Channelling inflammation: gasdermins in physiology and disease

Abstract

Gasdermins were recently identified as the mediators of pyroptosis - inflammatory cell death triggered by cytosolic sensing of invasive infection and danger signals. Upon activation, gasdermins form cell membrane pores, which release pro-inflammatory cytokines and alarmins and damage the integrity of the cell membrane. Roles for gasdermins in autoimmune and inflammatory diseases, infectious diseases, deafness and cancer are emerging, revealing potential novel therapeutic avenues. Here, we review current knowledge of the family of gasdermins, focusing on their mechanisms of action and roles in normal physiology and disease. Efforts to develop drugs to modulate gasdermin activity to reduce inflammation or activate more potent immune responses are highlighted.

Fig. 1. Key events in the history of gasdermins.

The purple boxes indicate milestone discoveries of gasdermin (GSDM) research.

Fig. 2. Gasdermins function as gatekeepers of pyroptosis.

In response to invasive pathogens, sterile danger signals or cytotoxic T cell attack, gasdermins (GSDMs) are activated by proteolytic cleavage, which releases the N-terminal (NT) fragment, which forms large cell membrane pores. The GSDM pore behaves as a gatekeeper for initiating downstream inflammatory cascades and pyroptotic cell death. Pyroptotic cells form large balloon-like membrane structures. Small intracellular molecules, including cytokines and cellular alarmins, are released through GSDM pores, causing inflammation. Some cells, termed ‘hyperactivated’, repair GSDM pores by shedding the damaged membrane and survive, but still induce inflammation by releasing IL-1 family cytokines.

Fig. 3. Molecular mechanisms that activate gasdermins.

In response to apoptotic signalling, gasdermin C (GSDMC) and GSDME are processed by caspase 8 and caspase 3, respectively, converting apoptosis into pyroptosis. Granzymes (Gzms) secreted by killer cells are delivered by perforin into target cells, where GzmA and GzmB can directly cleave and activate GSDMB and GSDME, respectively, to trigger pyroptotic cell death. In immune sentinel cells, both cytosolic canonical inflammasomes that respond to microbial infection or danger signals and the noncanonical inflammasome that responds directly to lipopolysaccharide (LPS) or endogenous oxidized phospholipids activate the inflammatory caspases (caspases 1, 4, 5 and 11), which cleave GSDMD and generate pore-forming N-terminal GSDMD (GSDMD-NT). TAK1 inhibition by the Yersinia effector protein YopJ or the small molecule 5z7 triggers caspase 8-dependent GSDMD cleavage and activation. GSDMD can also be directly processed and activated by neutrophil elastase (ELANE) and cathepsin G. In addition to protease-mediated release of active GSDM-NT, mutations in Gsdma3 lead to abolition of C-terminal GSDM inhibition and trigger GSMDA3 pore-forming activity. Diagram at bottom right indicates the proteases known to cleave and activate each of the gassdermins (yellow, caspases; purple, lymphocyte granzymes; blue, myeloid cell granule proteases). ALR, AIM-2 like receptor; CLR, C type lectin receptor; DAMP, damage-associated molecular pattern; NLR, NOD-like receptor; MOMP, mitochondrial outer membrane permeabilization; PAMP, pathogen-associated molecular pattern; TLR, Toll-like receptor.

Fig. 4. Mechanisms of gasdermin auto-inhibition, processing and pore formation.

a | Cartoon representation of gasdermin (GSDM) pore formation. Interaction between the N-terminal and C-terminal fragments of a GSDM (GSDM-NT, in purple and GSDM-CT, in yellow) auto-inhibits protein activation. Active caspase 1, a dimer of the p10 (in pale blue)–p20 (in silver) complex, recognizes two copies of GSDM through the GSDM-CT. This exosite recognition mechanism places the long interdomain linker (in grey) between GSDM-NT and GSDM-CT near the active site of caspase 1 (red balls). Active caspase 4 and caspase 11 use a similar mechanism to process GSDMD. Linker cleavage liberates GSDM-NT for pore formation at the membrane. The fully formed GSDM pore features a large cytosolic rim in addition to a transmembrane β-barrel. b | Structural illustration of GSDM at each stage in pore formation depicted in part a. In auto-inhibited GSDM (PDB: 5B5R for GSDMA3), GSDM-NT contacts GSDM-CT at two interfaces. Interface I is formed primarily by charged interactions of the α1 helix and the β1–β2 loop of GSDM-NT with the α5–α8–α12 helix cluster of GSDM-CT. Interface II is formed mainly by hydrophobic interactions between α4 of GSDM-NT and α9 and α11 of GSDM-CT. GSDMD recognition (PDB: 6KN0 and 6VIE) by caspase 1 is mediated by a substrate-binding exosite, comprised of two anti-parallel β-strands — βIII of caspase 1-p20 and βIII′ of caspase 1-p10. The exosite binds to α6 and α8 of GSDMD-CT through primarily hydrophobic interactions. The GSDMD NT–CT linker is positioned near the active site Cys285 of caspase 1 for cleavage. The GSDM pore contains roughly 27 subunits according to the structure of the GSDMA3 pore (PDB: 6CB8). With each subunit contributing four transmembrane β-strands, the membrane-inserted β-barrel contains 108 β-strands. The inner and outer diameters of the pore are approximately 180 Å and 280 Å, respectively. A soluble rim, rich in α-helices and loops, decorates the β-barrel on the cytosolic side. c | GSDM-NT conformations before (left) and after (right) pore formation. The positively charged lipid-binding helix α1 (in orange, like a thumb) is masked by GSDM-CT in the auto-inhibited conformation, and in the membrane-inserted conformation it interacts with lipids with negatively charged head groups found on the inner leaflet of the plasma membrane. Drastic conformational changes of GSDM-NT occur at extension domains 1 and 2 (ED1, in magenta and ED2, in red), which are located at the β3–β4–β5 and β7–α4–β8 regions in auto-inhibited GSDM-NT, respectively. Through pore formation, ED1 and ED2 become the two transmembrane β-hairpins (HP1, in magenta and HP2, in red, like four fingers), respectively.

Fig. 5. Role and function of gasdermins in human disease.

Autoactive gasdermin A (GSDMA) triggers pyroptosis in hair follicle cells and causes alopecia in mice but so far no human link to alopecia has been reported. GSDMB is implicated in autoimmune diseases such as childhood asthma and inflammatory bowel disease (IBD), but the underlying mechanism remains unclear. Granzyme A (GzmA) in killer lymphocytes activates GSDMB in target cells to cause pyroptosis. TNF can activate caspase 8 to trigger GSDMC-mediated pyroptosis in tumours. Upon sensing infection or danger, assembly of inflammasomes activates inflammatory caspases to cleave GSDMD and trigger pyroptosis, initiating immune responses. However, excessive pyroptosis leads to lethal sepsis, cytokine release syndrome (CRS) and severe inflammation and tissue damage. Autoactive pyrin or NLRP3 constitutively activate GSDMD and cause pyroptosis leading to familial Mediterranean fever (FMF) and neonatal-onset multisystem inflammatory disease (NOMID), respectively. Natural killer (NK) and CD8⁺ killer lymphocytes and chimeric antigen receptor (CAR) T cells activate pyroptosis in GSDME-expressing tumours when GzmB cleaves and activates GSDME and caspase 3. Alarmins released from pyroptotic tumour cells can also activate GSDMD-mediated pyroptosis in tumour-infiltrating macrophages and dendritic cells to enhance antigen presentation and functional activation of tumour-infiltrating lymphocytes (TILs). Secondary activation of macrophages can also mediate CRS during CAR T cell therapy. Chemotherapy drugs and radiotherapy activate caspase 3 in GSDME-expressing cells to convert apoptosis into pyroptosis, increasing their effectiveness at suppressing tumour growth by activating antitumour immunity; at the same time normal tissue cell damage and cytotoxicity is increased by activating GSDME-dependent pyroptosis especially in gut epithelial cells and haematopoietic precursor cells. Autoactive GSDME induces cochlear hair cell pyroptosis, leading to nonsyndromic hearing impairment. Loss-of-function mutation of DFNB59 also causes recessive nonsyndromic hearing loss. CAPS, cryopyrin-associated periodic syndromes; MS, multiple sclerosis; SS, systemic sclerosis.

Fig. 6. Strategies to treat gasdermin-associated diseases.

a | Steps in gasdermin (GSDM) activation that can be targeted, and the mechanisms used by existing GSDMD inhibitors^,. Other yet unused potential mechanisms are shown with a question mark. b | Cellular delivery of GSDM for cancer cell killing and activation of antitumour immunity. See text for details. CT, C-terminal; NT, N-terminal.