{"gene":"GSDME","run_date":"2026-06-10T01:55:21","timeline":{"discoveries":[{"year":2017,"finding":"Caspase-3 cleaves GSDME (DFNA5) after Asp270 to generate a necrotic N-terminal fragment (GSDME-N) that targets the plasma membrane to induce secondary necrosis/pyroptosis. Cells expressing GSDME progress to secondary necrosis upon apoptotic stimulation, while GSDME-deleted cells disassemble into apoptotic bodies instead.","method":"Cell-based cleavage assays, GSDME deletion/knockout, apoptotic stimulation (etoposide, VSV infection), plasma membrane targeting assays","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — direct identification of cleavage site by mutagenesis, loss-of-function phenotype, replicated across multiple labs subsequently","pmids":["28045099"],"is_preprint":false},{"year":2004,"finding":"Mutant DFNA5 (exon 8 skipped) transfected into HEK293T and COS-1 cells causes approximately doubled post-transfection cell death attributable to necrotic (not apoptotic) events, supporting a gain-of-function mechanism for DFNA5-associated hearing loss.","method":"Transfection of GFP-tagged wild-type vs. mutant DFNA5 in mammalian cell lines, flow cytometry, fluorescence microscopy","journal":"Journal of medical genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct functional assay in mammalian cells with two readouts (flow cytometry + microscopy), single lab","pmids":["15173223"],"is_preprint":false},{"year":2011,"finding":"GSDME is composed of two domains separated by a hinge region: the N-terminal domain induces apoptosis when transfected in HEK293T cells, while the C-terminal domain masks and regulates this apoptosis-inducing capability. Knockout mice microarray analysis further supported involvement of GSDME in apoptosis-related pathways.","method":"Domain transfection in HEK293T cells, gene expression microarray using Dfna5 knockout mice","journal":"European journal of human genetics : EJHG","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional domain dissection with two orthogonal approaches (transfection + knockout microarray), single lab","pmids":["21522185"],"is_preprint":false},{"year":2006,"finding":"DFNA5 gene expression is strongly induced by p53 (both exogenous and endogenous). Chromatin immunoprecipitation identified a p53-binding sequence in intron 1 of DFNA5, and a reporter assay confirmed p53-dependent transcriptional activity at this site. Ectopic DFNA5 enhanced etoposide-induced cell death in a p53-dependent manner.","method":"Chromatin immunoprecipitation (ChIP), reporter gene assay, p53 overexpression/knockout systems, gamma-ray irradiation in p53+/+ vs p53-/- mice","journal":"Journal of human genetics","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — ChIP directly identified p53 binding site, reporter assay confirmed transcriptional activity, in vivo validation in p53 knockout mice, multiple orthogonal methods","pmids":["16897187"],"is_preprint":false},{"year":2015,"finding":"Mutant DFNA5 (exon 8 deleted) induces programmed cell death through MAPK-related pathways in human cell lines (MAP kinase activity upregulated; inhibition partially attenuated cell death) and through mitochondrial pathways in yeast (cytochrome c oxidase genes upregulated). Both models showed downregulation of protein sorting/folding mechanisms.","method":"Microarray gene expression analysis in HEK293T cells and S. cerevisiae, MAPK pathway inhibition assays","journal":"Frontiers in cellular neuroscience","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — two model systems with microarray plus pharmacological inhibition, single lab","pmids":["26236191"],"is_preprint":false},{"year":2004,"finding":"Morpholino knockdown of dfna5 in zebrafish disrupts ugdh (UDP-glucose dehydrogenase) expression in the developing ear and pharyngeal arches, resulting in strongly reduced hyaluronic acid levels in developing semicircular canals and disorganized ear/cartilage development.","method":"Morpholino antisense knockdown in zebrafish, in situ hybridization for ugdh, HA detection in developing semicircular canals","journal":"Development (Cambridge, England)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct loss-of-function in model organism with molecular readout (ugdh expression, HA levels), single lab","pmids":["14736743"],"is_preprint":false},{"year":2020,"finding":"GSDME-C domain is palmitoylated during chemotherapy-induced pyroptosis. 2-Bromopalmitate (2-BP) inhibits GSDME-C palmitoylation and chemotherapy-induced pyroptosis. Mutation of palmitoylation sites on GSDME diminishes pyroptosis. 2-BP treatment increased the interaction between GSDME-C and GSDME-N. ZDHHC-2, 7, 11, and 15 interact with and palmitoylate GSDME.","method":"Palmitoylation assays, 2-BP pharmacological inhibition, palmitoylation site mutagenesis, co-immunoprecipitation with ZDHHC proteins, GSDME knockdown","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — mutagenesis of palmitoylation sites, co-IP with multiple ZDHHC proteins, pharmacological inhibition, single lab","pmids":["32332857"],"is_preprint":false},{"year":2022,"finding":"OTUD4 deubiquitinates and stabilizes GSDME protein, enhancing radiosensitivity in nasopharyngeal carcinoma by promoting GSDME-dependent pyroptosis. OTUD4 expression correlates with GSDME levels in NPC biopsies.","method":"Immunoprecipitation assays, mass spectrometry, in vitro and in vivo functional assays, immunohistochemistry","journal":"Journal of experimental & clinical cancer research : CR","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP with mass spectrometry identification, in vitro and in vivo validation, single lab","pmids":["36411454"],"is_preprint":false},{"year":2023,"finding":"CDC20, an E3 ubiquitin ligase component, targets GSDME for ubiquitination-mediated proteolysis in a degron-dependent manner, negatively regulating pyroptosis. Knockdown of CDC20 increases GSDME abundance and promotes transition from apoptosis to pyroptosis.","method":"Cycloheximide chase assay, immunoprecipitation, ubiquitination assay, RNA sequencing, qRT-PCR, western blotting, syngeneic mouse models","journal":"Experimental hematology & oncology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ubiquitination assay plus Co-IP, cycloheximide chase, in vivo confirmation, single lab","pmids":["37528490"],"is_preprint":false},{"year":2023,"finding":"Mannose metabolism increases N-acetylglucosamine-6-phosphate (GlcNAc-6P), which binds AMPK and facilitates AMPK phosphorylation by LKB1. Activated AMPK phosphorylates GSDME at Thr6, blocking caspase-3-induced GSDME cleavage and thereby repressing pyroptosis. This was confirmed in AMPK knockout and GSDME-T6E/T6A knock-in mice.","method":"Metabolite-AMPK binding assays, AMPK knockout mice, GSDME knock-in mice (T6E and T6A), in vitro cleavage assays, patient sample validation","journal":"Cell research","confidence":"High","confidence_rationale":"Tier 1 / Strong — identified specific phosphorylation site (Thr6), confirmed with knock-in mouse models (T6E phosphomimetic and T6A non-phosphorylatable), multiple orthogonal approaches including metabolite binding and in vivo genetic models","pmids":["37460805"],"is_preprint":false},{"year":2024,"finding":"Full-length (FL) GSDME can execute pyroptosis independent of proteolytic cleavage. UV-C-induced DNA damage activates nuclear PARP1, generating PAR polymers that activate PARP5 to PARylate GSDME, causing conformational change relieving autoinhibition. UV-C also promotes cytochrome c-catalysed cardiolipin peroxidation, elevating lipid ROS that is sensed by PARylated GSDME, leading to oxidative oligomerization and plasma membrane targeting of FL-GSDME.","method":"UV-C irradiation, PARP1/PARP5 inhibition and knockout, PARylation assays, lipid ROS detection, plasma membrane localization assays, oligomerization assays","journal":"Nature cell biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple orthogonal mechanistic experiments (PARylation, lipid ROS sensing, oligomerization, membrane targeting) establishing a novel cleavage-independent mechanism in a high-impact peer-reviewed journal","pmids":["38997456"],"is_preprint":false},{"year":2021,"finding":"GSDME regulates pore size during apoptosis-driven secondary necrosis. In anti-Fas-treated cells, GSDME accelerates cell lysis and controls passage of size-dependent dextrans through the plasma membrane. GSDME loss hampers influx of fluorescent dextrans, while efflux occurs independently of GSDME. GSDME does not affect phosphatidylserine exposure.","method":"SYTOX Blue staining, dextran influx/efflux assays with various molecular weights, GSDME knockout L929sAhFas cells, flow cytometry","journal":"Cellular and molecular life sciences : CMLS","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct pore-size characterization using size-fractionated dextrans with GSDME knockout controls, single lab","pmids":["34971436"],"is_preprint":false},{"year":2024,"finding":"Transcription factor Sp1 directly binds the GSDME promoter at the -36 to -28 site and promotes GSDME gene transcription. Sp1 knockdown or inhibition suppresses GSDME expression and reduces chemotherapy-induced pyroptosis. This regulation synergizes with STAT3 activity and antagonizes DNA methylation.","method":"Chromatin immunoprecipitation (ChIP), reporter assay, Sp1 knockdown, western blotting","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP identified direct promoter binding site, functional knockdown validated transcriptional effect, single lab","pmids":["38238307"],"is_preprint":false},{"year":2023,"finding":"STAT3 directly correlates with and positively regulates GSDME expression in macrophages during atherosclerosis development. Ox-LDL induces GSDME expression and GSDME-mediated pyroptosis in macrophages.","method":"GSDME/ApoE dual knockout mice, ox-LDL treatment of macrophages, STAT3 correlation analysis and functional experiments, single-cell transcriptomics","journal":"Nature communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo knockout model plus in vitro mechanistic analysis linking STAT3 to GSDME expression, single lab","pmids":["36807553"],"is_preprint":false},{"year":2021,"finding":"EMT-activating transcription factors ZEB1/2 directly bind the GSDME promoter and drive its transcriptional activation. EMT dictates reversible GSDME upregulation, and elevated GSDME undergoes proteolytic cleavage upon drug exposure to execute pyroptosis.","method":"Integrative bioinformatics, ChIP assay (ZEB1/2 binding to GSDME promoter), EMT induction/reversal experiments, drug treatment assays","journal":"Frontiers in cell and developmental biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP confirmed direct ZEB1/2 promoter binding, functional validation in multiple models, single lab","pmids":["34901025"],"is_preprint":false},{"year":2024,"finding":"ALKBH4 inhibits GSDME activation at the transcriptional level by suppressing H3K4me3 histone modification at the GSDME promoter region, thereby reducing 5-FU-induced pyroptosis in gastric cancer cells.","method":"ChIP assay for H3K4me3, ALKBH4 knockdown/overexpression, western blotting, colony formation, qRT-PCR","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP identified histone modification at GSDME promoter, functional ALKBH4 manipulation validated mechanistic link, single lab","pmids":["38902235"],"is_preprint":false},{"year":2024,"finding":"The non-N-terminal fragment of GSDME (GSDME-C) within macrophages combines with PDPK1, activating the PI3K-AKT pathway and facilitating M2-like macrophage polarization. The small-molecule Eliprodil inhibited PDPK1 phosphorylation mediated by GSDME.","method":"Co-immunoprecipitation, flow cytometry, Eliprodil pharmacological inhibition, single-cell sequencing, in vivo HCC models","journal":"Cellular & molecular immunology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP identifying GSDME-C/PDPK1 interaction, pharmacological inhibition, in vivo validation, single lab","pmids":["39496854"],"is_preprint":false},{"year":2024,"finding":"In platelets, caspase-3 cleaves GSDME to release GSDME-N, which targets the platelet plasma membrane forming pores and facilitating platelet granule release, promoting hyperactivity and thrombotic potential. Flotillin-2 was identified as a GSDME-N interactor that recruits GSDME-N to the platelet membrane.","method":"Human platelets and GSDME knockout mouse platelets, caspase-3 cleavage assays, membrane pore assays, Co-IP identifying flotillin-2 interaction, thrombosis models","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 / Strong — GSDME knockout mice, human platelet validation, identification of specific membrane-targeting interactor (flotillin-2) by Co-IP, multiple orthogonal readouts","pmids":["39378585"],"is_preprint":false},{"year":2024,"finding":"p53 directly mediates GSDME transcription; dual-luciferase reporter assay and ChIP-qPCR confirmed p53 binding to the GSDME promoter. ULK1 depletion additionally upregulates GSDME cleavage via ROS/NLRP3 signaling, synergizing with p53-driven basal pyroptosis.","method":"Dual-luciferase reporter assay, ChIP-qPCR, CRISPR/Cas9 kinome screen, ULK1 knockout/overexpression, GSDME knockdown, flow cytometry, LDH assay","journal":"Journal of experimental & clinical cancer research : CR","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP-qPCR and reporter assay confirmed p53 promoter binding, functional genetic validation, single lab","pmids":["39215364"],"is_preprint":false},{"year":2025,"finding":"GSDME-mediated pyroptosis in HCC is controlled by a STAT1-GSDME pyroptotic circuitry: HDAC inhibitor CXD101 promotes H3K27 hyperacetylation of IFNγ-responsive genes, driving STAT1-dependent antitumor immunity; cytotoxic lymphocytes recruited produce IFNγ/GZMB that promotes GSDME cleavage, and GSDME deletion abolishes antitumor efficacy equivalently to STAT1 knockout.","method":"ChIP-sequencing, single-cell multiomics, genetic GSDME/STAT1 knockout, orthotopic mouse models, co-culture systems, chromatin accessibility analysis","journal":"Gut","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP-seq, single-cell multiomics, genetic models; GSDME mechanistic role confirmed by deletion experiments equivalent to STAT1 KO; single lab","pmids":["39486886"],"is_preprint":false},{"year":2023,"finding":"GALNT6 promotes GSDME degradation via O-glycosylation, reducing GSDME-mediated pyroptosis in pancreatic ductal adenocarcinoma cells.","method":"GALNT6 knockdown, western blotting, immunofluorescence, ELISA, scanning electron microscopy","journal":"Frontiers in oncology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, glycosylation mechanistic claim based on knockdown and protein level changes without direct glycosylation site mapping or in vitro reconstitution","pmids":["36925932"],"is_preprint":false},{"year":2024,"finding":"GSDME deficiency in mice reduces citrullination at Arg-114 (R114) of dynamin-related protein 1 (Drp1), impairing Drp1 stability and its ability to redistribute to mitochondria for mitophagy; mutation of Drp1-R114 reduces its stability and promotes its degradation under MASH stress.","method":"RNA sequencing, quantitative proteomics, Drp1-R114 mutagenesis, GSDME knockout mice, myeloid-specific GSDME reintroduction, immunofluorescence","journal":"Cell death and differentiation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — site-specific mutagenesis of Drp1-R114, GSDME KO plus cell-type-specific reintroduction, proteomics-based mechanism; single lab","pmids":["39009654"],"is_preprint":false},{"year":2019,"finding":"Diverse targeted therapies (KRAS, EGFR, ALK inhibitors) engage the mitochondrial intrinsic apoptotic pathway, and the mobilized caspase-3 cleaves GSDME (encoded by DFNA5), which permeabilizes the cytoplasmic membrane and executes cell-lytic pyroptosis in addition to apoptosis in lung cancer cells.","method":"Immunoblot, phase-contrast imaging, scanning electron microscopy, flow cytometry, xenograft models, IHC of patient lung cancer tissues","journal":"Clinical cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple drug classes tested, multiple cell death readouts, in vivo xenograft validation and patient tissue analysis, single lab","pmids":["30061362"],"is_preprint":false},{"year":2019,"finding":"In DOX-treated cardiomyocytes, Bnip3 upregulation promotes caspase-3 activation and subsequent GSDME cleavage, leading to pyroptotic cell death. Silencing Bnip3 blunts cardiomyocyte pyroptosis by reducing caspase-3 activation and GSDME cleavage.","method":"siRNA knockdown of GSDME and Bnip3, caspase-3 inhibition, western blot, LDH assay, flow cytometry, echocardiography in mouse model","journal":"Life sciences","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pathway epistasis via knockdown of sequential components, in vivo mouse validation, single lab","pmids":["31862454"],"is_preprint":false},{"year":2024,"finding":"Cathepsin L degrades BMPR2 via the lysosomal pathway, reducing BMPR2 signaling and inducing caspase-3/GSDME-mediated endothelial cell pyroptosis to promote pulmonary hypertension. Restoring BMPR2 signaling prevents cathepsin L's pro-pyroptotic role.","method":"siRNA, lentiviral constructs, specific inhibitors, genetic cathepsin L ablation in PH rats, measurement of BMPR2/caspase-3/GSDME pathway in human PAH samples and experimental models","journal":"Hypertension (Dallas, Tex. : 1979)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo genetic ablation, human sample validation, and mechanistic rescue with BMPR2 restoration; single lab","pmids":["39403807"],"is_preprint":false},{"year":2023,"finding":"TNF-α triggers GSDME-mediated pyroptosis in myotubes through activating caspase-8 and caspase-3. TNF-α assembles TNF Complex IIb (rather than Complex IIa) to activate caspase-8, which then activates caspase-3 to cleave GSDME, leading to loss of myotubes.","method":"Caspase-8 and caspase-3 inhibitors, GSDME knockdown, comparison of TNF Complex IIa vs IIb assembly, western blotting, aged sarcopenia mouse model","journal":"Cell death discovery","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pharmacological dissection of caspase cascade, complex assembly comparison, GSDME knockdown functional validation, single lab","pmids":["36823174"],"is_preprint":false}],"current_model":"GSDME (DFNA5) is a gasdermin family pore-forming protein whose N-terminal domain permeabilizes the plasma membrane to execute pyroptosis; its primary activation mechanism is caspase-3-mediated cleavage at Asp270 (converting apoptotic signals to secondary necrosis/pyroptosis), but full-length GSDME can also be activated cleavage-independently via PARP1/PARP5-mediated PARylation coupled to lipid ROS sensing. GSDME activity is regulated at multiple levels: transcriptionally by p53, Sp1, ZEB1/2, and STAT3; epigenetically by DNA methylation and H3K4me3 (controlled by ALKBH4); post-translationally by caspase-3 cleavage, AMPK phosphorylation at Thr6 (which blocks cleavage), palmitoylation of the C-domain by ZDHHC enzymes (which modulates autoinhibition), ubiquitination by CDC20 (targeting it for degradation), and deubiquitination by OTUD4 (stabilizing it). The liberated GSDME-N fragment is recruited to the plasma membrane partly via flotillin-2 interaction, forms size-selective pores, and drives inflammatory cytokine release; the C-terminal fragment retains autoinhibitory function and also interacts with PDPK1 to activate PI3K-AKT signaling in macrophages."},"narrative":{"mechanistic_narrative":"GSDME (DFNA5) is a gasdermin-family pore-forming protein that converts apoptotic signaling into lytic, inflammatory cell death (pyroptosis/secondary necrosis) by permeabilizing the plasma membrane [PMID:28045099]. Its canonical activation is caspase-3 cleavage after Asp270, which liberates an N-terminal fragment (GSDME-N) that targets the plasma membrane; cells expressing GSDME progress to secondary necrosis upon apoptotic stimulation, whereas GSDME-null cells disassemble into apoptotic bodies [PMID:28045099]. The protein is bipartite, with the N-terminal domain executing membrane damage and the C-terminal domain masking and autoinhibiting this activity [PMID:21522185]. At the membrane GSDME-N forms size-selective pores that govern the influx of macromolecules and accelerate cell lysis [PMID:34971436], and its membrane recruitment is mediated in part by flotillin-2 [PMID:39378585]. This caspase-3/GSDME axis is engaged downstream of diverse apoptotic triggers, including targeted anticancer therapies acting through the mitochondrial intrinsic pathway [PMID:30061362] and caspase-8-dependent TNF Complex IIb signaling [PMID:36823174]. GSDME can also be activated independently of cleavage: DNA damage drives PARP1/PARP5-mediated PARylation of full-length GSDME, relieving autoinhibition and coupling to lipid-ROS sensing to trigger oxidative oligomerization and membrane targeting [PMID:38997456]. GSDME abundance and activation are controlled at multiple layers — transcriptionally by p53 [PMID:16897187, PMID:39215364], Sp1 [PMID:38238307], STAT3 [PMID:36807553] and ZEB1/2 [PMID:34901025], epigenetically through ALKBH4-regulated H3K4me3 [PMID:38902235]; post-translationally by AMPK phosphorylation at Thr6 that blocks caspase-3 cleavage [PMID:37460805], C-domain palmitoylation by ZDHHC enzymes that modulates autoinhibition [PMID:32332857], and opposing ubiquitin-dependent turnover via CDC20 and stabilization by the deubiquitinase OTUD4 [PMID:36411454, PMID:37528490]. Beyond pore formation, the GSDME-C fragment interacts with PDPK1 in macrophages to activate PI3K-AKT signaling and promote M2-like polarization [PMID:39496854]. Loss-of-function and gain-of-function studies link GSDME to autosomal dominant hearing loss (DFNA5), where exon-8-skipped mutant protein produces a necrotic gain-of-function cell-death phenotype [PMID:15173223].","teleology":[{"year":2004,"claim":"Established that disease-associated DFNA5 acts through a gain-of-function cytotoxic mechanism rather than haploinsufficiency, reframing how DFNA5 mutations cause hearing loss.","evidence":"Transfection of wild-type vs. exon-8-deleted mutant DFNA5 in mammalian cells with flow cytometry and microscopy","pmids":["15173223"],"confidence":"Medium","gaps":["Molecular identity of the death-executing fragment not yet defined","Mechanism by which the mutant is constitutively active unresolved"]},{"year":2004,"claim":"First in vivo developmental role addressed: whether dfna5 functions in ear morphogenesis, linking it to ugdh-dependent matrix synthesis.","evidence":"Morpholino knockdown in zebrafish with ugdh in situ hybridization and hyaluronic acid detection","pmids":["14736743"],"confidence":"Medium","gaps":["Connection between a pore-forming death protein and ugdh regulation mechanistically unexplained","Relationship to the later-defined pyroptotic function unclear"]},{"year":2006,"claim":"Answered how GSDME expression is induced in response to genotoxic stress, placing it downstream of p53 as a death effector.","evidence":"ChIP and reporter assays identifying a p53 site in intron 1, validated in p53-knockout mice after irradiation","pmids":["16897187"],"confidence":"High","gaps":["Did not define the post-transcriptional activation step","Effector mechanism of the induced protein not yet known"]},{"year":2011,"claim":"Defined the bipartite architecture, showing the N-terminal domain is the death-inducing module and the C-terminal domain is autoinhibitory.","evidence":"Domain transfection in HEK293T cells plus Dfna5-knockout mouse expression microarray","pmids":["21522185"],"confidence":"Medium","gaps":["The physiological protease liberating the N-domain not identified","Membrane-permeabilization activity vs. apoptosis distinction unresolved"]},{"year":2017,"claim":"Identified the activating cleavage event, establishing GSDME as a caspase-3 substrate whose N-fragment drives secondary necrosis/pyroptosis.","evidence":"Cleavage-site mutagenesis (Asp270), knockout, and plasma-membrane targeting assays after apoptotic stimulation","pmids":["28045099"],"confidence":"High","gaps":["Pore architecture and selectivity not characterized","Membrane recruitment partners unknown"]},{"year":2019,"claim":"Connected GSDME activation to clinically relevant apoptotic inputs—targeted therapies and Bnip3/TNF signaling—showing it converts drug- and cytokine-induced apoptosis into pyroptosis across tissues.","evidence":"Drug treatment, Bnip3/caspase manipulation, knockdowns, xenograft and cardiomyocyte/myotube models","pmids":["30061362","31862454"],"confidence":"Medium","gaps":["Tissue-specific determinants of pyroptosis vs. apoptosis outcome unclear","Quantitative threshold of GSDME needed for lysis undefined"]},{"year":2021,"claim":"Characterized the pore biophysically, showing GSDME governs size-selective macromolecular passage and accelerates lysis during secondary necrosis.","evidence":"Size-fractionated dextran influx/efflux and SYTOX assays in GSDME-knockout cells","pmids":["34971436"],"confidence":"Medium","gaps":["Structural pore stoichiometry not resolved","Single cell-line system"]},{"year":2021,"claim":"Expanded transcriptional control by showing EMT-driving ZEB1/2 directly activate GSDME, linking cell-state plasticity to pyroptotic competence.","evidence":"ChIP for ZEB1/2 promoter binding plus EMT induction/reversal and drug-treatment assays","pmids":["34901025"],"confidence":"Medium","gaps":["Reversibility mechanism at chromatin not defined","Interplay with other transcription factors untested here"]},{"year":2020,"claim":"Revealed lipid-modification control of autoinhibition: ZDHHC-mediated palmitoylation of GSDME-C modulates pyroptotic activation.","evidence":"Palmitoylation assays, site mutagenesis, 2-BP inhibition and Co-IP with multiple ZDHHC enzymes","pmids":["32332857"],"confidence":"Medium","gaps":["Precise effect of palmitoylation on N/C-domain interaction mechanistically incomplete","Which ZDHHC dominates in vivo unknown"]},{"year":2022,"claim":"Identified deubiquitination as a stabilizing layer, with OTUD4 raising GSDME levels to enhance radiosensitivity.","evidence":"Co-IP/mass spectrometry, functional in vitro/in vivo assays and patient IHC in nasopharyngeal carcinoma","pmids":["36411454"],"confidence":"Medium","gaps":["Ubiquitin linkage type and sites not mapped","Opposing ligase at the time unidentified"]},{"year":2023,"claim":"Established opposing ubiquitin-dependent turnover, with CDC20 driving degron-dependent GSDME degradation to gate the apoptosis-to-pyroptosis transition.","evidence":"Cycloheximide chase, ubiquitination assays, Co-IP and syngeneic mouse models","pmids":["37528490"],"confidence":"Medium","gaps":["Degron sequence detail limited","Regulation of CDC20-GSDME by cell-cycle state unexplored"]},{"year":2023,"claim":"Discovered metabolic phospho-regulation: GlcNAc-6P/AMPK phosphorylation of GSDME at Thr6 blocks caspase-3 cleavage, repressing pyroptosis.","evidence":"Metabolite-AMPK binding, AMPK-knockout and GSDME T6E/T6A knock-in mice with in vitro cleavage assays","pmids":["37460805"],"confidence":"High","gaps":["Structural basis of how Thr6 phosphorylation occludes the Asp270 site unresolved","Generality across cell types untested"]},{"year":2023,"claim":"Extended transcriptional and functional reach to macrophage biology and atherosclerosis via STAT3-driven GSDME expression.","evidence":"GSDME/ApoE double-knockout mice, ox-LDL macrophage treatment and STAT3 functional analysis","pmids":["36807553"],"confidence":"Medium","gaps":["Direct STAT3 promoter binding not shown here","Cell-autonomy vs. systemic effects partly conflated"]},{"year":2024,"claim":"Defined a cleavage-independent activation route, showing PARP1/PARP5 PARylation of full-length GSDME relieves autoinhibition and couples to lipid-ROS sensing for oligomerization.","evidence":"UV-C irradiation with PARP1/PARP5 inhibition/knockout, PARylation, lipid-ROS, oligomerization and membrane-targeting assays","pmids":["38997456"],"confidence":"High","gaps":["PARylation site(s) on GSDME not mapped","How lipid ROS is structurally sensed unresolved"]},{"year":2024,"claim":"Identified flotillin-2 as the membrane-recruitment partner for GSDME-N and extended pore function to platelet hyperactivity and thrombosis.","evidence":"Human and GSDME-knockout mouse platelets, cleavage and pore assays, Co-IP and thrombosis models","pmids":["39378585"],"confidence":"High","gaps":["Structural basis of flotillin-2/GSDME-N interaction unknown","Whether flotillin-2 recruitment generalizes to other cell types untested"]},{"year":2024,"claim":"Revealed a non-pore-forming signaling role: GSDME-C binds PDPK1 to activate PI3K-AKT and promote M2-like macrophage polarization.","evidence":"Co-IP, Eliprodil inhibition, single-cell sequencing and in vivo HCC models","pmids":["39496854"],"confidence":"Medium","gaps":["Structural interface of GSDME-C/PDPK1 undefined","Balance between pyroptotic and signaling roles in vivo unclear"]},{"year":2024,"claim":"Added further regulatory and downstream nodes: Sp1 promoter activation, ALKBH4-controlled H3K4me3 repression, p53/ULK1 control, and a GSDME-dependent Drp1-R114 citrullination/mitophagy axis.","evidence":"ChIP/reporter assays, ALKBH4 and ULK1 manipulation, GSDME-knockout/reintroduction and Drp1-R114 mutagenesis across cancer and MASH models","pmids":["38238307","38902235","39215364","39009654"],"confidence":"Medium","gaps":["Mechanism linking GSDME to Drp1 citrullination not fully defined","Integration of these regulators into a single hierarchy untested"]},{"year":2024,"claim":"Showed GSDME is the lytic effector arm of cytotoxic-lymphocyte/IFNγ antitumor immunity, with STAT1-GSDME circuitry required for HDAC-inhibitor efficacy.","evidence":"ChIP-seq, single-cell multiomics, genetic GSDME/STAT1 knockout and orthotopic HCC models","pmids":["39486886"],"confidence":"Medium","gaps":["Which protease (GZMB vs caspase-3) dominates cleavage in this setting partly open","Direct STAT1 regulation of GSDME locus vs. immune-cell recruitment effects entangled"]},{"year":null,"claim":"How the multiple activation inputs (caspase-3 cleavage, PARylation, lipid-ROS sensing) and antagonistic post-translational marks (Thr6 phosphorylation, palmitoylation, ubiquitination, O-glycosylation) are integrated at the structural level to set the threshold between apoptosis and pyroptosis remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No high-resolution structure of activated GSDME-N pore or of the autoinhibited full-length protein in the corpus","PARylation and modification sites not jointly mapped","Quantitative model of competing modifications absent"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[0,11,17]},{"term_id":"GO:0008289","term_label":"lipid binding","supporting_discovery_ids":[10]},{"term_id":"GO:0140313","term_label":"molecular sequestering activity","supporting_discovery_ids":[2]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[0,11,17,10]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[10]}],"pathway":[{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[0,22,25]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[13,16,19]}],"complexes":[],"partners":["ZDHHC2","ZDHHC7","OTUD4","CDC20","PDPK1","FLOT2","PARP1","AMPK"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"O60443","full_name":"Gasdermin-E","aliases":["Inversely correlated with estrogen receptor expression 1","ICERE-1","Non-syndromic hearing impairment protein 5"],"length_aa":496,"mass_kda":54.6,"function":"Precursor of a pore-forming protein that converts non-inflammatory apoptosis to pyroptosis (PubMed:27281216, PubMed:28459430, PubMed:33852854, PubMed:35594856, PubMed:36607699). This form constitutes the precursor of the pore-forming protein: upon cleavage, the released N-terminal moiety (Gasdermin-E, N-terminal) binds to membranes and forms pores, triggering pyroptosis (PubMed:28459430) Pore-forming protein produced by cleavage by CASP3 or granzyme B (GZMB), which converts non-inflammatory apoptosis to pyroptosis or promotes granzyme-mediated pyroptosis, respectively (PubMed:27281216, PubMed:28459430, PubMed:32188940, PubMed:33852854, PubMed:35594856). After cleavage, moves to the plasma membrane, homooligomerizes within the membrane and forms pores of 10-15 nanometers (nm) of inner diameter, allowing the release of mature interleukins (IL1B and IL16) and triggering pyroptosis (PubMed:28459430, PubMed:32188940, PubMed:33852854, PubMed:35594856). Binds to inner leaflet lipids, bisphosphorylated phosphatidylinositols, such as phosphatidylinositol (4,5)-bisphosphate (PubMed:28459430). 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1979)","url":"https://pubmed.ncbi.nlm.nih.gov/39403807","citation_count":13,"is_preprint":false},{"pmid":"40432601","id":"PMC_40432601","title":"Pyroptosis-Inducing Platinum(IV) Prodrugs via GSDME Pathway for Chemoimmunotherapy and Metastasis Inhibition in Triple-Negative Breast Cancer.","date":"2025","source":"Advanced science (Weinheim, Baden-Wurttemberg, Germany)","url":"https://pubmed.ncbi.nlm.nih.gov/40432601","citation_count":13,"is_preprint":false},{"pmid":"40523886","id":"PMC_40523886","title":"Repurposing MDM2 inhibitor RG7388 for TP53-mutant NSCLC: a p53-independent pyroptotic mechanism via ROS/p-p38/NOXA/caspase-3/GSDME axis.","date":"2025","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/40523886","citation_count":13,"is_preprint":false},{"pmid":"37761020","id":"PMC_37761020","title":"GSDME in Endothelial Cells: Inducing Vascular Inflammation and Atherosclerosis via Mitochondrial Damage and STING Pathway Activation.","date":"2023","source":"Biomedicines","url":"https://pubmed.ncbi.nlm.nih.gov/37761020","citation_count":13,"is_preprint":false},{"pmid":"39378585","id":"PMC_39378585","title":"GSDME-mediated pyroptosis contributes to chemotherapy-induced platelet hyperactivity and thrombotic potential.","date":"2024","source":"Blood","url":"https://pubmed.ncbi.nlm.nih.gov/39378585","citation_count":12,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":55659,"output_tokens":6694,"usd":0.133693,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":15609,"output_tokens":5429,"usd":0.106885,"stage2_stop_reason":"end_turn"},"total_usd":0.240578,"stage1_batch_id":"msgbatch_01DUPACZi7AAWwuZiUjseQF4","stage2_batch_id":"msgbatch_018yt7chE3f48PEFtDwtj6AE","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2017,\n      \"finding\": \"Caspase-3 cleaves GSDME (DFNA5) after Asp270 to generate a necrotic N-terminal fragment (GSDME-N) that targets the plasma membrane to induce secondary necrosis/pyroptosis. Cells expressing GSDME progress to secondary necrosis upon apoptotic stimulation, while GSDME-deleted cells disassemble into apoptotic bodies instead.\",\n      \"method\": \"Cell-based cleavage assays, GSDME deletion/knockout, apoptotic stimulation (etoposide, VSV infection), plasma membrane targeting assays\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — direct identification of cleavage site by mutagenesis, loss-of-function phenotype, replicated across multiple labs subsequently\",\n      \"pmids\": [\"28045099\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Mutant DFNA5 (exon 8 skipped) transfected into HEK293T and COS-1 cells causes approximately doubled post-transfection cell death attributable to necrotic (not apoptotic) events, supporting a gain-of-function mechanism for DFNA5-associated hearing loss.\",\n      \"method\": \"Transfection of GFP-tagged wild-type vs. mutant DFNA5 in mammalian cell lines, flow cytometry, fluorescence microscopy\",\n      \"journal\": \"Journal of medical genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct functional assay in mammalian cells with two readouts (flow cytometry + microscopy), single lab\",\n      \"pmids\": [\"15173223\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"GSDME is composed of two domains separated by a hinge region: the N-terminal domain induces apoptosis when transfected in HEK293T cells, while the C-terminal domain masks and regulates this apoptosis-inducing capability. Knockout mice microarray analysis further supported involvement of GSDME in apoptosis-related pathways.\",\n      \"method\": \"Domain transfection in HEK293T cells, gene expression microarray using Dfna5 knockout mice\",\n      \"journal\": \"European journal of human genetics : EJHG\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional domain dissection with two orthogonal approaches (transfection + knockout microarray), single lab\",\n      \"pmids\": [\"21522185\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"DFNA5 gene expression is strongly induced by p53 (both exogenous and endogenous). Chromatin immunoprecipitation identified a p53-binding sequence in intron 1 of DFNA5, and a reporter assay confirmed p53-dependent transcriptional activity at this site. Ectopic DFNA5 enhanced etoposide-induced cell death in a p53-dependent manner.\",\n      \"method\": \"Chromatin immunoprecipitation (ChIP), reporter gene assay, p53 overexpression/knockout systems, gamma-ray irradiation in p53+/+ vs p53-/- mice\",\n      \"journal\": \"Journal of human genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — ChIP directly identified p53 binding site, reporter assay confirmed transcriptional activity, in vivo validation in p53 knockout mice, multiple orthogonal methods\",\n      \"pmids\": [\"16897187\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Mutant DFNA5 (exon 8 deleted) induces programmed cell death through MAPK-related pathways in human cell lines (MAP kinase activity upregulated; inhibition partially attenuated cell death) and through mitochondrial pathways in yeast (cytochrome c oxidase genes upregulated). Both models showed downregulation of protein sorting/folding mechanisms.\",\n      \"method\": \"Microarray gene expression analysis in HEK293T cells and S. cerevisiae, MAPK pathway inhibition assays\",\n      \"journal\": \"Frontiers in cellular neuroscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — two model systems with microarray plus pharmacological inhibition, single lab\",\n      \"pmids\": [\"26236191\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Morpholino knockdown of dfna5 in zebrafish disrupts ugdh (UDP-glucose dehydrogenase) expression in the developing ear and pharyngeal arches, resulting in strongly reduced hyaluronic acid levels in developing semicircular canals and disorganized ear/cartilage development.\",\n      \"method\": \"Morpholino antisense knockdown in zebrafish, in situ hybridization for ugdh, HA detection in developing semicircular canals\",\n      \"journal\": \"Development (Cambridge, England)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct loss-of-function in model organism with molecular readout (ugdh expression, HA levels), single lab\",\n      \"pmids\": [\"14736743\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"GSDME-C domain is palmitoylated during chemotherapy-induced pyroptosis. 2-Bromopalmitate (2-BP) inhibits GSDME-C palmitoylation and chemotherapy-induced pyroptosis. Mutation of palmitoylation sites on GSDME diminishes pyroptosis. 2-BP treatment increased the interaction between GSDME-C and GSDME-N. ZDHHC-2, 7, 11, and 15 interact with and palmitoylate GSDME.\",\n      \"method\": \"Palmitoylation assays, 2-BP pharmacological inhibition, palmitoylation site mutagenesis, co-immunoprecipitation with ZDHHC proteins, GSDME knockdown\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — mutagenesis of palmitoylation sites, co-IP with multiple ZDHHC proteins, pharmacological inhibition, single lab\",\n      \"pmids\": [\"32332857\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"OTUD4 deubiquitinates and stabilizes GSDME protein, enhancing radiosensitivity in nasopharyngeal carcinoma by promoting GSDME-dependent pyroptosis. OTUD4 expression correlates with GSDME levels in NPC biopsies.\",\n      \"method\": \"Immunoprecipitation assays, mass spectrometry, in vitro and in vivo functional assays, immunohistochemistry\",\n      \"journal\": \"Journal of experimental & clinical cancer research : CR\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP with mass spectrometry identification, in vitro and in vivo validation, single lab\",\n      \"pmids\": [\"36411454\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"CDC20, an E3 ubiquitin ligase component, targets GSDME for ubiquitination-mediated proteolysis in a degron-dependent manner, negatively regulating pyroptosis. Knockdown of CDC20 increases GSDME abundance and promotes transition from apoptosis to pyroptosis.\",\n      \"method\": \"Cycloheximide chase assay, immunoprecipitation, ubiquitination assay, RNA sequencing, qRT-PCR, western blotting, syngeneic mouse models\",\n      \"journal\": \"Experimental hematology & oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ubiquitination assay plus Co-IP, cycloheximide chase, in vivo confirmation, single lab\",\n      \"pmids\": [\"37528490\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Mannose metabolism increases N-acetylglucosamine-6-phosphate (GlcNAc-6P), which binds AMPK and facilitates AMPK phosphorylation by LKB1. Activated AMPK phosphorylates GSDME at Thr6, blocking caspase-3-induced GSDME cleavage and thereby repressing pyroptosis. This was confirmed in AMPK knockout and GSDME-T6E/T6A knock-in mice.\",\n      \"method\": \"Metabolite-AMPK binding assays, AMPK knockout mice, GSDME knock-in mice (T6E and T6A), in vitro cleavage assays, patient sample validation\",\n      \"journal\": \"Cell research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — identified specific phosphorylation site (Thr6), confirmed with knock-in mouse models (T6E phosphomimetic and T6A non-phosphorylatable), multiple orthogonal approaches including metabolite binding and in vivo genetic models\",\n      \"pmids\": [\"37460805\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Full-length (FL) GSDME can execute pyroptosis independent of proteolytic cleavage. UV-C-induced DNA damage activates nuclear PARP1, generating PAR polymers that activate PARP5 to PARylate GSDME, causing conformational change relieving autoinhibition. UV-C also promotes cytochrome c-catalysed cardiolipin peroxidation, elevating lipid ROS that is sensed by PARylated GSDME, leading to oxidative oligomerization and plasma membrane targeting of FL-GSDME.\",\n      \"method\": \"UV-C irradiation, PARP1/PARP5 inhibition and knockout, PARylation assays, lipid ROS detection, plasma membrane localization assays, oligomerization assays\",\n      \"journal\": \"Nature cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple orthogonal mechanistic experiments (PARylation, lipid ROS sensing, oligomerization, membrane targeting) establishing a novel cleavage-independent mechanism in a high-impact peer-reviewed journal\",\n      \"pmids\": [\"38997456\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"GSDME regulates pore size during apoptosis-driven secondary necrosis. In anti-Fas-treated cells, GSDME accelerates cell lysis and controls passage of size-dependent dextrans through the plasma membrane. GSDME loss hampers influx of fluorescent dextrans, while efflux occurs independently of GSDME. GSDME does not affect phosphatidylserine exposure.\",\n      \"method\": \"SYTOX Blue staining, dextran influx/efflux assays with various molecular weights, GSDME knockout L929sAhFas cells, flow cytometry\",\n      \"journal\": \"Cellular and molecular life sciences : CMLS\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct pore-size characterization using size-fractionated dextrans with GSDME knockout controls, single lab\",\n      \"pmids\": [\"34971436\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Transcription factor Sp1 directly binds the GSDME promoter at the -36 to -28 site and promotes GSDME gene transcription. Sp1 knockdown or inhibition suppresses GSDME expression and reduces chemotherapy-induced pyroptosis. This regulation synergizes with STAT3 activity and antagonizes DNA methylation.\",\n      \"method\": \"Chromatin immunoprecipitation (ChIP), reporter assay, Sp1 knockdown, western blotting\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP identified direct promoter binding site, functional knockdown validated transcriptional effect, single lab\",\n      \"pmids\": [\"38238307\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"STAT3 directly correlates with and positively regulates GSDME expression in macrophages during atherosclerosis development. Ox-LDL induces GSDME expression and GSDME-mediated pyroptosis in macrophages.\",\n      \"method\": \"GSDME/ApoE dual knockout mice, ox-LDL treatment of macrophages, STAT3 correlation analysis and functional experiments, single-cell transcriptomics\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo knockout model plus in vitro mechanistic analysis linking STAT3 to GSDME expression, single lab\",\n      \"pmids\": [\"36807553\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"EMT-activating transcription factors ZEB1/2 directly bind the GSDME promoter and drive its transcriptional activation. EMT dictates reversible GSDME upregulation, and elevated GSDME undergoes proteolytic cleavage upon drug exposure to execute pyroptosis.\",\n      \"method\": \"Integrative bioinformatics, ChIP assay (ZEB1/2 binding to GSDME promoter), EMT induction/reversal experiments, drug treatment assays\",\n      \"journal\": \"Frontiers in cell and developmental biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP confirmed direct ZEB1/2 promoter binding, functional validation in multiple models, single lab\",\n      \"pmids\": [\"34901025\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"ALKBH4 inhibits GSDME activation at the transcriptional level by suppressing H3K4me3 histone modification at the GSDME promoter region, thereby reducing 5-FU-induced pyroptosis in gastric cancer cells.\",\n      \"method\": \"ChIP assay for H3K4me3, ALKBH4 knockdown/overexpression, western blotting, colony formation, qRT-PCR\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP identified histone modification at GSDME promoter, functional ALKBH4 manipulation validated mechanistic link, single lab\",\n      \"pmids\": [\"38902235\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"The non-N-terminal fragment of GSDME (GSDME-C) within macrophages combines with PDPK1, activating the PI3K-AKT pathway and facilitating M2-like macrophage polarization. The small-molecule Eliprodil inhibited PDPK1 phosphorylation mediated by GSDME.\",\n      \"method\": \"Co-immunoprecipitation, flow cytometry, Eliprodil pharmacological inhibition, single-cell sequencing, in vivo HCC models\",\n      \"journal\": \"Cellular & molecular immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP identifying GSDME-C/PDPK1 interaction, pharmacological inhibition, in vivo validation, single lab\",\n      \"pmids\": [\"39496854\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"In platelets, caspase-3 cleaves GSDME to release GSDME-N, which targets the platelet plasma membrane forming pores and facilitating platelet granule release, promoting hyperactivity and thrombotic potential. Flotillin-2 was identified as a GSDME-N interactor that recruits GSDME-N to the platelet membrane.\",\n      \"method\": \"Human platelets and GSDME knockout mouse platelets, caspase-3 cleavage assays, membrane pore assays, Co-IP identifying flotillin-2 interaction, thrombosis models\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — GSDME knockout mice, human platelet validation, identification of specific membrane-targeting interactor (flotillin-2) by Co-IP, multiple orthogonal readouts\",\n      \"pmids\": [\"39378585\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"p53 directly mediates GSDME transcription; dual-luciferase reporter assay and ChIP-qPCR confirmed p53 binding to the GSDME promoter. ULK1 depletion additionally upregulates GSDME cleavage via ROS/NLRP3 signaling, synergizing with p53-driven basal pyroptosis.\",\n      \"method\": \"Dual-luciferase reporter assay, ChIP-qPCR, CRISPR/Cas9 kinome screen, ULK1 knockout/overexpression, GSDME knockdown, flow cytometry, LDH assay\",\n      \"journal\": \"Journal of experimental & clinical cancer research : CR\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP-qPCR and reporter assay confirmed p53 promoter binding, functional genetic validation, single lab\",\n      \"pmids\": [\"39215364\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"GSDME-mediated pyroptosis in HCC is controlled by a STAT1-GSDME pyroptotic circuitry: HDAC inhibitor CXD101 promotes H3K27 hyperacetylation of IFNγ-responsive genes, driving STAT1-dependent antitumor immunity; cytotoxic lymphocytes recruited produce IFNγ/GZMB that promotes GSDME cleavage, and GSDME deletion abolishes antitumor efficacy equivalently to STAT1 knockout.\",\n      \"method\": \"ChIP-sequencing, single-cell multiomics, genetic GSDME/STAT1 knockout, orthotopic mouse models, co-culture systems, chromatin accessibility analysis\",\n      \"journal\": \"Gut\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP-seq, single-cell multiomics, genetic models; GSDME mechanistic role confirmed by deletion experiments equivalent to STAT1 KO; single lab\",\n      \"pmids\": [\"39486886\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"GALNT6 promotes GSDME degradation via O-glycosylation, reducing GSDME-mediated pyroptosis in pancreatic ductal adenocarcinoma cells.\",\n      \"method\": \"GALNT6 knockdown, western blotting, immunofluorescence, ELISA, scanning electron microscopy\",\n      \"journal\": \"Frontiers in oncology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, glycosylation mechanistic claim based on knockdown and protein level changes without direct glycosylation site mapping or in vitro reconstitution\",\n      \"pmids\": [\"36925932\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"GSDME deficiency in mice reduces citrullination at Arg-114 (R114) of dynamin-related protein 1 (Drp1), impairing Drp1 stability and its ability to redistribute to mitochondria for mitophagy; mutation of Drp1-R114 reduces its stability and promotes its degradation under MASH stress.\",\n      \"method\": \"RNA sequencing, quantitative proteomics, Drp1-R114 mutagenesis, GSDME knockout mice, myeloid-specific GSDME reintroduction, immunofluorescence\",\n      \"journal\": \"Cell death and differentiation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — site-specific mutagenesis of Drp1-R114, GSDME KO plus cell-type-specific reintroduction, proteomics-based mechanism; single lab\",\n      \"pmids\": [\"39009654\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Diverse targeted therapies (KRAS, EGFR, ALK inhibitors) engage the mitochondrial intrinsic apoptotic pathway, and the mobilized caspase-3 cleaves GSDME (encoded by DFNA5), which permeabilizes the cytoplasmic membrane and executes cell-lytic pyroptosis in addition to apoptosis in lung cancer cells.\",\n      \"method\": \"Immunoblot, phase-contrast imaging, scanning electron microscopy, flow cytometry, xenograft models, IHC of patient lung cancer tissues\",\n      \"journal\": \"Clinical cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple drug classes tested, multiple cell death readouts, in vivo xenograft validation and patient tissue analysis, single lab\",\n      \"pmids\": [\"30061362\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"In DOX-treated cardiomyocytes, Bnip3 upregulation promotes caspase-3 activation and subsequent GSDME cleavage, leading to pyroptotic cell death. Silencing Bnip3 blunts cardiomyocyte pyroptosis by reducing caspase-3 activation and GSDME cleavage.\",\n      \"method\": \"siRNA knockdown of GSDME and Bnip3, caspase-3 inhibition, western blot, LDH assay, flow cytometry, echocardiography in mouse model\",\n      \"journal\": \"Life sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pathway epistasis via knockdown of sequential components, in vivo mouse validation, single lab\",\n      \"pmids\": [\"31862454\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Cathepsin L degrades BMPR2 via the lysosomal pathway, reducing BMPR2 signaling and inducing caspase-3/GSDME-mediated endothelial cell pyroptosis to promote pulmonary hypertension. Restoring BMPR2 signaling prevents cathepsin L's pro-pyroptotic role.\",\n      \"method\": \"siRNA, lentiviral constructs, specific inhibitors, genetic cathepsin L ablation in PH rats, measurement of BMPR2/caspase-3/GSDME pathway in human PAH samples and experimental models\",\n      \"journal\": \"Hypertension (Dallas, Tex. : 1979)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo genetic ablation, human sample validation, and mechanistic rescue with BMPR2 restoration; single lab\",\n      \"pmids\": [\"39403807\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"TNF-α triggers GSDME-mediated pyroptosis in myotubes through activating caspase-8 and caspase-3. TNF-α assembles TNF Complex IIb (rather than Complex IIa) to activate caspase-8, which then activates caspase-3 to cleave GSDME, leading to loss of myotubes.\",\n      \"method\": \"Caspase-8 and caspase-3 inhibitors, GSDME knockdown, comparison of TNF Complex IIa vs IIb assembly, western blotting, aged sarcopenia mouse model\",\n      \"journal\": \"Cell death discovery\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pharmacological dissection of caspase cascade, complex assembly comparison, GSDME knockdown functional validation, single lab\",\n      \"pmids\": [\"36823174\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"GSDME (DFNA5) is a gasdermin family pore-forming protein whose N-terminal domain permeabilizes the plasma membrane to execute pyroptosis; its primary activation mechanism is caspase-3-mediated cleavage at Asp270 (converting apoptotic signals to secondary necrosis/pyroptosis), but full-length GSDME can also be activated cleavage-independently via PARP1/PARP5-mediated PARylation coupled to lipid ROS sensing. GSDME activity is regulated at multiple levels: transcriptionally by p53, Sp1, ZEB1/2, and STAT3; epigenetically by DNA methylation and H3K4me3 (controlled by ALKBH4); post-translationally by caspase-3 cleavage, AMPK phosphorylation at Thr6 (which blocks cleavage), palmitoylation of the C-domain by ZDHHC enzymes (which modulates autoinhibition), ubiquitination by CDC20 (targeting it for degradation), and deubiquitination by OTUD4 (stabilizing it). The liberated GSDME-N fragment is recruited to the plasma membrane partly via flotillin-2 interaction, forms size-selective pores, and drives inflammatory cytokine release; the C-terminal fragment retains autoinhibitory function and also interacts with PDPK1 to activate PI3K-AKT signaling in macrophages.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"GSDME (DFNA5) is a gasdermin-family pore-forming protein that converts apoptotic signaling into lytic, inflammatory cell death (pyroptosis/secondary necrosis) by permeabilizing the plasma membrane [#0]. Its canonical activation is caspase-3 cleavage after Asp270, which liberates an N-terminal fragment (GSDME-N) that targets the plasma membrane; cells expressing GSDME progress to secondary necrosis upon apoptotic stimulation, whereas GSDME-null cells disassemble into apoptotic bodies [#0]. The protein is bipartite, with the N-terminal domain executing membrane damage and the C-terminal domain masking and autoinhibiting this activity [#2]. At the membrane GSDME-N forms size-selective pores that govern the influx of macromolecules and accelerate cell lysis [#11], and its membrane recruitment is mediated in part by flotillin-2 [#17]. This caspase-3/GSDME axis is engaged downstream of diverse apoptotic triggers, including targeted anticancer therapies acting through the mitochondrial intrinsic pathway [#22] and caspase-8-dependent TNF Complex IIb signaling [#25]. GSDME can also be activated independently of cleavage: DNA damage drives PARP1/PARP5-mediated PARylation of full-length GSDME, relieving autoinhibition and coupling to lipid-ROS sensing to trigger oxidative oligomerization and membrane targeting [#10]. GSDME abundance and activation are controlled at multiple layers — transcriptionally by p53 [#3, #18], Sp1 [#12], STAT3 [#13] and ZEB1/2 [#14], epigenetically through ALKBH4-regulated H3K4me3 [#15]; post-translationally by AMPK phosphorylation at Thr6 that blocks caspase-3 cleavage [#9], C-domain palmitoylation by ZDHHC enzymes that modulates autoinhibition [#6], and opposing ubiquitin-dependent turnover via CDC20 and stabilization by the deubiquitinase OTUD4 [#7, #8]. Beyond pore formation, the GSDME-C fragment interacts with PDPK1 in macrophages to activate PI3K-AKT signaling and promote M2-like polarization [#16]. Loss-of-function and gain-of-function studies link GSDME to autosomal dominant hearing loss (DFNA5), where exon-8-skipped mutant protein produces a necrotic gain-of-function cell-death phenotype [#1].\",\n  \"teleology\": [\n    {\n      \"year\": 2004,\n      \"claim\": \"Established that disease-associated DFNA5 acts through a gain-of-function cytotoxic mechanism rather than haploinsufficiency, reframing how DFNA5 mutations cause hearing loss.\",\n      \"evidence\": \"Transfection of wild-type vs. exon-8-deleted mutant DFNA5 in mammalian cells with flow cytometry and microscopy\",\n      \"pmids\": [\"15173223\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular identity of the death-executing fragment not yet defined\", \"Mechanism by which the mutant is constitutively active unresolved\"]\n    },\n    {\n      \"year\": 2004,\n      \"claim\": \"First in vivo developmental role addressed: whether dfna5 functions in ear morphogenesis, linking it to ugdh-dependent matrix synthesis.\",\n      \"evidence\": \"Morpholino knockdown in zebrafish with ugdh in situ hybridization and hyaluronic acid detection\",\n      \"pmids\": [\"14736743\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Connection between a pore-forming death protein and ugdh regulation mechanistically unexplained\", \"Relationship to the later-defined pyroptotic function unclear\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Answered how GSDME expression is induced in response to genotoxic stress, placing it downstream of p53 as a death effector.\",\n      \"evidence\": \"ChIP and reporter assays identifying a p53 site in intron 1, validated in p53-knockout mice after irradiation\",\n      \"pmids\": [\"16897187\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define the post-transcriptional activation step\", \"Effector mechanism of the induced protein not yet known\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Defined the bipartite architecture, showing the N-terminal domain is the death-inducing module and the C-terminal domain is autoinhibitory.\",\n      \"evidence\": \"Domain transfection in HEK293T cells plus Dfna5-knockout mouse expression microarray\",\n      \"pmids\": [\"21522185\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"The physiological protease liberating the N-domain not identified\", \"Membrane-permeabilization activity vs. apoptosis distinction unresolved\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Identified the activating cleavage event, establishing GSDME as a caspase-3 substrate whose N-fragment drives secondary necrosis/pyroptosis.\",\n      \"evidence\": \"Cleavage-site mutagenesis (Asp270), knockout, and plasma-membrane targeting assays after apoptotic stimulation\",\n      \"pmids\": [\"28045099\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Pore architecture and selectivity not characterized\", \"Membrane recruitment partners unknown\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Connected GSDME activation to clinically relevant apoptotic inputs—targeted therapies and Bnip3/TNF signaling—showing it converts drug- and cytokine-induced apoptosis into pyroptosis across tissues.\",\n      \"evidence\": \"Drug treatment, Bnip3/caspase manipulation, knockdowns, xenograft and cardiomyocyte/myotube models\",\n      \"pmids\": [\"30061362\", \"31862454\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Tissue-specific determinants of pyroptosis vs. apoptosis outcome unclear\", \"Quantitative threshold of GSDME needed for lysis undefined\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Characterized the pore biophysically, showing GSDME governs size-selective macromolecular passage and accelerates lysis during secondary necrosis.\",\n      \"evidence\": \"Size-fractionated dextran influx/efflux and SYTOX assays in GSDME-knockout cells\",\n      \"pmids\": [\"34971436\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural pore stoichiometry not resolved\", \"Single cell-line system\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Expanded transcriptional control by showing EMT-driving ZEB1/2 directly activate GSDME, linking cell-state plasticity to pyroptotic competence.\",\n      \"evidence\": \"ChIP for ZEB1/2 promoter binding plus EMT induction/reversal and drug-treatment assays\",\n      \"pmids\": [\"34901025\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Reversibility mechanism at chromatin not defined\", \"Interplay with other transcription factors untested here\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Revealed lipid-modification control of autoinhibition: ZDHHC-mediated palmitoylation of GSDME-C modulates pyroptotic activation.\",\n      \"evidence\": \"Palmitoylation assays, site mutagenesis, 2-BP inhibition and Co-IP with multiple ZDHHC enzymes\",\n      \"pmids\": [\"32332857\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Precise effect of palmitoylation on N/C-domain interaction mechanistically incomplete\", \"Which ZDHHC dominates in vivo unknown\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Identified deubiquitination as a stabilizing layer, with OTUD4 raising GSDME levels to enhance radiosensitivity.\",\n      \"evidence\": \"Co-IP/mass spectrometry, functional in vitro/in vivo assays and patient IHC in nasopharyngeal carcinoma\",\n      \"pmids\": [\"36411454\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Ubiquitin linkage type and sites not mapped\", \"Opposing ligase at the time unidentified\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Established opposing ubiquitin-dependent turnover, with CDC20 driving degron-dependent GSDME degradation to gate the apoptosis-to-pyroptosis transition.\",\n      \"evidence\": \"Cycloheximide chase, ubiquitination assays, Co-IP and syngeneic mouse models\",\n      \"pmids\": [\"37528490\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Degron sequence detail limited\", \"Regulation of CDC20-GSDME by cell-cycle state unexplored\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Discovered metabolic phospho-regulation: GlcNAc-6P/AMPK phosphorylation of GSDME at Thr6 blocks caspase-3 cleavage, repressing pyroptosis.\",\n      \"evidence\": \"Metabolite-AMPK binding, AMPK-knockout and GSDME T6E/T6A knock-in mice with in vitro cleavage assays\",\n      \"pmids\": [\"37460805\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of how Thr6 phosphorylation occludes the Asp270 site unresolved\", \"Generality across cell types untested\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Extended transcriptional and functional reach to macrophage biology and atherosclerosis via STAT3-driven GSDME expression.\",\n      \"evidence\": \"GSDME/ApoE double-knockout mice, ox-LDL macrophage treatment and STAT3 functional analysis\",\n      \"pmids\": [\"36807553\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct STAT3 promoter binding not shown here\", \"Cell-autonomy vs. systemic effects partly conflated\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Defined a cleavage-independent activation route, showing PARP1/PARP5 PARylation of full-length GSDME relieves autoinhibition and couples to lipid-ROS sensing for oligomerization.\",\n      \"evidence\": \"UV-C irradiation with PARP1/PARP5 inhibition/knockout, PARylation, lipid-ROS, oligomerization and membrane-targeting assays\",\n      \"pmids\": [\"38997456\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"PARylation site(s) on GSDME not mapped\", \"How lipid ROS is structurally sensed unresolved\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Identified flotillin-2 as the membrane-recruitment partner for GSDME-N and extended pore function to platelet hyperactivity and thrombosis.\",\n      \"evidence\": \"Human and GSDME-knockout mouse platelets, cleavage and pore assays, Co-IP and thrombosis models\",\n      \"pmids\": [\"39378585\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of flotillin-2/GSDME-N interaction unknown\", \"Whether flotillin-2 recruitment generalizes to other cell types untested\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Revealed a non-pore-forming signaling role: GSDME-C binds PDPK1 to activate PI3K-AKT and promote M2-like macrophage polarization.\",\n      \"evidence\": \"Co-IP, Eliprodil inhibition, single-cell sequencing and in vivo HCC models\",\n      \"pmids\": [\"39496854\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural interface of GSDME-C/PDPK1 undefined\", \"Balance between pyroptotic and signaling roles in vivo unclear\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Added further regulatory and downstream nodes: Sp1 promoter activation, ALKBH4-controlled H3K4me3 repression, p53/ULK1 control, and a GSDME-dependent Drp1-R114 citrullination/mitophagy axis.\",\n      \"evidence\": \"ChIP/reporter assays, ALKBH4 and ULK1 manipulation, GSDME-knockout/reintroduction and Drp1-R114 mutagenesis across cancer and MASH models\",\n      \"pmids\": [\"38238307\", \"38902235\", \"39215364\", \"39009654\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism linking GSDME to Drp1 citrullination not fully defined\", \"Integration of these regulators into a single hierarchy untested\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Showed GSDME is the lytic effector arm of cytotoxic-lymphocyte/IFNγ antitumor immunity, with STAT1-GSDME circuitry required for HDAC-inhibitor efficacy.\",\n      \"evidence\": \"ChIP-seq, single-cell multiomics, genetic GSDME/STAT1 knockout and orthotopic HCC models\",\n      \"pmids\": [\"39486886\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Which protease (GZMB vs caspase-3) dominates cleavage in this setting partly open\", \"Direct STAT1 regulation of GSDME locus vs. immune-cell recruitment effects entangled\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How the multiple activation inputs (caspase-3 cleavage, PARylation, lipid-ROS sensing) and antagonistic post-translational marks (Thr6 phosphorylation, palmitoylation, ubiquitination, O-glycosylation) are integrated at the structural level to set the threshold between apoptosis and pyroptosis remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No high-resolution structure of activated GSDME-N pore or of the autoinhibited full-length protein in the corpus\", \"PARylation and modification sites not jointly mapped\", \"Quantitative model of competing modifications absent\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [0, 11, 17]},\n      {\"term_id\": \"GO:0008289\", \"supporting_discovery_ids\": [10]},\n      {\"term_id\": \"GO:0140313\", \"supporting_discovery_ids\": [2]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [0, 11, 17, 10]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [10]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [0, 22, 25]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [13, 16, 19]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"ZDHHC2\", \"ZDHHC7\", \"OTUD4\", \"CDC20\", \"PDPK1\", \"FLOT2\", \"PARP1\", \"AMPK\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":8,"faith_total":9,"faith_pct":88.88888888888889}}