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.
Highlights
NLR family, pyrin domain-containing 3 (NLRP3) is the most extensively studied among the inflammasome family of caspase-1-activating complexes and is critical to the innate immune response to infection, damage and pathophysiological dysfunction.[1]
We propose that mitochondrial ROS generation is a downstream effect of potassium efflux-dependent calcium influx and defines a coordinated, ion flux-driven regulation of the NLRP3 inflammasome via oxidative signaling
The release of both active components was abolished in the presence of high extracellular potassium as well as the selective, competitive, P2X7 receptor antagonist A438079.20 The requirement for potassium efflux in inflammasome-mediated pyroptotic cell death was confirmed by propidium iodide staining and live-cell imaging (Figure 1b)
Summary
NLR family, pyrin domain-containing 3 (NLRP3) is the most extensively studied among the inflammasome family of caspase-1-activating complexes and is critical to the innate immune response to infection, damage and pathophysiological dysfunction.[1]. The asymmetric distribution of ions in cellular compartments establishes a gradient such that, under conditions of membrane permeability, ions rapidly diffuse across the gradient without energy input.[8] Cells benefit from asymmetric ion distribution by using it to affect rapid processes such as neuronal action potentials.[8] Recent work has implicated potassium flux as the common trigger in regulating NLRP3 inflammasome activity.[9] it has been understood for over two decades that potassium flux regulates the processing of interleukin (IL)-1β, a downstream effect of inflammasome activation.[10,11] While potassium is the most commonly studied ion posited to regulate the NLRP3 pathway, calcium flux has gained popularity in recent years because intervention in calcium mobilization has inhibitory effects on inflammasome activity.[12,13,14] Both ions are permeant to the non-specific cation channel formed by plasma membrane expressed purinergic receptor P2X, ligand-gated ion channel, 7 (P2X7) which is activated by external ATP. We propose that mitochondrial ROS generation is a downstream effect of potassium efflux-dependent calcium influx and defines a coordinated, ion flux-driven regulation of the NLRP3 inflammasome via oxidative signaling
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
