Caspase 3 proteolytically inactivates gasdermin Ec and inhibits pyroptosis in an early vertebrate

• A new gasdermin E homolog was identified in swordtail fish. • GSDMEc executes pyroptosis via N-terminal domain. • Caspase 3 destructively cleaves GSDMEc and inhibits pyroptosis. Gasdermin represents a family of pore-forming proteins that execute programmed necrotic cell death, pyroptosis. As an ancient family member, gasdermin E (GSDME) exists ubiquitously from fish to mammals. Recently, a new GSDME homolog, designated GSDMEc, was identified exclusively in fish. However, the function and regulation of GSDMEc remain elusive. In the present study, we identified a GSDMEc ortholog in swordtail fish, which exhibits canonical gasdermin two-domain architecture. Similar to its pyroptosis-inducing homologs, swordtail fish GSDMEc could execute pyroptosis via conserved N-terminal domain, whereas the C-terminal domain exerts auto-inhibitory effect on N-terminal domain to maintain the full-length protein non-cytotoxicity. We further explored the proteolytic cleavage of GSDMEc, and found that caspase 3 specifically cleaved GSDMEc at the site within the N-terminal domain, and generated a truncated N-terminal fragment. In accordance with this observation, co-expression of caspase 3 and GSDMEc did not result in pyroptosis, while co-expression of caspase 3 impeded pyroptosis induced by the N-terminal domain of GSDMEc. Collectively, our study reveals the negative regulation of caspase 3/GSDMEc-mediated pyroptosis in swordtail fish, which provides insight into gasdermin functionality and pyroptosis regulation in early vertebrates.