Caspase-11 detection of intracellular lipopolysaccharide (LPS) from invasive Gram-negative bacteria mediates noncanonical activation of the NLRP3 inflammasome. While avirulent bacteria do not invade the cytosol, their presence in tissues necessitates clearance and immune system mobilization. Despite sharing LPS, only live avirulent Gram-negative bacteria activate the NLRP3 inflammasome. Here, we found that bacterial mRNA, which signals bacterial viability, was required alongside LPS for noncanonical activation of the NLRP3 inflammasome in macrophages. Concurrent detection of bacterial RNA by NLRP3 and binding of LPS by pro-caspase-11 mediated a pro-caspase-11–NLRP3 interaction before caspase-11 activation and inflammasome assembly. LPS binding to pro-caspase-11 augmented bacterial mRNA-dependent assembly of the NLRP3 inflammasome, while bacterial viability and an assembled NLRP3 inflammasome were necessary for activation of LPS-bound pro-caspase-11. Thus, the pro-caspase-11–NLRP3 interaction nucleated a scaffold for their interdependent activation explaining their functional reciprocal exclusivity. Our findings inform new vaccine adjuvant combinations and sepsis therapy. Blander and colleagues show that concurrent detection of LPS and bacterial RNA triggers the interaction of procaspase-11 with NLRP3, upstream of the activation of either receptor and before NLRP3–ASC oligomerization.

Caspase-11 interaction with NLRP3 potentiates the noncanonical activation of the NLRP3 inflammasome