Neutrophil IL-1β secretion induced by ExoS expressing Pseudomonas aeruginosa is dependent on NLRP3 and Gasdermin D

Abstract

Macrophages infected with Gram-negative bacteria expressing flagellin or Type III secretion system (T3SS) structural proteins are known to activate the NLRC4 inflammasome, resulting in caspase-1 and Gasdermin D (GSDMD) cleavage, IL-1β secretion and pyroptotic cell death. We examined the role of these mediators in IL-1β secretion by neutrophils infected with Pseudomonas aeruginosa strain PAO1 that expresses the Type III secretion system (T3SS) effectors ExoS and ExoT. We found that IL-1β secretion by neutrophils was dependent on expression of the T3SS needle and translocon proteins. Although pro-GSDMD and pro-GSDME were processed in PAO1 infected neutrophils, only GSDMD was required for IL-1β secretion. PAO1 – induced IL-1β secretion by macrophages was NLRC4 dependent, IL-1β secretion by neutrophils utilized NLRC4 only in the absence of P. aeruginosa exoenzymes. Instead, PAO1 – induced IL-1β secretion required NLRP3, which mediated by ExoS ADP ribosyl transferase activity. Overall, these findings reveal fundamental differences between neutrophils and macrophages in IL-1β secretion in response to pathogenic bacteria.

Pyroptosis in neutrophils: Multimodal integration of inflammasome and regulated cell death signaling pathways

“…Conversely, Chauhan et al 75 used GSDMD‐deficient neutrophils to demonstrate GSDMD was completely dispensable for the activation of NETosis by phorbol ester, the well‐established inducer of NADPH oxidase‐dependent NETosis. We have also observed identical rates and extents of phorbol ester‐induced NETosis in neutrophils isolated from wildtype C57/Bl6 mice, Gsdmd −/− mice, or Gsdme −/− mice 189 . Together, these recent reports suggest a contingent, rather than obligatory, role for GSDMD in regulating NETosis.…”

“…We have also observed identical rates and extents of phorbol ester-induced NETosis in neutrophils isolated from wildtype C57/Bl6 mice, Gsdmd −/− mice, or Gsdme −/− mice. 189 Together, these recent reports suggest a contingent, rather than obligatory, role for GSDMD in regulating NETosis. Clearly, more investigation is required to further clarify the signaling pathways that underlie this contingent role for GSDMD in an important component of neutrophil-mediated innate immunity.…”

“…We recently demonstrated that P. aeruginosa activates the NLRP3 rather than the NLRC4 inflammasome in murine neutrophils (though not macrophages) when infected with Pseudomonas aeruginosa expressing the ExoS-ADPRT virulence factor delivered via the T3SS. 189 A critical factor in determining this skewing to NLRP3-versus NLRC4-inflammasomes (as described by Santoni et al 79 ) is priming the neutrophils with LPS before infection, or using elicited neutrophils from the inflamed peritoneal cavity, both of which conditions upregulate NLRP3 expression. We also observed that GSDMD is required for IL-1β release and pyroptosis in the P. aeruginosa-infected neutrophils but is entirely dispensable for NETosis.…”