Irgm2 and Gate‐16 cooperatively dampen Gram‐negative bacteria‐induced caspase‐11 response

Abstract

Inflammatory caspase‐11 (rodent) and caspases‐4/5 (humans) detect the Gram‐negative bacterial component LPS within the host cell cytosol, promoting activation of the non‐canonical inflammasome. Although non‐canonical inflammasome‐induced pyroptosis and IL‐1‐related cytokine release are crucial to mount an efficient immune response against various bacteria, their unrestrained activation drives sepsis. This suggests that cellular components tightly control the threshold level of the non‐canonical inflammasome in order to ensure efficient but non‐deleterious inflammatory responses. Here, we show that the IFN‐inducible protein Irgm2 and the ATG8 family member Gate‐16 cooperatively counteract Gram‐negative bacteria‐induced non‐canonical inflammasome activation, both in cultured macrophages and in vivo. Specifically, the Irgm2/Gate‐16 axis dampens caspase‐11 targeting to intracellular bacteria, which lowers caspase‐11‐mediated pyroptosis and cytokine release. Deficiency in Irgm2 or Gate16 induces both guanylate binding protein (GBP)‐dependent and GBP‐independent routes for caspase‐11 targeting to intracellular bacteria. Our findings identify molecular effectors that fine‐tune bacteria‐activated non‐canonical inflammasome responses and shed light on the understanding of the immune pathways they control.

Phospholipid peroxidation fuels ExoU phospholipase-dependent cell necrosis and supportsPseudomonas aeruginosa-driven pathology

“…Murine Bone Marrow-Derived Macrophages (BMDMs) from bone marrow progenitors were differentiated in DMEM (Invitrogen) supplemented with 10% v/v FCS (Thermo Fisher Scientific), 10% v/v MCSF (L929 cell supernatant), 10 mM HEPES (Invitrogen), and nonessential amino acids (Invitrogen) for 7 days as previously described (Eren et al, 2020).…”

“…Immortalized murine bone-marrow derived macrophages have been described previously (Eren et al, 2020). U937 cells were cultured in RPMI glutamax medium containing 10% FBS, 100 IU/ml penicillin and 10 μg/ml streptomycin, 1 mM sodium pyruvate (Life Technologies), and 50 μM 2-ME (Sigma-Aldrich).…”

“…Monocytes were isolated from Peripheral Blood Mononuclear Cells (PBMCs) from the buffy coat of healthy donors obtained from the EFS Toulouse Purpan (France) as described previously (Troegeler et al, 2014). Briefly, PBMCs were isolated by centrifugation using standard Ficoll-Paque density (GE Healthcare) (Eren et al, 2020). The blood was diluted 1:1 in phosphate-buffered saline (PBS) pre-warmed to 37°C and carefully layered over the Ficoll-Paque gradient.…”

“…Cell lysate generation has been described previously (Eren et al, 2020). Briefly, proteins were loaded in 12% SDS-PAGE gels and then transferred on PVDF membranes.…”

Therapeutic implications of inflammasome in inflammatory bowel disease

“…Moreover, the observed effects of IRGM2 were independent of NLRP3 activation. 142,143 Further studies are warranted to explore the role of IRGM in IBD pathogenesis and utilize IRGM as a therapeutic target for treating intestinal inflammation. NEK7 a component of NLRP3 is also shown to modulate NLRP3 activation and subsequently induce pyroptosis in DSS-induced chronic colitis in mice.…”