Abstract
The innate immune system acts as the first line of defense against infection. One key component of the innate immune response to gram-negative bacterial infections is inflammasome activation. The caspase-11 (CASP11)-nucleotide-binding oligomerization domain-like receptor pyrin domain-containing 3 (NLRP3) inflammasome is activated by cytosolic lipopolysaccharide, a gram-negative bacterial cell wall component, to trigger pyroptosis and host defense during infection. Although several cellular signaling pathways have been shown to regulate CASP11-NLRP3 inflammasome activation in response to lipopolysaccharide, the upstream molecules regulating CASP11 activation during infection with live pathogens remain unclear. Here, we report that the understudied caspase-6 (CASP6) contributes to the activation of the CASP11-NLRP3 inflammasome in response to infections with gram-negative bacteria. Using in vitro cellular systems with bone marrow-derived macrophages and 293T cells, we found that CASP6 can directly process CASP11 by cleaving at Asp59 and Asp285, the CASP11 auto-cleavage sites, which could contribute to the activation of CASP11 during gram-negative bacterial infection. Thus, the loss of CASP6 led to impaired CASP11-NLRP3 inflammasome activation in response to gram-negative bacteria. These results demonstrate that CASP6 potentiates activation of the CASP11-NLRP3 inflammasome to produce inflammatory cytokines during gram-negative bacterial infections.
Highlights
The inflammasome is a multiprotein complex which canonically consists of a sensor, the adaptor molecule apoptosis-associated specklike protein containing a CARD (ASC) and the inflammatory caspase, caspase-1 (CASP1) [1,2]
To investigate whether caspase-6 activation. TRIF-dependent signaling (CASP6) had a role in transcription factor IRF8 contributes to the the regulation of the CASP11-NLRP3 induction of the type I interferon (IFN) response inflammasome during gram-negative bacterial during gram-negative bacterial infection and infections, we infected bone marrow-derived promotes CASP11-NLRP3 inflammasome macrophages (BMDMs) isolated from WT and activation without affecting the CASP11
We found that the p26 fragment was produced when CASP1, CASP3, CASP6, or CASP8 were individually co-expressed with the catalytically dead CASP11 (Figure 5B)
Summary
CASP6 has recently been shown to promote activation of the ZBP1-NLRP3 inflammasome in response to IAV infection [19]. Consistent with the mRNA levels, the protein expression of NLRP3, CASP11 and pro– IL-1β was not impaired in E. coli or C. rodentium infected Casp6–/– BMDMs (Figures 2C and 2D). The expression of the adaptor protein of the CASP11-NLRP3 inflammasome, ASC, was comparable between WT and Casp6–/– BMDMs during both E. coli and C. rodentium infection (Figures 2C and 2D). Casp6CA/CA and Casp6–/– BMDMs compared with with GSDMD resulted in cleavage of GSDMD; that in WT BMDMs in response to E. coli and C. overexpression of CASP6 with rodentium infections (Figures 4C and 4D). Mutating Asp to an Ala in CASP11 indicate that the catalytic activity of CASP6 reduced the production of the p26 fragment and contributes to gram-negative bacteria-induced led to cleavage at alternative sites (Figure 5D), cell death and CASP11-NLRP3 inflammasome suggesting that CASP6 can cleave after Asp in activation. 4D), we hypothesized that CASP6 may be directly involved in CASP11 processing
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