<i>Salmonella</i> Flagellin Activates NAIP/NLRC4 and Canonical NLRP3 Inflammasomes in Human Macrophages

Abstract

Infection of human macrophages with <i>Salmonella enterica</i> serovar Typhimurium (<i>S.</i> Typhimurium) leads to inflammasome activation. Inflammasomes are multiprotein complexes facilitating caspase-1 activation and subsequent gasdermin D-mediated cell death and IL-1β and IL-18 cytokine release. The NAIP/NLRC4 inflammasome is activated by multiple bacterial protein ligands, including flagellin from the flagellum and the needle protein PrgI from the <i>S.</i> Typhimurium type III secretion system. In this study, we show that transfected ultrapure flagellin from <i>S</i> Typhimurium induced cell death and cytokine secretion in THP-1 cells and primary human monocyte-derived macrophages. In THP-1 cells, NAIP/NLRC4 and NLRP3 played redundant roles in inflammasome activation during infection with <i>S.</i> Typhimurium. Knockout of <i>NAIP</i> or <i>NLRC4</i> in THP-1 cells revealed that flagellin, but not PrgI, now activated the NLRP3 inflammasome through a reactive oxygen species- and/or cathepsin-dependent mechanism that was independent of caspase-4/5 activity. In conclusion, our data suggest that NLRP3 can be activated by flagellin to act as a "safety net" to maintain inflammasome activation under conditions of suboptimal NAIP/NLRC4 activation, as observed in THP-1 cells, possibly explaining the redundant role of NLRP3 and NAIP/NLRC4 during <i>S.</i> Typhimurium infection.