The tetrapeptide sequence of IL-18 and IL-1ß regulates their recruitment and activation by inflammatory caspases.

Inflammasomes are multiprotein signaling complexes that activate the innate immune system. Canonical inflammasomes recruit and activate caspase-1, which then cleaves and activates IL-1ß and IL-18, as well as gasdermin D (GSDMD) to induce pyroptosis. In contrast, non-canonical inflammasomes, caspases-4/-5 (CASP4/5) in humans and caspase-11 (CASP11) in mice, are known to cleave GSDMD, but their role in direct processing of other substrates besides GSDMD has remained unknown. Here, we show that CASP4/5 but not CASP11 can directly cleave and activate IL-18. However, CASP4/5/11 can all cleave IL-1ß to generate a 27-kDa fragment that deactivates IL-1ß signaling. Mechanistically, we demonstrate that the sequence identity of the tetrapeptide sequence adjacent to the caspase cleavage site regulates IL-18 and IL-1ß recruitment and activation. Altogether, we have identified new substrates of the non-canonical inflammasomes and reveal key mechanistic details regulating inflammation that may aid in developing new therapeutics for immune-related disorders.

The tetrapeptide sequence of IL-1ß regulates its recruitment and activation by inflammatory caspases.

The mammalian innate immune system uses germline-encoded cytosolic pattern-recognition receptors (PRRs) to detect intracellular danger signals. At least six of these PRRs are known to form multiprotein complexes called inflammasomes which activate cysteine proteases known as caspases. Canonical inflammasomes recruit and activate caspase-1 (CASP1), which in turn cleaves and activates inflammatory cytokines such as IL-1ß and IL-18, as well as the pore forming protein, gasdermin D (GSDMD), to induce pyroptotic cell death. In contrast, non-canonical inflammasomes, caspases-4/-5 (CASP4/5) in humans and caspase-11 (CASP11) in mice, are activated by intracellular LPS to cleave GSDMD, but their role in direct processing of inflammatory cytokines has not been established. Here we show that active CASP4/5 directly cleave IL-18 to generate the active species. Surprisingly, we also discovered that CASP4/5/11 cleave IL-1ß at D27 to generate a 27 kDa fragment that is predicted to be inactive and cannot signal to the IL-1 receptor. Mechanistically, we discovered that the sequence identity of the P4-P1 tetrapeptide sequence adjacent to the caspase cleavage site (D116) regulates the recruitment and processing of IL-1ß by inflammatory caspases to generate the bioactive species. Thus, we have identified new substrates of the non-canonical inflammasomes and reveal key mechanistic details regulating inflammation.