Mus musculus breed:R26-Cas9 Knock In
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PROVIDER: PRJNA739880 | ENA |
REPOSITORIES: ENA
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Project description:Control of gasdermin D oligomerization and pyroptosis by the Ragulator-Rag-mTORC1 pathway
| PRJNA742124 | ENA
Project description:The process of pyroptosis is mediated by inflammasomes and a downstream effector known as gasdermin D (GSDMD). Upon cleavage by inflammasome-associated caspases, the N-terminal domain of GSDMD forms membrane pores that promote cytolysis. Numerous proteins promote GSDMD cleavage, but none are known to be required for pore formation after GSDMD cleavage. Herein, we report a forward genetic screen that identified the Ragulator-Rag complex as being necessary for GSDMD pore formation and pyroptosis in macrophages. Mechanistic analysis revealed that Ragulator-Rag is not required for GSDMD cleavage upon inflammasome activation, but rather promotes GSDMD oligomerization in the plasma membrane. Defects in GSDMD oligomerization and pore formation can be rescued by mitochondrial poisons that stimulate reactive oxygen species (ROS) production, and ROS modulation impacts the ability of inflammasome pathways to promote pore formation downstream of GSDMD cleavage. These findings reveal an unexpected link between key regulators of immunity (inflammasome-GSDMD) and metabolism (Ragulator-Rag).
2021-06-29 | GSE179089 | GEO
Project description:Pyroptosis by Caspase11/4-Gasdermin-D Pathway in Alcoholic Hepatitis
| PRJNA381041 | ENA
Project description:Gasdermin D (GSDMD) is the executioner of pyroptosis, which is important for host defense against pathogen infection. After activation, caspase-mediated cleavage of GSDMD liberates an N-terminal fragment (GSDMD-NT), which oligomerizes and forms pores in the plasma membrane, leading to cell death and subsequent release of proinflammatory cytokines. How this process is spatiotemporally controlled to promote pyroptosis in cells has been a fundamental, unaddressed question. Here, we identify GSDMD as a substrate for reversible S-palmitoylation on cysteine 192 (Cys192) in response to lipopolysaccharide (LPS) stimulation. We found that the palmitoyl acyltransferase DHHC7palmitoylates GSDMD to direct its cleavage by caspases in pyroptosis by promoting the interaction of GSDMD and caspases. We further show that after GSDMD cleavage, palmitoylation of GSDMD-NTpromotes its plasma membrane translocation and binding, and then acyl protein thioesterase 2 (APT2) depalmitoylates GSDMD-NT to unmask the Cys192 residue to promote oxidation-mediated oligomerization and pyroptosis. Perturbation of either palmitoylation or depalmitoylation suppresses pyroptosis, extends the survival of mice from LPS-induced lethal septic shock and sensitizes mice to bacterial infection. Thus. our findings reveal a model through which a palmitoylation-depalmitoylationrelay spatially and temporally controls GSDMD activation in pyroptosis.
2024-02-25 | GSE252911 | GEO
Project description:Gasdermin D (GSDMD) is the executioner of pyroptosis, which is important for host defense against pathogen infection. After activation, caspase-mediated cleavage of GSDMD liberates an N-terminal fragment (GSDMD-NT), which oligomerizes and forms pores in the plasma membrane, leading to cell death and subsequent release of proinflammatory cytokines. How this process is spatiotemporally controlled to promote pyroptosis in cells has been a fundamental, unaddressed question. Here, we identify GSDMD as a substrate for reversible S-palmitoylation on cysteine 192 (Cys192) in response to lipopolysaccharide (LPS) stimulation. We found that the palmitoyl acyltransferase DHHC7palmitoylates GSDMD to direct its cleavage by caspases in pyroptosis by promoting the interaction of GSDMD and caspases. We further show that after GSDMD cleavage, palmitoylation of GSDMD-NTpromotes its plasma membrane translocation and binding, and then acyl protein thioesterase 2 (APT2) depalmitoylates GSDMD-NT to unmask the Cys192 residue to promote oxidation-mediated oligomerization and pyroptosis. Perturbation of either palmitoylation or depalmitoylation suppresses pyroptosis, extends the survival of mice from LPS-induced lethal septic shock and sensitizes mice to bacterial infection. Thus. our findings reveal a model through which a palmitoylation-depalmitoylationrelay spatially and temporally controls GSDMD activation in pyroptosis.
2024-02-25 | GSE252893 | GEO
Project description:The Rag GTPases recruit the master kinase mTORC1 to lysosomes to regulate cell growth and proliferation in response to amino acid availability. The nucleotide state of Rag heterodimers is critical for their association with mTORC1. Our cryo-EM structure of RagA/RagC in complex with mTORC1 shows the details of RagA/C binding to the RAPTOR subunit of mTORC1 and explains why only the RagAGTP/RagCGDP nucleotide state binds mTORC1. Previous kinetic studies suggested that GTP binding to one Rag locks the heterodimer to prevent GTP binding to the other. Our crystal structures and dynamics show the mechanism for this locking, and explain how oncogenic hotspot mutations disrupt this process. In contrast to allosteric activation by RHEB, Rag heterodimer binding does not change mTORC1 conformation and activates mTORC1 by targeting it to lysosomes.
2019-10-11 | PXD015778 | Pride
Project description:This submission is associated with a paper by Hesketh et al. that identifies the GATOR-Rag GTPase pathway as a negative regulator of mTORC1 activation by lysosome-derived amino acids.
2019-07-30 | MSV000084159 | MassIVE
Project description:Immune checkpoint blockade (ICB) therapy has demonstrated promising clinical results in oncology and is currently one of the most sought-after immunotherapies for tumors. However, there are a number of issues with ICB therapy, including low response rates and a lack of effective efficacy predictors. Gasdermin-mediated pyroptosis is a typical form of inflammatory death mode, but it is unclear whether ICB treatment can trigger its occurrence or whether gasdermin expression has any effect on the response rate to ICB treatment. Herein, we discovered that high expression of gasdermin protein was associated with a favorable tumor immune microenvironment and a favorable prognosis in head and neck squamous cell carcinoma (HNSCC), and taking advantage of the mouse HNSCC cell line 4MOSC1 (responsive to CTLA-4 blockade) and 4MOSC2 (resistant to CTLA-4 blockade) orthotopic models, we also demonstrated that CTLA-4 blockade treatment induced gasdermin-mediated pyroptosis of tumor cells and that the expression of gasdermin was positively associated with the effectiveness of CTLA-4 blocking treatment. Mechanistically, through RNA sequencing, flow cytometry, ELISA, Western blotting and RT-PCR, we discovered that CTLA-4 blockade activated CD8+ T cells in the tumor and increased the levels of the cytokines TNF-α and IFN-γ in the tumor microenvironment, and that TNF-α and IFN-γ synergistically activated the STAT1/IRF1 axis, inducing tumor cell pyroptosis and the release of large amounts of inflammatory substances and chemokines such as CXCL10.
2023-04-01 | GSE201367 | GEO
Project description:The tumor immune microenvironment is complex in composition, function and dynamic, influencing the evolution of tumor, prognosis and response to immunotherapy. Recent studies have significantly advanced our knowledge of the roles of Gasdermin protein-mediated tumoral pyroptosis activated by caspases and granzymes from cytotoxic lymphocytes in facilitating the killing of tumor cells. However, the pyroptosis of immune cells in tumor microenvironment and their affecting the reprogram of the tumor microenvironment remains poorly understood. In this study, we find that Gasdermin D (GSDMD), among Gasdermin family proteins, is significantly positively correlated with the immune checkpoint signature, and inversely linked to the overall survival of cancer patients by the analysis with TCGA clinical data. Using conditional knockout of GSDMD together with immunofluorescence co-labeling and single-cell RNA sequencing (scRNA-seq), we demonstrate that GSDMD mainly on antigen-presenting cells (APCs) such as macrophages and dendritic cells (DCs) in tumor microenvironment, restrains anti-tumor immunity under the treatment of PD-L1 antibody. Loss of GSDMD in APCs enhanced Type I interferon-stimulated gene (ISG) program of such cells, which promotes antigen presentation of macrophages and DCs, and facilitates the production of CD8+ effector and functional T cell. We further demonstrate that GSDMD deficiency increases anti-tumor immunity in a cyclic GMP–AMP synthase (cGAS)-dependent manner. Moreover, pharmacological inhibition of GSDMD-mediated pyroptosis significantly improves anti-tumor immunity in combination with PD-L1 antibody immunotherapy. Together, our findings reveal an important role for GSDMD-mediated pyroptosis of APCs such as macrophages and DCs in regulating CD8+ T cell function and underscore the potential of GSDMD blockade in promoting anti-tumor immunity, which may help the development of cancer immunotherapy.
2022-10-05 | GSE198550 | GEO
Project description:Acidic tumor microenvironment (TME)-evoked MRC nanoparticles (MRC NPs) co-delivering immune agonist RGX-104 and photosensitizer chlorine e6 (Ce6) are reported for pyroptosis-mediated immunotherapy. RGX-104 remodels TME by transcriptional activation of ApoE to regress myeloid-derived suppressor cells’ (MDSCs) activity, which neatly creates foreshadowing for intensifying pyroptosis. Considering Ce6-triggered photodynamic therapy (PDT) can strengthen oxidative stress and organelles destruction to increase immunogenicity, immunomodulatory-photodynamic MRC nanodrugs will implement an aforementioned two-pronged strategy to enhance gasdermin E (GSDME)-dependent pyroptosis. RNA-seq analysis of MRC at the cellular level is introduced to first elucidate the intimate relationship between RGX-104 acting on LXR/ApoE axis and pyroptosis, where RGX-104 provides the prerequisite for pyroptosis participating in antitumor therapy. Briefly, MRC with favorable biocompatibility tackles the obstacle of hydrophobic drugs delivery, and becomes a powerful pyroptosis inducer to reinforce immune efficacy. MRC-elicited pyroptosis in combination with anti-PD-1 blockade therapy boosts immune response in solid tumors, successfully arresting invasive metastasis and extending survival based on remarkable antitumor immunity. MRC may initiate a new window for immuno-photo pyroptosis stimulators augmenting pyroptosis-based immunotherapy. Owing to the transcriptome sequencing on 4T1 cells exposed to various treatment, pyroptosis mechanism of MRC was thoroughly analyzed via comparing the expression discrepancy among PBS, MRC, and MRC+L groups.
2023-08-13 | GSE240328 | GEO