Enteritidis antitoxin DinJ inhibits NLRP3-dependent canonical inflammasome activation in macrophages.

The inflammasome is a pivotal component of the innate immune system, acting as a multiprotein complex that plays an essential role in detecting and responding to microbial infections. Enteritidis have evolved multiple mechanisms to regulate inflammasome activation and evade host immune system clearance. Through screening . Enteritidis C50336Δ transposon mutant library, we found that the insertion mutant of increased inflammasome activation. In this study, we demonstrated the genetic connection between the antitoxin DinJ and the toxin YafQ in . Enteritidis, confirming their co-transcription. The deletion mutant ΔΔ increased cell death and IL-1β secretion in J774A.1 cells. Western blotting analysis further showed elevated cleaved Caspase-1 product (p10 subunits) and IL-1β secretion in cells infected with ΔΔ compared to cells infected with Δ. DinJ was found to inhibit canonical inflammasome activation using primary bone marrow-derived macrophages (BMDMs) from C57BL/6 mice. Furthermore, DinJ specifically inhibited NLRP3 inflammasome activation, as demonstrated in BMDMs from and mice. Fluorescence resonance energy transfer (FRET) experiments confirmed the translocation of DinJ into host cells during infection. Finally, we revealed that DinJ could inhibit the secretion of IL-1β and IL-18 , contributing to . Enteritidis evading host immune clearance. In summary, our findings provide insights into the role of DinJ in modulating the inflammasome response during . Enteritidis infection, highlighting its impact on inhibiting inflammasome activation and immune evasion.