institute, Arizona State University, 1001S. McAllister Avenue, PO Box 877101, Tempe, AZ 85287-7101, USA. Tel: 480 727 9397; Fax: 480 965 2337; E-mail: Deirdre.Meldrum@asu.edu

Received 8 May 2015; Revised 27 August 2015; Accepted 31 August 2015

Edited by G Amarante-Mendes

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Abstract

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 P2X7 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 P2X7-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.

Abbreviations:

NLRP3, NLR family, pyrin domain-containing 3; IL-1β, interleukin-1 beta; P2X7, purinergic receptor P2X, ligand-gated ion channel, 7; ROS, reactive oxygen species; mROS, mitochondrial reactive oxygen species; TXNIP, thioredoxin-interacting protein; LPS, lipopolysaccharide