Succination inactivates gasdermin D and blocks pyroptosis.
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ABSTRACT:
SUBMITTER: Humphries F
PROVIDER: S-EPMC8744141 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Pyroptosis is the process of inflammatory cell death. The primary function of pyroptosis is to induce strong inflammatory responses that defend the host against microbe infection. Excessive pyroptosis, however, leads to several inflammatory diseases, including sepsis and autoimmune disorders. Pyroptosis can be canonical or noncanonical. Upon microbe infection, the canonical pathway responds to pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs), while the noncanonical pathway responds to intracellular lipopolysaccharides (LPS) of Gram-negative bacteria. The last step of pyroptosis requires the cleavage of gasdermin D (GsdmD) at D275 (numbering after human GSDMD) into N- and C-termini by caspase 1 in the canonical pathway and caspase 4/5/11 (caspase 4/5 in humans, caspase 11 in mice) in the noncanonical pathway. Upon cleavage, the N-terminus of GsdmD (GsdmD-N) forms a transmembrane pore that releases cytokines such as IL-1β and IL-18 and disturbs the regulation of ions and water, eventually resulting in strong inflammation and cell death. Since GsdmD is the effector of pyroptosis, promising inhibitors of GsdmD have been developed for inflammatory diseases. This review will focus on the roles of GsdmD during pyroptosis and in diseases.
| S-EPMC8463274 | biostudies-literature
Project description:Human Gasdermin D (GSDMD) is a key mediator of pyroptosis, a pro-inflammatory form of cell death occurring downstream of inflammasome activation as part of the innate immune defence. Upon cleavage by inflammatory caspases in the cytosol, the N-terminal domain of GSDMD forms pores in the plasma membrane resulting in cytokine release and eventually cell death. Targeting GSDMD is an attractive way to dampen inflammation. In this study, six GSDMD targeting nanobodies are characterized in terms of their binding affinity, stability, and effect on GSDMD pore formation. Three of the nanobodies inhibit GSDMD pore formation in a liposome leakage assay, although caspase cleavage was not perturbed. We determine the crystal structure of human GSDMD in complex with two nanobodies at 1.9 Å resolution, providing detailed insights into the GSDMD-nanobody interactions and epitope binding. The pore formation is sterically blocked by one of the nanobodies that binds to the oligomerization interface of the N-terminal domain in the multi-subunit pore assembly. Our biochemical and structural findings provide tools for studying inflammasome biology and build a framework for the design of GSDMD targeting drugs.
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Project description:Gasdermin D (GSDMD)-mediated pyroptosis and downstream inflammation are important self-protection mechanisms against stimuli and infections. Hosts can defend against intracellular bacterial infections by inducing cell pyroptosis, which triggers the clearance of pathogens. However, pyroptosis is a double-edged sword. Numerous studies have revealed the relationship between abnormal GSDMD activation and various inflammatory diseases, including sepsis, coronavirus disease 2019 (COVID-19), neurodegenerative diseases, nonalcoholic steatohepatitis (NASH), inflammatory bowel disease (IBD), and malignant tumors. GSDMD, a key pyroptosis-executing protein, is linked to inflammatory signal transduction, activation of various inflammasomes, and the release of downstream inflammatory cytokines. Thus, inhibiting GSDMD activation is considered an effective strategy for treating related inflammatory diseases. The study of the mechanism of GSDMD activation, the formation of GSDMD membrane pores, and the regulatory strategy of GSDMD-mediated pyroptosis is currently a hot topic. Moreover, studies of the structure of caspase-GSDMD complexes and more in-depth molecular mechanisms provide multiple strategies for the development of GSDMD inhibitors. This review will mainly discuss the structures of GSDMD and GSDMD pores, activation pathways, GSDMD-mediated diseases, and the development of GSDMD inhibitors.
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Project description:SARS-CoV-2 is an emerging coronavirus that causes dysfunctions in multiple human cells and tissues. Studies have looked at the entry of SARS-CoV-2 into host cells mediated by the viral spike protein and human receptor ACE2. However, less is known about the cellular immune responses triggered by SARS-CoV-2 viral proteins. Here, we show that the nucleocapsid of SARS-CoV-2 inhibits host pyroptosis by blocking Gasdermin D (GSDMD) cleavage. SARS-CoV-2-infected monocytes show enhanced cellular interleukin 1b (IL-1b) expression, but reduced IL-1b secretion. While SARS-CoV-2 infection promotes activation of the NLRP3 inflammasome and caspase-1, GSDMD cleavage and pyroptosis are inhibited in infected human monocytes. SARS-CoV-2 nucleocapsid protein associates with GSDMD in cells and inhibits GSDMD cleavage in vitro and in vivo. The nucleocapsid binds the GSDMD linker region and hinders GSDMD processing by caspase-1. These insights into how SARS-CoV-2 antagonizes cellular inflammatory responses may open new avenues for treating COVID-19 in the future.
| S-SCDT-EMBOJ-2021-108249 | biostudies-other
Project description:Bacillus cereus is well known as a causative agent of foodborne gastrointestinal diseases and systemic non-gastrointestinal diseases. We have recently identified a pathogenic B. cereus (named H2) from a deep-sea cold-seep. H2 possesses the pyroptosis-inducing capacity and contains a number of enterotoxins including cytotoxin K (CytK). In the present work, we examined the cytotoxicity of the CytK of H2 to human macrophages. CytK bound macrophages by interaction with the plasma membrane and caused cellular structure damage. CytK-cell interaction triggered rapid pyroptosis mediated by caspase 1-activated gasdermin D (GSDMD). CytK-induced pyroptosis required NLRP3 inflammasome activation, K+ efflux, and intracellular Ca2+ accumulation. CytK exhibited apparent binding to several cytomembrane lipids, in particular phosphatidic acid, which proved to be essential to CytK-elicited cell death. Together, these results add new insights into the cytotoxic mechanism of CytK.
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Project description:BackgroundThe frequent emergence of drug resistance to chemotherapy is a major obstacle for the treatment of ovarian cancer. There is a need for novel drugs to fulfill this challenge. Pyroptosis-inducing drugs can inhibit tumor growth. However, their roles in ovarian cancer have not been demonstrated.MethodsWe tested the effectiveness of a novel drug, BI 2536, which we found in colorectal cancer. Cell proliferation, cell cycle, and drug-induced apoptosis and pyroptosis were tested. In vivo treatments were performed using a cell-derived xenograft model.ResultsBI 2536 significantly inhibited the proliferation of ovarian cancer cells and induced cell cycle arrest at the G2/M phases. After BI 2536 treatment, DNA fragmentation and PS exposure on the outside of apoptotic cells were detected. Moreover, the pyroptotic phenotype of ovarian cancer cells along with the release of LDH and HMGB1 were observed, indicating the leakage of cells. Western blot analysis verified that BI 2536 induced GSDME-mediated pyroptosis. Pyroptosis was abolished after additional treatment with Z-DEVD-FMK, a caspase-3 inhibitor. Thus, BI 2536 induced pyroptosis in ovarian cancer through the caspase-3/GSDME pathway. In vivo experiments further demonstrated the antitumoral effect and ability of BI 2536 to accumulate CD8+ T cells in ovarian cancer.ConclusionIn this study, we identified BI 2536 as an effective anti-ovarian cancer drug that inhibits proliferation, arrests the cell cycle, induces apoptosis and pyroptosis, and leads to the accumulation of CD8+ T cells in tumor sites. Drug-induced pyroptosis may have promising prospects for reducing side effects and activating immune responses.
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Project description:Inflammatory caspases (caspases 1, 4, 5 and 11) are activated in response to microbial infection and danger signals. When activated, they cleave mouse and human gasdermin D (GSDMD) after Asp276 and Asp275, respectively, to generate an N-terminal cleavage product (GSDMD-NT) that triggers inflammatory death (pyroptosis) and release of inflammatory cytokines such as interleukin-1β. Cleavage removes the C-terminal fragment (GSDMD-CT), which is thought to fold back on GSDMD-NT to inhibit its activation. However, how GSDMD-NT causes cell death is unknown. Here we show that GSDMD-NT oligomerizes in membranes to form pores that are visible by electron microscopy. GSDMD-NT binds to phosphatidylinositol phosphates and phosphatidylserine (restricted to the cell membrane inner leaflet) and cardiolipin (present in the inner and outer leaflets of bacterial membranes). Mutation of four evolutionarily conserved basic residues blocks GSDMD-NT oligomerization, membrane binding, pore formation and pyroptosis. Because of its lipid-binding preferences, GSDMD-NT kills from within the cell, but does not harm neighbouring mammalian cells when it is released during pyroptosis. GSDMD-NT also kills cell-free bacteria in vitro and may have a direct bactericidal effect within the cytosol of host cells, but the importance of direct bacterial killing in controlling in vivo infection remains to be determined.
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Project description:Alcoholic hepatitis (AH) continues to be a disease with high mortality and no efficacious medical treatment. Although severe AH is presented as acute on chronic liver failure, what underlies this transition from chronic alcoholic steatohepatitis (ASH) to AH, is largely unknown. To address this question, unbiased RNA-seq and proteomic analyses were performed on livers of the recently developed AH mouse model which exhibits the shift to AH from chronic ASH upon weekly alcohol binge, and these results are compared with gene expression profiling data from AH patients. This cross-analysis has identified Casp11 (CASP4 in man) as a commonly upregulated gene known to be involved in non-canonical inflammasome pathway. Immunoblotting confirms CASP11/4 activation in AH mice but not in chronic ASH. Gasdermin-D (GSDMD) which induces pyroptosis (lytic cell death caused by bacterial infection) downstream of CASP11/4 activation, is also activated in AH livers. CASP11 deficiency reduces GSDMD activation, bacterial load in the liver, and the severity of AH. Conversely, the deficiency of IL-18, the key anti-microbial cytokine, aggravates hepatic bacterial load, GSDMD activation, and AH. Further, hepatocyte-specific expression of constitutively active GSDMD worsens hepatocellular lytic death and PMN inflammation. These results implicate pyroptosis induced by CASP11/4-GSDMD pathway in the pathogenesis of AH.
2017-03-31 | GSE97234 | GEO
Project description:Sepsis is a life-threatening condition caused by pathogen infection and associated with pyroptosis. Pyroptosis occurs upon activation of proinflammatory caspases and their subsequent cleavage of gasdermin D (GSDMD), resulting in GSDMD N-terminal fragments that form membrane pores to induce cell lysis. Here, we show that antioxidant defense enzyme glutathione peroxidase 4 (GPX4) and its ability to decrease lipid peroxidation, negatively regulate macrophage pyroptosis, and septic lethality in mice. Conditional Gpx4 knockout in myeloid lineage cells increases lipid peroxidation-dependent caspase-11 activation and GSDMD cleavage. The resultant N-terminal GSDMD fragments then trigger macrophage pyroptotic cell death in a phospholipase C gamma 1 (PLCG1)-dependent fashion. Administration of the antioxidant vitamin E that reduces lipid peroxidation, chemical inhibition of PLCG1, or genetic Caspase-11 deletion or Gsdmd inactivation prevents polymicrobial sepsis in Gpx4-/- mice. Collectively, this study suggests that lipid peroxidation drives GSDMD-mediated pyroptosis and hence constitutes a potential therapeutic target for lethal infection.
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Project description:The syncytiotrophoblast is a human epithelial cell that is bathed in maternal blood on the maternal-facing surface of the human placenta. It therefore acts as a barrier and exchange interface between the mother and fetus. Syncytiotrophoblast dysfunction is a feature of pregnancy pathologies, like preeclampsia. Dysfunctional syncytiotrophoblasts display a loss of microvilli, which is a marker of aberrant apical-basal polarization, but little data exist about the regulation of syncytiotrophoblast polarity. Atypical PKC isoforms are conserved polarity regulators. Thus, we hypothesized that aPKC isoforms regulate syncytiotrophoblast polarity. Using human placental explant culture and primary trophoblasts, we found that loss of aPKC activity or expression induces syncytiotrophoblast gasdermin-E-dependent pyroptosis, a form of programmed necrosis. We also establish that TNF-α induces an isoform-specific decrease in aPKC expression and gasdermin-E-dependent pyroptosis. Therefore, aPKCs are homeostatic regulators of the syncytiotrophoblast function and a pathogenically relevant pro-inflammatory cytokine leads to the induction of programmed necrosis at the maternal-fetal interface. Hence, our results have important implications for the pathobiology of placental disorders like preeclampsia.
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