Entamoeba histolytica ‐Induced Activation of Caspase‐4 Regulates Gasdermin D Cleavage to Mediate IL‐1β Secretion in Macrophages

A hallmark of Entamoeba histolytica (Eh) invasion in the gut is acute intestinal inflammation dominated by secretion pro-inflammatory cytokines. Eh contact with macrophages activates caspase-1 recruitment NLRP3 inflammasome complex a Gal-lectin and cysteine proteases 5 (EhCP-A5)-dependent manner, resulting maturation interleukin (IL)-1β IL-18. Although it well-defined that activation important mediating IL-1β IL-18, role caspase-4 play overall response less identified. We have recently shown caspase-1/4 macrophage cleaved gasdermin D (GSDMD) leading to insertion N-terminal effector domain into cell membrane, driving both pore formation without causing significant death. In this study, we interrogated requirements mechanism for Eh-induced cleaving GSDMD bioactive release. Similar caspase-1, was dependent on EhCP-A5 binding triggered ATP release acted as second signal. To determine if interacted canonical regulate GSDMD-mediated cytokine release, CRISPR-Cas9 gene editing 4, ASC, THP-1 cells were utilized. Unlike within 10 min independent NLRP3, caspase-1. absence significantly up regulated enhanced cleavage induce robust secretion. played major triggering quantified HEK-Blue™ reporter assay pyroptosis. This marked contrast positive control, LPS + Nigericin induced high expression cleavage, secretion, massive These results suggest “hyperactivated macrophages” led pyroptosis disease pathogenesis.