Person:

Xia, Shiyu

Inflammasomes are innate immune organelles that activate inflammatory caspases, which proteolytically process gasdermin D (GSDMD). In addition, caspase-1, one of the inflammatory caspases, cleaves inactive interleukin (IL)-1 family precursors to generate mature cytokines such as IL-1β and IL-18. Cleaved GSDMD forms transmembrane pores to allow IL-1 release and drive lytic cell death known as pyroptosis. Here we report cryo-electron microscopy structures of the GSDMD pore and prepore, which reveal the different conformational states and extensive membrane-binding elements including a hydrophobic anchor and positively charged patches. The GSDMD pore conduit is predominantly negatively charged. By contrast, IL-1 precursors possess an acidic domain that is proteolytically removed by caspase-1. Unlysed liposomes permeabilized by GSDMD pores release positively charged and neutral cargoes faster than negatively charged ones of similar sizes, and favourably allow the passage of IL-1β and IL-18 in comparison to their precursors. Concordantly, GSDMD-perforated living but not pyroptotic macrophages preferentially release mature IL-1β. Mutations of acidic residues in GSDMD compromise this preference, hindering intracellular retention of the precursor and secretion of the mature cytokine. Therefore, the GSDMD pore mediates IL-1 release via electrostatic filtering, which suggests the importance of charge in addition to size in cargo transport across this large channel.