LRRC8A is essential for hypotonicity-, but not for DAMP-induced NLRP3 inflammasome activation.

Jack P Green,Shi Yu,David Brough,Tessa Swanton,James Cook,James Beswick,Antony D Adamson,Neil E Humphreys,Catherine Lawrence,Richard Bryce,Sally Freeman,Lucy V Morris,Lina Y El-Sharkawy

doi:10.7554/elife.59704

Abstract

The NLRP3 inflammasome is a multi-molecular protein complex that converts inactive cytokine precursors into active forms of IL-1β and IL-18. The NLRP3 inflammasome is frequently associated with the damaging inflammation of non-communicable disease states and is considered an attractive therapeutic target. However, there is much regarding the mechanism of NLRP3 activation that remains unknown. Chloride efflux is suggested as an important step in NLRP3 activation, but which chloride channels are involved is still unknown. We used chemical, biochemical, and genetic approaches to establish the importance of chloride channels in the regulation of NLRP3 in murine macrophages. Specifically, we identify LRRC8A, an essential component of volume-regulated anion channels (VRAC), as a vital regulator of hypotonicity-induced, but not DAMP-induced, NLRP3 inflammasome activation. Although LRRC8A was dispensable for canonical DAMP-dependent NLRP3 activation, this was still sensitive to chloride channel inhibitors, suggesting there are additional and specific chloride sensing and regulating mechanisms controlling NLRP3.

Highlights

Inflammation is an important protective host-response to infection and injury, and yet is detrimental during non-communicable diseases (Dinarello et al, 2012)
The discovery that the Cl- channel regulating the regulatory volume decrease (RVD) (VRAC) was composed of LRRC8 sub-units, and that LRRC8A was essential for channel activity, offered us the opportunity to investigate the direct importance of volume-regulated anion channels (VRAC) in the regulation of NLRP3
Hypotonicity induces cell swelling which is corrected by the VRAC-dependent RVD (Qiu et al, 2014; Voss et al, 2014)

Summary

Introduction

Inflammation is an important protective host-response to infection and injury, and yet is detrimental during non-communicable diseases (Dinarello et al, 2012). When the macrophages are exposed to molecules produced by damaged cells or toxins from bacteria, Green et al discovered that other previously unidentified chloride channels are involved in activating the NLRP3 inflammasome. These results suggest that it might be possible to develop drugs to prevent the activation of the NLRP3 inflammasome that selectively target specific sets of chloride channels depending on which stimuli are causing the inflammation. Such a selective approach would minimise the side effects associated with drugs that generically suppress all NLRP3 activity by directly binding to NLRP3 itself. We provide genetic evidence for the importance of Cl- in regulating NLRP3 via the VRAC dependence of the hypotonicity response, and suggest the presence of additional Cl- sensing mechanisms regulating NLRP3 in response to DAMPs

Results

Discussion

Materials and methods