CASP5 inflammasome assembly

CASP5 inflammasome assembly Caspase-5 (CASP5), an inflammatory caspase closely related to caspase-4, is involved in the innate immune response triggered by cytosolic lipopolysaccharide (LPS), a component of the outer membrane of Gram-negative bacteria (Shi J et al., 2014, 2015; reviewed by Eckhart L & Fischer H, 2024). CASP5 expression is low at baseline but strongly induced by pro-inflammatory stimuli such as LPS through TLR4-dependent NF-κB signaling (Lin XY et al., 2000; Eckhart L et al., 2006; Viganò E et al., 2015). In human peripheral blood mononuclear cells (PBMCs), LPS stimulation results in a >10-fold increase in CASP5 expression levels, a greater induction than that observed for CASP1 or CASP4 (Eckhart L et al., 2006). Similarly, CASP5 shows higher LPS-inducibility in human monocytes at both mRNA and protein levels compared to CASP4 (Viganò E et al., 2015; Cheng Y et al., 2023). Both interferon γ (IFNγ) and IFNα/β may contribute to CASP5 upregulation by activating JAK-STAT-dependent transcription (Lin XY et al., 2000; Eckhart L et al., 2006; Rooney M et al., 2024). Promoter analysis reveals conserved NF-κB and STAT binding motifs (Eckhart L et al., 2006), and expression is highest in immune-related tissues including blood, spleen, lung, and colon, as per GTEx data (Eckhart L & Fischer H 2024). CASP5, like CASP4, is activated when intracellular bacterial LPS binds to its N-terminal caspase activation and recruitment domain (CARD), promoting oligomerization and proximity-induced activation of CASP5 (Shi J et al., 2014). CASP5 undergoes autocatalytic cleavage at aspartate residues, yielding large and small subunits that assemble into the active CASP5 heterotetramer (Shi J et al., 2014; Viganò et al., 2015). Activated CASP5 can then cleave gasdermin D (GSDMD), which is also a substrate of CASP1, CASP4, 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). CASP5 also processes pro-IL-18 and pro-IL-1β. Like CASP4, CASP5 efficiently cleaves pro-interleukin-18 (pro-IL-18) at aspartic acid residue D36 to generate its mature, biologically active form (Shi X et al., 2023; Devant P et al., 2023; Exconde PM et al., 2023; reviewed by Exconde PM, 2024). Structural and biochemical analyses revealed that this cleavage relies on a bivalent recognition mechanism, in which pro-IL-18 binds CASP4/CASP5 through two interfaces: the protease exosite binds a hydrophobic pocket within pro-IL-18, while the active site of caspase 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, CASP5- and CASP4-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).Cytosolic delivery of LPS can occur via endocytosis of outer membrane vesicles (OMVs) released by Gram-negative bacteria (Wacker MA et al., 2017; Bitto NJ et al., 2018; reviewed in Barker JH and Weiss JP, 2019; Page MJ et al., 2022), or through phagolysosomal rupture following bacterial uptake. Guanylate-binding protein 1 (GBP1) directly binds LPS on cytosol-exposed bacteria and, along with other GBPs (GBP2, GBP3 and GBP4, forms a coat on the bacterial surface, creating a platform that recruits CASP4 to facilitate inflammasome activation (Santos JC et al., 2020; Wandel MP et al., 2020). While CASP5 likely follows a similar GBP-mediated recruitment mechanism, there is currently no direct evidence for a physical interaction between CASP5 and GBPs, and this interaction is therefore not shown here.