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) 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 monomers, oligomers, and rings in an asymmetric plasma membrane mimetic. We identify distinct interaction motifs of GSDMD with phosphatidylinositol-4,5-bisphosphate (PI(4,5)P) 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 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 arcs and lipid efflux from partial rings. We find that high-order GSDMD oligomers can crack under the line tension of 86 pN created by an open membrane edge to form the slit pores or closed GSDMD rings seen in atomic force microscopy experiments. Our simulations provide a detailed view of key steps in GSDMD-induced plasma membrane pore formation, including sublytic pores that explain nonselective ion flux during early pyroptosis.