Review
Biological mechanisms and therapeutic relevance of the gasdermin family

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Abstract

Innate immunity enables host defense against pathogens and endogenous danger through inflammasomes, which are supramolecular complexes that recognize the threats and activate the immune response. Inflammasome activation often leads to pyroptosis, a highly inflammatory and lytic form of cell death, as a means of killing infected cells and releasing IL-1 family cytokines that communicate with other cells. Dysregulated inflammasome signaling results in a wide range of immune disorders including gout, sepsis, and hepatitis. Discovered as a direct killer molecule in pyroptosis, gasdermin D (GSDMD) is a pore-forming protein that represents a novel family with diverse cellular functions and pathological roles. This review summarizes current opinions in the biological mechanisms and therapeutic values of the GSDM family, particularly of GSDMD. Detailed mechanisms of auto-inhibition and pore formation by the GSDM family are presented, followed by a brief summary of the progress in the development of GSDM-targeting therapeutics.

Introduction

This review first gives a brief account of the discovery of gasdermin D (GSDMD), which represents a novel pore-forming protein family known as gasdermins (GSDMs). The GSDM family is predominantly expressed in the GI tract, skin, and immune cells and implicated in many diseases including inflammatory disorders and cancer (Table 1). The GSDM family was mentioned in literature two decades ago as a gene family associated with alopecia in mice and hearing loss in humans (Saeki et al., 2000a). The name “gasdermin” came from expression profiling of GSDMA in mice which showed predominant localization in the gastrointestinal (GI) tract and the dermis. In humans, the family contains six members, GSDMA through GSDMF. In mice, GSDMB is absent, and GSDMA and GSDMC have multiple isoforms, which are GSDMA1-3 and GSDMC1-4, respectively. As of now, the activating enzymes and biological functions have been elucidated for GSDMD, GSDME, and GSDMB. In this review, the functions, pore formation mechanism, and therapeutic potentials of GSDMs will be covered with an emphasis on insights from GSDM structures.

To understand GSDMs entails knowledge about inflammasomes, which are cytosolic protein complexes that function as signaling organelles in innate immunity (Lamkanfi and Dixit, 2014). In response to pathogen- and damage-associated molecular patterns (PAMPs and DAMPs), sensors such as nucleotide-binding domain and leucine-rich repeat-containing proteins (NLRs) and AIM2-like receptors (ALRs) assemble into multimeric protein platforms (Hu et al., 2015; Tenthorey et al., 2017; Zhang et al., 2015), which in turn recruit adaptors such as apoptosis-associated, speck-like protein containing a caspase recruitment domain (ASC) (Lu et al., 2014) and effectors such as caspase-1 through homotypic death domain interactions (Shen et al., 2019). Caspase-1 promotes the proteolytic maturation of IL-1 family cytokines including IL-1β and IL-18. The NLR/ALR-containing sensor-adaptor-effector systems are often referred to as canonical inflammasomes, with the most studied example being the NLRP3 inflammasome. By contrast, the presence of oxidized lipids or exogeneous lipids such as lipopolysaccharides (LPS) from Gram-negative bacteria in the cytosol directly activates caspase-11 (mouse homolog of human caspase-4/5), which represents the non-canonical inflammasome (Kayagaki et al., 2013; Zanoni et al., 2016). Caspase-1 in the canonical inflammasomes and caspase-4, -5, and -11 in the noncanonical inflammasome are cysteine proteases collectively known as inflammatory caspases. The recruitment of these caspases to inflammasomes facilitates their oligomerization, higher-order assembly, and proximity-induced auto-activation (Wu, 2013). Direct downstream effects of active inflammatory caspases include IL-1 release and pyroptosis, a highly inflammatory and lytic form of programmed cell death. Yet, while these phenomena have long been observed, the direct executioner of IL-1 release and pyroptosis remained a mystery until very recently.

Section snippets

GSDMD, a direct executioner of pyroptosis

GSDMD, the first extensively studied member of the GSDM family, was discovered in the context of inflammasome biology and first functionally linked to pyroptosis. In 2015, an N-ethyl-N-nitrosourea (ENU)-based screen in mice identified mutations in the Gsdmd gene that impaired IL-1β secretion from macrophages following LPS challenge (Kayagaki et al., 2015). In line with this discovery, a CRISPR-Cas9-based screen in murine bone-marrow derived macrophages (BMDMs) identified GSDMD as involved in

Auto-inhibition of GSDM-NT by GSDM-CT

GSDMs are neither strictly membrane proteins nor soluble proteins, and the solubility and localization of GSDMs are dependent on whether the proteins are proteolytically activated. While full-length GSDMs are auto-inhibited andsoluble, cleaved GSDMs bind lipids in the membranes to form pores and possess cytotoxic effects.

Prior to proteolytic activation, the GSDM family except for GSDMF feature a two-domain organization, with the functional, pore-forming fragment GSDM-NT kept inactive by the

High-resolution snapshot of an active-state GSDM

While structures of full-length GSDMs provided valuable visualization of the auto-inhibition architecture of the family, the exact mechanism of GSDM pore formation requires the structural determination of GSDMs in their active states. However, even in the golden age of structural biology thanks to cryo-electron microscopy (cryo-EM), to solve the high-resolution structure of a GSDM pore is not an easy task. In crystallizing GSDMD, the loop-rich and flexible nature of GSDMD-NT necessitated

Pharmacological interventions

Physiological inflammation may recruit phagocytes cells to the site of infection for host defense, or directly kill compromised host cells in the case of pyroptosis. However, dysregulated inflammation can lead to severe pathologies, including inflammatory disorders such as gout, inflammatory bowel disease, sepsis, and alcoholic and non-alcoholic hepatitis (Khanova et al., 2018; Lieberman et al., 2019; So and Martinon, 2017; Xu et al., 2018) (Table 1). Precise control of the degree of

Conclusions and future prospects

A rich body of research has begun over the past few years on the pyroptotic role of GSDMD, the first functionally elucidated member of the GSDM family. Adding to these insights are discoveries of the lysis-independent, non-pyroptotic functions of GSDMD, other regulators of GSDMD activation, and mechanisms for downregulating GSDMD developed by both host cells and pathogens. Clearly, there is still a long way ahead to elucidate the biological functions of the GSDM family, which extend far beyond

Funding

This work was supported by NIH/NIAID grant 1R01AI139914 to Hao Wu at Harvard Medical School and Boston Children's Hospital.

Declaration of competing interest

The author confirms that there are no conflicts of interest.

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      After inflammasome activation, caspase-1 cleaves the precursor of IL-1β and IL-18 and converts them into matured forms [14–16]. At the same time, proinflammatory caspases cleave gasdermins (GSMDs), a key family of pore-forming proteins that participate in the activation of pyroptosis [18–20]. The cleaved N-terminal domain of GSMDs is cytotoxic to cells and translocates to the cell membrane, where it interacts with membrane lipids and assembles pores with various subunits that induce pyroptotic cell death [18,19].

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