Activated CASP4 can then cleave gasdermin D (GSDMD), which is also a substrate of CASP1, CASP5, and Casp11, a murine homolog of human CASP4/CASP5 (Shi J et al., 2014, 2015; Kayagaki N et al., 2015; Zhao Y et al., 2018; Wang K et al., 2020). The resulting N-terminal fragment of GSDMD oligomerizes to form pores in the cell membrane, leading to pyroptosis in mammals (Liu X et al., 2016; Ding J et al., 2016; Sborgi L et al., 2016; Aglietti RA et al., 2016). In addition, CASP4 and CASP5 cleave pro-interleukin-18 (pro-IL-18) at aspartic acid D36 with high efficiency, producing the mature, active cytokine (Shi X et al., 2023; Exconde PM et al., 2023; Devant P et al., 2023; reviewed by Exconde PM, 2024). Structural analyses revealed that this cleavage relies on a bivalent recognition mechanism, in which pro-IL-18 binds caspase-4 through two interfaces: the protease exosite binds a hydrophobic pocket within pro-IL-18, while the active site of CASP4 engages charged residues located within and adjacent to the tetrapeptide recognition motif in the pro-domain (Shi X et al., 2023; Devant P et al., 2023). In contrast, CASP4- and CASP5-mediated cleavage of pro-IL-1β at D27 produces an inactive fragment that lacks receptor-stimulating activity (Exconde PM et al., 2023; reviewed by Exconde PM, 2024). An alternative CASP4-mediated cleavage at D116, the canonical pro-IL-1β activation site, has been observed but occurs with lower efficiency comparing to pro-IL-18 processing (Bibo-Verdugo B et al., 2020; Chan AH et al., 2023; Devant P et al., 2023).

Intracellular bacterial pathogens have evolved strategies to suppress host inflammatory responses. For example, Shigella flexneri secretes the type III secretion system (T3SS) effector OspC3, which catalyzes ADP-riboxanation of CASP4, thereby inhibiting LPS-induced, CASP4-mediated pyroptosis (Li Z et al., 2021; Hou Y et al., 2023).