The tetrapeptide sequence of IL-1β 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-1β 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-1β 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-1β 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.