Sublytic gasdermin-D pores captured in atomistic molecular simulations

Gasdermin-D (GSDMD) is the ultimate effector of pyroptosis, a form of programmed cell death associated with pathogen invasion and inflammation. After proteolytic cleavage by caspases, the GSDMD N-terminal domain (GSDMD^NT) assembles on the inner leaflet of the plasma membrane and induces the formation of membrane pores. We use atomistic molecular dynamics simulations to study GSDMD^NT monomers, oligomers, and rings in an asymmetric plasma membrane mimetic. We identify distinct interaction motifs of GSDMD^NT with phosphatidylinositol-4,5-bisphosphate (PI(4,5)P₂) and phosphatidylserine (PS) headgroups and describe their conformational dependence. Oligomers are stabilized by shared lipid binding sites between neighboring monomers acting akin to double-sided tape. We show that already small GSDMD^NT oligomers support stable, water-filled, and ion-conducting membrane pores bounded by curled beta-sheets. In large-scale simulations, we resolve the process of pore formation from GSDMD^NT arcs and lipid efflux from partial rings. We find that high-order GSDMD^NT oligomers can crack under the line tension of 86 pN created by an open membrane edge to form the slit pores or closed GSDMD^NT rings seen in atomic force microscopy experiments. Our simulations provide a detailed view of key steps in GSDMD^NT-induced plasma membrane pore formation, including sublytic pores that explain nonselective ion flux during early pyroptosis.