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Inflammasomes are cytosolic protein complexes that activate caspase-1, an inflammatory protease that converts pro-interleukin (IL)-1β and IL-18 into their secreted, bioactive forms (1); and mechanisms regulating inflammasome activation have attracted [...]

Intracellular lipopolysaccharide from Gram-negative bacteria including Escherichia coli, Salmonella typhimurium, Shigella flexneri, and Burkholderia thailandensis activates mouse caspase-11, causing pyroptotic cell death, interleukin-1β processing, and lethal septic shock. How caspase-11 executes these downstream signalling events is largely unknown. Here we show that gasdermin D is essential for caspase-11-dependent pyroptosis and interleukm-1β maturation. A forward genetic screen with ethyl-N-nitrosourea-mutagenized mice links Gsdmd to the intracellular lipopolysaccharide response. Macrophages from [Gsdmd.sup.-/-] mice generated by gene targeting also exhibit defective pyroptosis and interleukin-1β secretion induced by cytoplasmic lipopolysaccharide or Gram-negative bacteria. In addition, [Gsdmd.sup.-/-] mice are protected from a lethal dose of lipopolysaccharide. Mechanistically, caspase-11 cleaves gasdermin D, and the resulting amino-terminal fragment promotes both pyroptosis and NLRP3-dependent activation of caspase-1 in a cell-intrinsic manner. Our data identify gasdermin D as a critical target of caspase-11 and a key mediatorof the host response against Gram-negative bacteria., Cytoplasmic caspase-11 (also known as caspase-4) defines the non-canonical inflammasome that is activated by various Gram-negative bacterial infections and causes infected cells to die by pyroptosis (1-4). Caspase-11 also triggers [...]

T-helpertype 17 ([T.sub.H]17) cells that produce the cytokines interleukin-17A (IL-17A) and IL-17F are implicated in the pathogenesis of several autoimmune diseases (1,2). The differentiation of [T.sub.H]17 cells is regulated by [...]

Lipopolysaccharide from Gram-negative bacteria is sensed in the host cell cytoplasm by a non-canonical inflammasome pathway that ultimately results in caspase-11 activation and cell death (1-3). In mouse macrophages, activation [...]

NLRC4 is a cytosolic member of the NOD-like receptor family that is expressed in innate immune cells. It senses indirectly bacterial flagellin and type III secretion systems, and responds by [...]

Inflammasomes are cytosolic multiprotein complexes assembled by intracellular nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) and they initiate innate immune responses to invading pathogens and danger signals by activating caspase-1 (ref. [...]

Caspase-1 activation by inflammasome scaffolds comprised of intracellular nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) and the adaptor ASC is believed to be essential for production of the pro-inflammatory cytokines interleukin (IL)-1β and IL-18 during the innate immune response (1-5). Here we show, with C57BL/6 Casp11 gene-targeted mice, that caspase-11 (also known as caspase-4) (6-8) is critical for caspase-1 activation and IL-1β production in macrophages infected with Escherichia coli, Citrobacter rodentium or Vibrio cholerae. Strain 129 mice, like [Casp11.sup.-/-] mice, exhibited defects in IL-1β production and harboured a mutation in the Casp11 locus that attenuated caspase-11 expression. This finding is important because published targeting of the Casp1 gene was done using strain 129 embryonic stem cells (9,10). Casp1 and Casp11 are too close in the genome to be segregated by recombination; consequently, the published [Casp1.sup.-/-] mice lack both caspase-11 and caspase-1. Interestingly, [Casp11.sup.-/-] macrophages secreted IL-1β normally in response to ATP and monosodium urate, indicating that caspase-11 is engaged by a non-canonical inflammasome. [Casp1.sup.-/-][Casp11.sup.129mt/129mt] macrophages expressing caspase-11 from a C57BL 6 bacterial artificial chromosome transgene failed to secrete IL-1β regardless of stimulus, confirming an essential role for caspase-1 in IL-1β production. Caspase-11 rather than caspase-1, however, was required for non-canonical inflammasome-triggered macrophage cell death, indicating that caspase-11 orchestrates both caspase-1-dependent and -independent outputs. Caspase-1 activation by non-canonical stimuli required NLRP3 and ASC, but caspase-11 processing and cell death did not, implying that there is a distinct activator of caspase-11. Lastly, loss of caspase-11 rather than caspase-1 protected mice from a lethal dose of lipopolysaccharide. These data highlight a unique pro-inflammatory role for caspase-11 in the innate immune response to clinically significant bacterial infections., Many bacterial toxins promote inflammasome activation (11-15), and this is also true for cholera toxin B (CTB), a component of the AB5 holotoxin complex (Fig. 1a and Supplementary Fig. 1). [...]

Macrophages respond to cytosolic nucleic acids by activating cysteine protease caspase-1 within a complex called the inflammasome. Subsequent cleavage and secretion of proinflammatory cytokines IL-1[beta] and IL-18 are critical for innate immunity. Here, we show that macrophages from mice lacking absent in melanoma 2 (AIM2) cannot sense cytosolic double-stranded DNA and fail to trigger inflammasome assembly. Caspase-1 activation in response to intracellular pathogen Francisella tularensis also required AIM2. Immunofluorescence microscopy of macrophages infected with F. tularensis revealed striking colocalization of bacterial DNA with endogenous AIM2 and inflammasome adaptor ASC. By contrast, type I IFN (IFN-[alpha] and -[beta]) secretion in response to F. tularensis did not require AIM2. IFN-I did, however, boost AIM2-dependent caspase-1 activation by increasing AIM2 protein levels. Thus, inflammasome activation was reduced in infected macrophages lacking either the IFN-I receptor or stimulator of interferon genes (STING). Finally, AIM2-deficient mice displayed increased susceptibility to F. tularensis infection compared with wild-type mice. Their increased bacterial burden in vivo confirmed that AIM2 is essential for an effective innate immune response. inflammasome | innate immunity | interferon | apoptosis-associated specklike protein containing a caspase recruitment domain www.pnas.org/cgi/doi/10.1073/pnas.1003738107

Fas ligand (FasL) is a member of the tumor necrosis factor family and induces apoptosis in Fas (CD95)bearing target cells. In this study, we generated several mAbs that react with mouse FasL (mFasL) and characterized their functional properties. One of these mAbs, K10, specifically reacted with mFasL derived from C57BL/6 (B6) mice, but not that from BALB/c mice as estimated by surface staining and blocking of cytotoxic activities of mFasL transfectants, suggesting a polymorphism of mFasL. Sequence analysis of mFasL cDNA from several strains revealed that BALB/c and DBA/2 mice have three nucleotide differences from the known B6 and C3H sequences, which result in two amino acid substitutions (Thr-184 [approaches] Ala-184 and Glu-218 [approaches] Gly-218) in the extracellular region. Analysis of the K10 reactivity and genotyping by PCR-restriction fragment length polymorphism revealed that inbred mice segregate into the following two allotypes: mFasL.1 (B6, C3H, MRL, SJL, NOD, NZB, NZW) and mFasL.2 (BALB/c, DBA/1, DBA/2). Interestingly, COS7 cells expressing BALB/c FasL lysed Fas-bearing target cells more efficiently than those expressing B6 FasL. Furthermore, BALB/c-derived CDS-FasL fusion protein, which is composed of the extracellular domains of human CD8[Alpha] and mFasL, exhibited 9-fold higher specific activity than did B6-derived CD8-FasL. These results suggest that in mFasL.2 mice the Fas/FasL system works more effectively than in mFasL.1 mice.

Bellgrau and colleagues misinterpreted the concept of CD95 ligand (CD95 L) in graft rejection. A baby hamster kidney fibroblast cell line expressing transfected human CD95L complementary DNA was transplanted into nude mice. The injection of anti-CD95L reversed the rejection of the ligand transfectant, indicating that CD95L induces the rejection. In vivo experiments indicate that granulocytes are involved in the rejection process. CD95L binds to neutrophils and stimulates their cytotoxic mechanism, causing acute graft rejection.