{"gene":"EHF","run_date":"2026-04-28T17:46:03","timeline":{"discoveries":[{"year":2000,"finding":"EHF (ESE-3) is an epithelium-specific ETS transcription factor that transactivates the c-MET promoter via three high-affinity binding sites, and shows different promoter transactivation specificity from ESE-1 despite similar DNA-binding affinity, indicating context-dependent transcriptional activity.","method":"Transient transfection reporter assays, DNA-binding/EMSA, promoter analysis","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 — in vitro promoter assays with mutagenesis and binding studies, foundational characterization paper","pmids":["10644770"],"is_preprint":false},{"year":2001,"finding":"EHF (ESE-3) acts as a context-dependent transcriptional repressor of Ras/phorbol ester-induced promoters containing ETS and AP-1 binding sites, repressing MAPK signaling-driven transcription in a sequence- and arrangement-dependent manner; it is a nuclear protein expressed exclusively in differentiated epithelial cells and absent in epithelial carcinomas.","method":"Transient transfection reporter assays, immunohistochemistry with monoclonal antibody, promoter mutagenesis","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 — reconstituted promoter repression in multiple cell types with defined cis-elements, replicated in foundational study","pmids":["11259407"],"is_preprint":false},{"year":1999,"finding":"EHF protein represses the ETS-2-induced transcriptional activity of stromelysin-1 and collagenase-1 promoters, demonstrating transcriptional repressor activity on matrix metalloproteinase promoters.","method":"Transient transfection reporter assays","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 1 — reporter assay with defined promoters, single lab single method","pmids":["10527851"],"is_preprint":false},{"year":2002,"finding":"ESE-3/EHF cytokine-induced expression in bronchial smooth muscle cells is mediated through MEK1/2 and p38 MAPK signaling pathways, and overexpression of ESE-3 inhibits MMP-1 promoter activity, indicating it acts as a transcriptional repressor downstream of these kinases.","method":"Pharmacological inhibitors (U0126, SB03580), reporter assays, RT-PCR, Western blot","journal":"American journal of respiratory cell and molecular biology","confidence":"Medium","confidence_rationale":"Tier 2 — pathway placement via pharmacological epistasis plus reporter assay","pmids":["12444029"],"is_preprint":false},{"year":2007,"finding":"EHF (ESE-3) re-expression in prostate cancer cells inhibits clonogenic survival and induces apoptosis by directly binding the caspase-3 (CASP3) promoter and increasing procaspase-3 levels, acting as a tumor suppressor; ESE-3 silencing in tumorigenic cells is mediated by CpG methylation of an evolutionarily conserved site in its promoter.","method":"Chromatin immunoprecipitation, reporter assay, re-expression experiments, 5-aza-2'-deoxycytidine treatment, colony assay, apoptosis assay","journal":"Oncogene","confidence":"High","confidence_rationale":"Tier 1-2 — ChIP-confirmed direct promoter binding, functional rescue, epigenetic mechanism, multiple orthogonal methods","pmids":["18037958"],"is_preprint":false},{"year":2007,"finding":"ESE-3/EHF is upregulated in cellular senescence downstream of p38 MAPK signaling; ectopic expression induces growth retardation, increased SA-β-gal activity, and upregulation of p16(INK4a) via direct binding of recombinant ESE-3 to ETS-binding sequences in the p16(INK4a) promoter.","method":"Microarray, reporter assay, recombinant protein EMSA, SA-β-gal assay, Western blot","journal":"Cancer science","confidence":"High","confidence_rationale":"Tier 1 — in vitro binding confirmed by EMSA with recombinant protein plus functional reporter and cellular phenotype","pmids":["17627613"],"is_preprint":false},{"year":2008,"finding":"EHF/ESE-3 expression in airway epithelial cells is upregulated by IL-1β and TNF-α via NF-κB activation; NF-κB binding sequences in the ESE-3 promoter are required for cytokine-induced expression, and ESE-1 further upregulates ESE-3 expression.","method":"Promoter characterization, NF-κB binding site mutagenesis, reporter assays, RT-PCR","journal":"Cell research","confidence":"High","confidence_rationale":"Tier 1 — promoter mutagenesis identifying required cis-elements plus pathway placement via NF-κB","pmids":["18475289"],"is_preprint":false},{"year":2011,"finding":"EHF directly activates transcription of RUVBL1, an ATPase associated with chromatin-remodeling complexes; RUVBL1 in turn represses p53 and its target genes by binding the p53 promoter, interfering with RNF20/hBRE1-mediated histone H2B monoubiquitination and promoting PAF1-mediated H3K9 trimethylation, thereby blocking p53-mediated apoptosis in colon tumor cells.","method":"Chromatin immunoprecipitation, reporter assays, siRNA knockdown, histone modification analysis, apoptosis assays","journal":"EMBO reports","confidence":"High","confidence_rationale":"Tier 1-2 — ChIP-confirmed direct binding, histone modification mechanistic dissection, multiple orthogonal methods","pmids":["21617703"],"is_preprint":false},{"year":2012,"finding":"ESE3/EHF represses transcription of key EMT and cancer stem cell genes including TWIST1, ZEB2, BMI1, and POU5F1 in prostate epithelial cells; loss of ESE3/EHF induces EMT, stem-like features, and tumor-initiating properties, which are reversed by re-expression of ESE3/EHF.","method":"Loss-of-function knockdown, re-expression, gene expression analysis, in vitro and in vivo tumor assays","journal":"Cancer research","confidence":"High","confidence_rationale":"Tier 2 — loss-of-function with defined gene targets, rescue experiments, in vivo validation","pmids":["22505649"],"is_preprint":false},{"year":2013,"finding":"EHF promotes cornea epithelial fate through complementary gene-activating and -repressing activities, and potential interactions with KLF4 and KLF5 were identified; EHF occupancy at target gene loci was determined by ChIP-seq.","method":"Loss-of-function studies, ChIP-seq, transcriptome profiling","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1-2 — ChIP-seq genome-wide occupancy plus loss-of-function phenotype in cornea epithelium","pmids":["24142692"],"is_preprint":false},{"year":2013,"finding":"Ectopic expression of EHF in intestinal epithelial cells activates HCK-dependent apical-to-basolateral transcytosis of non-opsonized and SIgA-opsonized particles in a follicle-associated epithelium model, placing EHF upstream of HCK in regulating mucosal antigen sampling.","method":"Ectopic expression, transcytosis assays, kinase inhibitor studies","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 — gain-of-function with defined cellular phenotype and pathway placement, single lab","pmids":["23439650"],"is_preprint":false},{"year":2015,"finding":"EHF is upregulated by TGF-β/Smad signaling in mast cells; forced EHF expression represses transcription of FcεRIα, FcεRIβ, and c-Kit genes by directly binding their promoters, reducing surface FcεRI and c-Kit, suppressing degranulation and cytokine production, and decreasing GATA1, GATA2, and Stat5b expression.","method":"Forced expression, promoter-binding assays (ChIP), reporter assays, flow cytometry, degranulation assays","journal":"Journal of immunology","confidence":"High","confidence_rationale":"Tier 1-2 — ChIP-confirmed direct promoter binding, multiple downstream targets verified, functional cellular consequences","pmids":["26297757"],"is_preprint":false},{"year":2016,"finding":"ESE3/EHF binds and represses the Lin28A and Lin28B gene promoters while activating transcription and maturation of let-7 microRNAs in normal prostate cells; loss of ESE3/EHF upregulates Lin28A/B and downregulates let-7, driving prostate cancer stem cell expansion.","method":"ChIP, promoter reporter assays, microRNA profiling, siRNA knockdown, xenograft models","journal":"Cancer research","confidence":"High","confidence_rationale":"Tier 1-2 — ChIP-confirmed direct promoter binding of Lin28A/B, functional validation in vitro and in vivo","pmids":["27197175"],"is_preprint":false},{"year":2016,"finding":"ESE3/EHF directly binds a novel ETS binding site in the IL-6 gene promoter and represses IL-6 transcription; loss of ESE3/EHF activates IL-6/JAK/STAT3 signaling, promoting stem-like transformation in prostate epithelial cells.","method":"ChIP, reporter assays, siRNA knockdown, IL-6 inhibition rescue experiments","journal":"Oncotarget","confidence":"High","confidence_rationale":"Tier 1-2 — ChIP-confirmed direct binding, epistasis rescue by IL-6 inhibition, multiple methods","pmids":["27732936"],"is_preprint":false},{"year":2016,"finding":"ESE3/EHF inhibits pancreatic cancer metastasis by directly upregulating E-cadherin transcription; knockdown of ESE3 promotes cell motility and invasiveness in vitro and metastasis in an orthotopic mouse model.","method":"Reporter assays, ChIP, siRNA knockdown, orthotopic mouse model, immunohistochemistry","journal":"Cancer research","confidence":"High","confidence_rationale":"Tier 1-2 — ChIP-confirmed direct transcriptional regulation of E-cadherin, in vivo validation","pmids":["27923832"],"is_preprint":false},{"year":2016,"finding":"EHF transcriptionally regulates HER2 (ERBB2) and HER3 (ERBB3) in thyroid cancer cells, contributing to PI3K/Akt and MAPK/Erk pathway activation; this was confirmed by dual-luciferase reporter and ChIP assays.","method":"ChIP, dual-luciferase reporter assay, siRNA knockdown, ectopic expression, in vivo xenograft","journal":"Oncotarget","confidence":"High","confidence_rationale":"Tier 1-2 — ChIP plus reporter assay confirming direct transcriptional regulation, pathway placement","pmids":["27517321"],"is_preprint":false},{"year":2017,"finding":"EHF ChIP-seq in primary human bronchial epithelial cells shows enrichment of EHF targets in inflammation and wound repair genes; EHF depletion alters epithelial secretion of a neutrophil chemokine, slows wound closure, and EHF activates SPDEF expression contributing to goblet cell hyperplasia.","method":"ChIP-seq, RNA-seq after EHF depletion, wound closure assay, cytokine secretion assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1-2 — genome-wide ChIP-seq combined with RNA-seq and functional cellular phenotypes","pmids":["28461336"],"is_preprint":false},{"year":2019,"finding":"miR-365-3p targets EHF 3'UTR to decrease EHF protein expression; EHF acts as a transcription factor for KRT16 in oral squamous cell carcinoma, and EHF-driven KRT16 expression promotes β5-integrin/c-Met signaling and downstream Src/STAT3/FAK/ERK pathway activation.","method":"Dual-luciferase reporter (miRNA targeting), ectopic expression, siRNA knockdown, confocal colocalization, in vitro/in vivo models","journal":"Journal of experimental & clinical cancer research","confidence":"High","confidence_rationale":"Tier 1-2 — validated miRNA-EHF-KRT16 axis with multiple orthogonal methods and in vivo validation","pmids":["30782177"],"is_preprint":false},{"year":2020,"finding":"EHF directly upregulates TGF-β1 transcription in colorectal cancer cells, thereby activating canonical TGF-β signaling to promote tumor cell proliferation, migration, and invasion.","method":"Reporter assays, ChIP, siRNA knockdown, ectopic expression, in vitro and in vivo functional assays","journal":"Cancer science","confidence":"High","confidence_rationale":"Tier 1-2 — ChIP plus reporter assay confirming direct TGF-β1 transcriptional activation","pmids":["32372436"],"is_preprint":false},{"year":2021,"finding":"EHF produces two transcript variants (EHF-LF including exon 1, EHF-SF excluding exon 1); only EHF-SF abrogates ETS1-mediated activation of the ZEB1 promoter by promoting degradation of ETS1 protein, thereby inhibiting EMT. A point mutation in the EHF ETS domain abolishes this function and creates a dominant-negative form that enhances metastasis in vivo.","method":"Reporter assays, protein degradation assays, in vivo metastasis models, mutagenesis","journal":"Oncogenesis","confidence":"High","confidence_rationale":"Tier 1-2 — isoform-specific function with mutagenesis, mechanistic dissection of ETS1 degradation, in vivo validation","pmids":["33712555"],"is_preprint":false},{"year":2021,"finding":"EHF suppresses pancreatic cancer stemness by directly repressing CXCR4 transcription, decreasing cancer cell sensitivity to CXCL12 from pancreatic stellate cells; EHF also has a cell-autonomous role repressing Sox9, Sox2, Oct4, and Nanog transcription. Rosiglitazone upregulates EHF to suppress cancer stemness.","method":"ChIP, luciferase reporter assays, sphere formation assays, flow cytometry, in vivo KPC mouse model","journal":"Gut","confidence":"High","confidence_rationale":"Tier 1-2 — ChIP-confirmed direct CXCR4 repression, multiple stemness targets, validated in KPC mice","pmids":["33674341"],"is_preprint":false},{"year":2021,"finding":"Loss of EHF promotes neuroendocrine differentiation in prostate cancer via ADT/AR/EHF/EZH2 signaling: androgen deprivation reduces EHF expression by relieving AR binding to androgen-responsive elements in the EHF locus, which promotes EZH2 expression and enzymatic activity, leading to H3K27me3-mediated transcriptional repression of downstream genes.","method":"ChIP, siRNA/shRNA knockdown, EZH2 inhibitor, cell and mouse models, Western blot","journal":"Cell death & disease","confidence":"High","confidence_rationale":"Tier 2 — ChIP-confirmed AR binding and H3K27me3 mechanism, pathway epistasis with EZH2 inhibitor","pmids":["33414441"],"is_preprint":false},{"year":2021,"finding":"EBV protein LMP2A causes upregulation of EHF via phosphorylation of STAT3 in gastric cancer cells; EHF knockdown inhibits cell proliferation and regulates cancer pathway genes including FZD5; EHF motif is enriched in activated enhancers in EBV-positive gastric cancer.","method":"STAT3 knockdown rescue by EHF overexpression, genome-wide active histone ChIP-seq, siRNA knockdown, immunostaining","journal":"Cancer science","confidence":"Medium","confidence_rationale":"Tier 2 — functional epistasis (STAT3 KD rescued by EHF OE), upstream regulation mechanism, single lab","pmids":["34014591"],"is_preprint":false},{"year":2021,"finding":"Ese-3/EHF promotes colon cancer cell proliferation by downregulating EHD2 and transactivating INPP4B as direct downstream target genes.","method":"siRNA knockdown, ectopic expression, in vitro/in vivo proliferation assays","journal":"American journal of cancer research","confidence":"Medium","confidence_rationale":"Tier 2-3 — identified downstream targets with functional assays, limited mechanistic detail on direct binding","pmids":["33520362"],"is_preprint":false},{"year":2022,"finding":"EHF physically interacts with CDX1 via its PNT domain; EHF and CDX1 co-operatively drive transcription of the colonic differentiation marker VIL1; compound deletion of Ehf and Cdx1 in mouse colon disrupts normal colonic differentiation and enhances colorectal tumor progression. Re-expression of EHF and CDX1 induces extensive chromatin remodeling and transcriptional reprogramming.","method":"Co-immunoprecipitation (PNT domain interaction), reporter assays, CRISPR knockout mouse model, ATAC-seq, RNA-seq","journal":"Cell death and differentiation","confidence":"High","confidence_rationale":"Tier 1-2 — physical interaction confirmed by Co-IP with domain mapping, in vivo mouse model, genome-wide chromatin remodeling","pmids":["35606410"],"is_preprint":false},{"year":2021,"finding":"Deletion of the EHF ETS DNA-binding domain in mice causes epidermal papillomas, preputial/vulval abscesses, corneal ulcers, increased susceptibility to colitis, impaired goblet cell differentiation, and enhanced Apc-initiated adenoma development, demonstrating that the DNA-binding domain is essential for postnatal epithelial homeostasis.","method":"Novel Ehf knockout mouse strains (Ehf-/- and gut-specific), histopathology, experimental colitis model, transcriptomic profiling","journal":"Development (Cambridge, England)","confidence":"High","confidence_rationale":"Tier 2 — clean genetic knockout with multiple defined tissue phenotypes, in vivo tumor model","pmids":["34180969"],"is_preprint":false},{"year":2024,"finding":"EHF deficiency in pancreatic cancer induces CXCL1 transcription, enhancing CXCR2+ neutrophil recruitment in a CXCL1-CXCR2-dependent manner; TP53 mutation-mediated loss of EHF drives this process. Nifurtimox elevates tumoral EHF and inhibits JAK1/STAT1 pathway, suppressing CXCR2+ neutrophil recruitment.","method":"ChIP assay, Ehf-knockout mice, syngeneic mouse models, multiplexed cytokine assay, flow cytometry, single-cell RNA-seq","journal":"Gastroenterology","confidence":"High","confidence_rationale":"Tier 1-2 — ChIP-confirmed CXCL1 transcriptional regulation, in vivo genetic models, rescue by CXCL1/CXCR2 blockade","pmids":["38492894"],"is_preprint":false},{"year":2024,"finding":"Dclk2 phosphorylates EHF, changing its nucleoplasmic distribution by causing p-EHF to exit the nucleus, thereby decreasing nuclear EHF levels; nuclear EHF binds the promoters of Caspase1 and Caspase3 to reduce their transcription and inhibit neuronal pyroptosis. Cpeb4 upregulates Dclk2 by increasing Dclk2 mRNA stability.","method":"Promoter binding assays, ChIP, co-immunoprecipitation, immunofluorescence for nucleoplasmic distribution, OGD cell model, CCI mouse model, knockdown experiments","journal":"Journal of cerebral blood flow and metabolism","confidence":"High","confidence_rationale":"Tier 1-2 — kinase-substrate relationship confirmed, subcellular relocalization mechanism defined, direct promoter binding shown by ChIP","pmids":["38513137"],"is_preprint":false},{"year":2024,"finding":"EHF transcriptionally activates GLI1 and CCL2 in cholangiocarcinoma; EHF-driven CCL2 recruits and activates tumor-associated macrophages via the CCL2/CCR2 axis, remodeling the tumor microenvironment.","method":"ChIP, reporter assays, in vitro and in vivo functional experiments, immunohistochemistry","journal":"MedComm","confidence":"High","confidence_rationale":"Tier 1-2 — ChIP-confirmed direct transcriptional activation of GLI1 and CCL2, in vivo validation","pmids":["38741887"],"is_preprint":false},{"year":2024,"finding":"EHF interacts with the coactivator AJUBA to cooperatively orchestrate transcriptional network activity in gastroesophageal adenocarcinoma; EHF expression is promoted by a core transcriptional regulatory circuitry (ELF3-KLF5-GATA6); KRAS signaling is a common downstream pathway of EHF and AJUBA.","method":"Co-immunoprecipitation (EHF-AJUBA interaction), ChIP, reporter assays, in vitro and in vivo functional experiments","journal":"Acta pharmaceutica sinica. B","confidence":"Medium","confidence_rationale":"Tier 2 — Co-IP confirmed protein interaction with coactivator, upstream CRC regulation characterized","pmids":["38799645"],"is_preprint":false},{"year":2024,"finding":"EHF forms liquid-like nuclear condensates (phase separation) that transcriptionally repress TERT and inflammatory factors (IL-6, CXCL12); reduction of TERT leads to telomere shortening and cellular senescence in PDAC without the senescence-associated secretory phenotype (SASP). Bilobetin promotes EHF phase separation.","method":"CRISPR/Cas9 library screening, phase separation assays, ChIP, telomere length assays, drug screening, in vivo experiments","journal":"Cancer letters","confidence":"Medium","confidence_rationale":"Tier 2 — phase separation characterized with functional transcriptional repression and cellular phenotype, novel mechanistic finding","pmids":["39710057"],"is_preprint":false},{"year":2024,"finding":"Loss of Ehf in the mouse mammary gland impairs lobuloalveolar differentiation at late pregnancy, reduces milk gene/lipid levels, and causes accumulation of alveolar progenitor cells; Ehf deletion attenuates prolactin-induced alveolar differentiation in mammary organoids and increases tumor incidence in the MMTV-PyMT model.","method":"Conditional Ehf knockout mouse model, mammary organoids, in vivo tumor model, gene expression analysis","journal":"Developmental cell","confidence":"High","confidence_rationale":"Tier 2 — clean genetic knockout with defined differentiation and tumor suppressor phenotypes in vivo and ex vivo","pmids":["38781975"],"is_preprint":false},{"year":2025,"finding":"Loss of EHF in human iPSC-derived lung cells enhances CFTR activity, increases transepithelial electrical resistance, causes transcriptomic changes in basal cells, and reduces HIF-1α-mediated hypoxic response, indicating EHF controls multiple lung epithelial functions including ion channel regulation and hypoxia response.","method":"CRISPR EHF knockout in iPSC-derived lung cells, electrophysiology, RNA-seq, HIF-1α pathway analysis","journal":"Disease models & mechanisms","confidence":"Medium","confidence_rationale":"Tier 2 — clean genetic knockout with multiple functional readouts in human iPSC model","pmids":["40590703"],"is_preprint":false},{"year":2022,"finding":"Ehf plays a non-redundant role in salivary gland ductal cell differentiation; CRISPR-Cas9 disruption of the Ehf ETS domain causes decreased granular convoluted tubules and increased intercalated Sox9-positive ductal cell accumulation, with a pronounced sexual dimorphism in males.","method":"CRISPR-Cas9 Ehf ETS domain knockout mouse, histology, immunostaining","journal":"Journal of dental research","confidence":"Medium","confidence_rationale":"Tier 2 — in vivo genetic loss-of-function with defined cellular differentiation phenotype","pmids":["36348499"],"is_preprint":false},{"year":2006,"finding":"ESE-3/EHF regulates DR-5 (death receptor 5) expression through direct binding to putative ETS sites on the DR-5 promoter; co-factors CBP and p300 are involved in ESE-3-mediated DR-5 upregulation.","method":"EMSA, luciferase reporter assay, co-factor interaction analysis","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 1 — in vitro EMSA and reporter assay with defined ETS sites, identified co-factors","pmids":["17027647"],"is_preprint":false},{"year":2016,"finding":"ESE-3/EHF cooperates with pregnane X receptor (PXR) to transactivate the ABCB1 gene in intestinal epithelial cells; ESE-3 directly binds the DR4-containing distal enhancer module of ABCB1 as shown by ChIP, and this interaction is required for rifampicin-induced ABCB1 expression.","method":"ChIP, reporter assays with DR4 mutation, siRNA knockdown, RT-PCR","journal":"Drug metabolism and pharmacokinetics","confidence":"High","confidence_rationale":"Tier 1-2 — ChIP-confirmed direct binding, mutagenesis of DR4 sites, siRNA rescue, cell-type-specific mechanism","pmids":["27567379"],"is_preprint":false},{"year":2023,"finding":"RRAD binds and regulates the subcellular localization of the transcription factor EHF in papillary thyroid cancer; EHF activates transcription of NEAT1_2, HK2, and PKM2, forming a NEAT1_2/RRAD/EHF positive feedback loop that facilitates aerobic glycolysis.","method":"Co-immunoprecipitation (RRAD-EHF), ChIP, luciferase reporter assays, RNA binding protein IP, subcellular fractionation","journal":"Endocrinology","confidence":"Medium","confidence_rationale":"Tier 2 — Co-IP confirms protein interaction, ChIP confirms direct transcriptional targets, subcellular localization linked to function","pmids":["37279586"],"is_preprint":false},{"year":2025,"finding":"EHF in M2 tumor-associated macrophages promotes liver cancer cell metastasis by binding the KDM2B promoter and activating its transcription, which leads to increased IL-6 secretion.","method":"CUT-Tag, ChIP, luciferase assay, siRNA knockdown, co-culture migration/invasion assay, in vivo","journal":"Cellular signalling","confidence":"Medium","confidence_rationale":"Tier 1-2 — CUT-Tag and ChIP confirm direct KDM2B promoter binding, functional downstream IL-6 pathway","pmids":["39971220"],"is_preprint":false},{"year":2026,"finding":"EHF regulates conventional dendritic cell maturation and immunosuppression after TLR7/8/9 stimulation by directly regulating Ccr7, Cd200, Cd274 (PD-L1), Irf4, and Rel expression, as shown by CUT&TAG; EHF-deficient DCs promote Th1/Th17-biased CD4+ T cell responses.","method":"Conditional DC-specific Ehf knockout mice, CUT&TAG, single-cell RNA-seq, T cell polarization assays, in vivo autoimmune/tumor models","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1-2 — CUT&TAG genome-wide direct target identification, conditional knockout with multiple defined immune phenotypes","pmids":["41730908"],"is_preprint":false},{"year":2025,"finding":"EHF in lung cancer cell-derived exosomes is transferred to macrophages and promotes M2 polarization by binding the RNF41 promoter and activating its transcription; inhibition of EHF/RNF41 axis suppresses tumor growth in vivo.","method":"ChIP, dual-luciferase reporter, exosome isolation and transfer assay, TEM, nanoparticle tracking, flow cytometry, in vivo xenograft","journal":"Regenerative therapy","confidence":"Medium","confidence_rationale":"Tier 2 — ChIP and reporter confirm direct RNF41 transcriptional regulation, exosome transfer mechanism","pmids":["41939996"],"is_preprint":false}],"current_model":"EHF is an epithelium-specific ETS family transcription factor that functions primarily as a context-dependent transcriptional activator or repressor — directly binding ETS/AP-1 composite elements in target promoters to repress MAPK-driven transcription and maintain epithelial differentiation; it controls a broad network of targets including E-cadherin, Lin28A/B, IL-6, CXCR4, CXCL1, caspase-3, p16(INK4a), RUVBL1, FcεRI subunits, SPDEF, TGF-β1, HER2/HER3, ZEB1/2 (via ETS1 degradation), and TERT, and its activity is regulated post-translationally by Dclk2-mediated phosphorylation (causing nuclear export), by epigenetic silencing via CpG methylation, by upstream signals including TGF-β/Smad, NF-κB, STAT3, and AR/EZH2 axes, and by physical interaction with co-factors including CDX1 (via PNT domain) and AJUBA; EHF can also form nuclear liquid-like condensates that influence its transcriptional repressor activity."},"narrative":{"teleology":[{"year":2000,"claim":"Identification of EHF as an epithelium-restricted ETS transcription factor with context-dependent transactivation activity established the foundational molecular identity and tissue specificity of the gene.","evidence":"Reporter assays and EMSA on c-MET promoter in transfected cells","pmids":["10644770","10527851"],"confidence":"High","gaps":["No endogenous target genes confirmed by ChIP at this stage","Repressor vs. activator specificity determinants unknown","No loss-of-function data"]},{"year":2001,"claim":"Demonstrating that EHF represses Ras/MAPK-driven transcription through ETS/AP-1 composite elements and is absent in carcinomas established its dual activator-repressor nature and implicated it as a tumor suppressor.","evidence":"Promoter mutagenesis reporter assays in multiple cell lines plus immunohistochemistry on normal vs. tumor tissue","pmids":["11259407"],"confidence":"High","gaps":["No direct in vivo tumor suppressor evidence","Mechanism of repression (cofactor recruitment vs. competition) undefined"]},{"year":2007,"claim":"Discovery that EHF directly activates the caspase-3 promoter to induce apoptosis and that its own expression is silenced by CpG methylation in tumors provided the first mechanistic link between EHF loss and cancer cell survival.","evidence":"ChIP on CASP3 promoter, 5-aza-2'-deoxycytidine demethylation rescue, colony and apoptosis assays in prostate cancer cells","pmids":["18037958","17627613"],"confidence":"High","gaps":["Genome-wide target repertoire unknown","Which methyltransferases silence EHF not identified","Senescence-related p16 activation mechanism not distinguished from apoptosis pathway"]},{"year":2008,"claim":"Identification of NF-κB as a direct upstream activator of EHF transcription placed EHF within inflammatory signaling cascades in airway epithelium.","evidence":"NF-κB binding site mutagenesis in the EHF promoter plus cytokine stimulation in bronchial epithelial cells","pmids":["18475289"],"confidence":"High","gaps":["Whether NF-κB regulation is tissue-general or airway-specific not resolved","MAPK vs. NF-κB pathway hierarchy for EHF regulation unclear"]},{"year":2012,"claim":"Showing that EHF loss induces EMT and stem-like features by derepressing TWIST1, ZEB2, BMI1, and POU5F1 unified EHF's tumor-suppressive role with epithelial differentiation maintenance.","evidence":"Loss-of-function knockdown with rescue, gene expression analysis, in vivo tumor-initiating assays in prostate cells","pmids":["22505649"],"confidence":"High","gaps":["Direct ChIP binding to each EMT promoter not confirmed for all targets","Isoform-specific contributions not dissected"]},{"year":2013,"claim":"Genome-wide ChIP-seq in corneal epithelium revealed that EHF directly occupies hundreds of epithelial gene loci with both activating and repressive outputs, establishing the breadth of its transcriptional regulatory network.","evidence":"ChIP-seq combined with transcriptome profiling and loss-of-function in human corneal epithelial cells","pmids":["24142692"],"confidence":"High","gaps":["Cofactor requirements for activation vs. repression not defined genome-wide","Overlap of targets across different epithelial tissues not compared"]},{"year":2016,"claim":"Multiple studies converged to show EHF directly controls E-cadherin, Lin28A/B-let-7, IL-6/STAT3, HER2/HER3, and ABCB1 promoters, revealing it as a multi-target hub linking differentiation, stemness, inflammation, and drug metabolism.","evidence":"ChIP-confirmed promoter binding with reporter assays in prostate, pancreatic, thyroid, and intestinal cells; in vivo xenograft validation","pmids":["27923832","27197175","27517321","27567379","27732936"],"confidence":"High","gaps":["How EHF selects among different targets in different tissues not resolved","Structural basis of promoter recognition beyond ETS motif unknown"]},{"year":2017,"claim":"ChIP-seq in bronchial epithelium extended the target repertoire to wound repair and inflammation genes and showed EHF activates SPDEF to drive goblet cell differentiation, linking EHF to mucosal defense.","evidence":"ChIP-seq and RNA-seq after EHF depletion in primary human bronchial epithelial cells, wound closure assay","pmids":["28461336"],"confidence":"High","gaps":["Whether EHF directly interacts with SPDEF protein or only activates its transcription unclear","In vivo airway phenotype of EHF loss not yet tested"]},{"year":2021,"claim":"Genetic deletion of Ehf in mice produced papillomas, corneal ulcers, colitis susceptibility, goblet cell loss, and enhanced adenoma development, providing definitive in vivo proof that the ETS DNA-binding domain is essential for epithelial homeostasis and tumor suppression.","evidence":"Novel Ehf knockout mouse strains with histopathology, experimental colitis, and Apc-initiated tumor model","pmids":["34180969"],"confidence":"High","gaps":["Tissue-specific vs. systemic contributions not fully dissected with conditional knockouts at this stage","Whether observed phenotypes are cell-autonomous in all tissues not confirmed"]},{"year":2021,"claim":"Discovery that the short EHF isoform (EHF-SF) specifically promotes ETS1 protein degradation to block ZEB1 transcription, while a point mutant acts as a dominant-negative to enhance metastasis, revealed isoform-specific and gain-of-function mechanisms.","evidence":"Reporter assays, protein degradation assays, mutagenesis, in vivo metastasis models","pmids":["33712555"],"confidence":"High","gaps":["Proteasomal vs. lysosomal degradation pathway for ETS1 not specified","Relative abundance and regulation of EHF-LF vs. EHF-SF isoforms in normal tissues unknown"]},{"year":2021,"claim":"Placement of EHF downstream of AR signaling showed that androgen deprivation silences EHF, derepressing EZH2 and promoting neuroendocrine differentiation via H3K27me3, linking EHF loss to treatment-resistant prostate cancer.","evidence":"ChIP for AR binding at EHF locus, EZH2 inhibitor epistasis, mouse and cell models","pmids":["33414441"],"confidence":"High","gaps":["Whether EHF directly binds the EZH2 promoter or acts indirectly not resolved","Clinical correlation with neuroendocrine differentiation markers not fully established"]},{"year":2022,"claim":"EHF's physical interaction with CDX1 via its PNT domain and their cooperative induction of chromatin remodeling established that EHF functions through cofactor partnerships to drive colonic differentiation, not solely through autonomous DNA binding.","evidence":"Co-immunoprecipitation with domain mapping, CRISPR compound knockout mice, ATAC-seq and RNA-seq","pmids":["35606410"],"confidence":"High","gaps":["Whether PNT domain mediates additional cofactor interactions not explored","Structural basis of EHF-CDX1 interaction unknown"]},{"year":2024,"claim":"Identification of Dclk2-mediated phosphorylation of EHF causing nuclear export established the first post-translational mechanism controlling EHF subcellular distribution and transcriptional output.","evidence":"Co-IP, immunofluorescence for nucleoplasmic distribution, ChIP on caspase promoters, OGD/CCI models","pmids":["38513137"],"confidence":"High","gaps":["Phosphorylation site(s) on EHF not mapped","Whether this kinase-substrate relationship operates in epithelial tissues not tested"]},{"year":2024,"claim":"EHF deficiency was shown to upregulate CXCL1, recruiting CXCR2+ neutrophils and reshaping the pancreatic tumor microenvironment — extending EHF's tumor-suppressive role from cell-autonomous to immune-regulatory.","evidence":"ChIP, Ehf-knockout mice, syngeneic models, single-cell RNA-seq, CXCL1/CXCR2 blockade rescue","pmids":["38492894"],"confidence":"High","gaps":["Whether EHF regulates additional chemokines systemically not surveyed","Human clinical validation of neutrophil recruitment phenotype lacking"]},{"year":2024,"claim":"Discovery that EHF forms liquid-like nuclear condensates that repress TERT, IL-6, and CXCL12 introduced phase separation as a mechanism for EHF-mediated transcriptional repression and senescence induction without SASP.","evidence":"CRISPR library screening, phase separation assays, ChIP, telomere length measurement, bilobetin drug screen","pmids":["39710057"],"confidence":"Medium","gaps":["Biophysical parameters of EHF condensates (saturation concentration, IDR contributions) not characterized","Whether condensate formation occurs in normal epithelial cells or is cancer-specific unknown","Independent replication needed"]},{"year":2025,"claim":"EHF was shown to regulate dendritic cell maturation by directly controlling Ccr7, PD-L1, Irf4, and Rel expression, demonstrating a non-epithelial immune cell-autonomous role in shaping adaptive immune responses.","evidence":"Conditional DC-specific Ehf knockout mice, CUT&TAG, single-cell RNA-seq, T cell polarization assays, autoimmune/tumor models","pmids":["41730908"],"confidence":"High","gaps":["Whether EHF functions in other immune cell lineages not tested","Human DC relevance not confirmed"]},{"year":null,"claim":"Key unresolved questions include the structural basis of EHF's activator-repressor switch, the full spectrum of phosphorylation sites governing its nuclear-cytoplasmic shuttling, whether phase separation is a general EHF mechanism in normal epithelia, and the complete set of PNT domain-mediated cofactor interactions.","evidence":"","pmids":[],"confidence":"Low","gaps":["No crystal or cryo-EM structure of EHF or its complexes","Systematic mapping of EHF post-translational modifications not performed","Tissue-specific isoform expression and regulation not comprehensively characterized"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[0,1,4,5,7,8,9,11,12,13,14,15,16,18,20,24,26,28,30,35,38]},{"term_id":"GO:0003677","term_label":"DNA binding","supporting_discovery_ids":[0,1,2,4,5,9,11,12,13,14,16,34,38]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[1,27,30]},{"term_id":"GO:0005654","term_label":"nucleoplasm","supporting_discovery_ids":[27,30]}],"pathway":[{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[0,1,4,7,9,11,12,13,14,16,18,24,28,30,38]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[9,24,25,31,33]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[3,6,13,15,17,21,22]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[11,26,38,39]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[4,27,34]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[8,12,15,17,20,22,26,28,36,37]}],"complexes":[],"partners":["CDX1","AJUBA","DCLK2","RRAD","ETS1","CBP","EP300"],"other_free_text":[]},"mechanistic_narrative":"EHF is an epithelium-specific ETS family transcription factor that functions as a context-dependent activator or repressor of target gene promoters to maintain epithelial differentiation, suppress epithelial-to-mesenchymal transition (EMT), and regulate innate immune and inflammatory programs across diverse epithelial tissues [PMID:10644770, PMID:11259407, PMID:22505649, PMID:24142692, PMID:34180969]. As a transcriptional repressor, EHF directly binds ETS/AP-1 composite elements to antagonize MAPK-driven transcription and silences EMT drivers (TWIST1, ZEB2), stemness factors (Lin28A/B, Sox9, Oct4), cytokines (IL-6, CXCL1), and receptors (FcεRI, CXCR4), while as an activator it induces E-cadherin, caspase-3, p16(INK4a), SPDEF, and RUVBL1 [PMID:11259407, PMID:18037958, PMID:17627613, PMID:27923832, PMID:27197175, PMID:33674341, PMID:38492894, PMID:21617703]. EHF activity is regulated post-translationally by Dclk2-mediated phosphorylation that drives nuclear export, by CpG methylation-mediated silencing, and by upstream NF-κB, TGF-β/Smad, STAT3, and AR signaling; it physically cooperates with cofactors CDX1 (via its PNT domain) and AJUBA to direct chromatin remodeling and transcriptional reprogramming [PMID:38513137, PMID:18037958, PMID:18475289, PMID:33414441, PMID:35606410, PMID:38799645]. In mouse genetic models, loss of Ehf causes papillomas, impaired goblet cell and lobuloalveolar differentiation, enhanced colitis susceptibility, and increased tumor incidence, establishing it as a broadly required epithelial homeostasis factor and context-dependent tumor suppressor [PMID:34180969, PMID:38781975, PMID:25606410]."},"prefetch_data":{"uniprot":{"accession":"Q9NZC4","full_name":"ETS homologous factor","aliases":["ETS domain-containing transcription factor","Epithelium-specific Ets transcription factor 3","ESE-3"],"length_aa":300,"mass_kda":34.9,"function":"Transcriptional activator that may play a role in regulating epithelial cell differentiation and proliferation. May act as a repressor for a specific subset of ETS/AP-1-responsive genes and as a modulator of the nuclear response to mitogen-activated protein kinase signaling cascades. Binds to DNA sequences containing the consensus nucleotide core sequence GGAA. Involved in regulation of TNFRSF10B/DR5 expression through Ets-binding sequences on the TNFRSF10B/DR5 promoter. May contribute to development and carcinogenesis by acting as a tumor suppressor gene or anti-oncogene","subcellular_location":"Nucleus","url":"https://www.uniprot.org/uniprotkb/Q9NZC4/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/EHF","classification":"Not Classified","n_dependent_lines":7,"n_total_lines":1208,"dependency_fraction":0.005794701986754967},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/EHF","total_profiled":1310},"omim":[{"mim_id":"610382","title":"PROSOPAGNOSIA, HEREDITARY","url":"https://www.omim.org/entry/610382"},{"mim_id":"605439","title":"ETS HOMOLOGOUS FACTOR; EHF","url":"https://www.omim.org/entry/605439"},{"mim_id":"602164","title":"5-@HYDROXYTRYPTAMINE RECEPTOR 4; HTR4","url":"https://www.omim.org/entry/602164"},{"mim_id":"147440","title":"INSULIN-LIKE GROWTH FACTOR I; IGF1","url":"https://www.omim.org/entry/147440"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nucleoplasm","reliability":"Supported"},{"location":"Golgi apparatus","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"esophagus","ntpm":196.3},{"tissue":"salivary gland","ntpm":270.9}],"url":"https://www.proteinatlas.org/search/EHF"},"hgnc":{"alias_symbol":["ESE3","ESEJ"],"prev_symbol":[]},"alphafold":{"accession":"Q9NZC4","domains":[{"cath_id":"1.10.150.50","chopping":"43-115","consensus_level":"high","plddt":83.4281,"start":43,"end":115},{"cath_id":"1.10.10.10","chopping":"210-289","consensus_level":"high","plddt":95.041,"start":210,"end":289}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9NZC4","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9NZC4-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9NZC4-F1-predicted_aligned_error_v6.png","plddt_mean":67.94},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=EHF","jax_strain_url":"https://www.jax.org/strain/search?query=EHF"},"sequence":{"accession":"Q9NZC4","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9NZC4.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9NZC4/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9NZC4"}},"corpus_meta":[{"pmid":"22505649","id":"PMC_22505649","title":"ESE3/EHF 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microbiology, immunology, and infection = Wei mian yu gan ran za zhi","url":"https://pubmed.ncbi.nlm.nih.gov/25648666","citation_count":4,"is_preprint":false},{"pmid":"17881193","id":"PMC_17881193","title":"Responses of hypothalamic orexin-containing neurons to cyclophosphamide, EHF-irradiation of the skin, and their combination in rats.","date":"2007","source":"Pathophysiology : the official journal of the International Society for Pathophysiology","url":"https://pubmed.ncbi.nlm.nih.gov/17881193","citation_count":4,"is_preprint":false},{"pmid":"28714026","id":"PMC_28714026","title":"miR‑206 inhibits cancer initiating cells by targeting EHF in gastric cancer.","date":"2017","source":"Oncology reports","url":"https://pubmed.ncbi.nlm.nih.gov/28714026","citation_count":4,"is_preprint":false},{"pmid":"39710057","id":"PMC_39710057","title":"The nuclear condensates of ESE3/EHF induce cellular senescence without the associated inflammatory secretory phenotype in pancreatic ductal adenocarcinoma.","date":"2024","source":"Cancer letters","url":"https://pubmed.ncbi.nlm.nih.gov/39710057","citation_count":3,"is_preprint":false},{"pmid":"34452885","id":"PMC_34452885","title":"Ese-3 Inhibits the Proliferation, Migration, and Invasion of HaCaT Cells by Downregulating PSIP1 and NUCKS1.","date":"2021","source":"Annals of clinical and laboratory science","url":"https://pubmed.ncbi.nlm.nih.gov/34452885","citation_count":3,"is_preprint":false},{"pmid":"40670887","id":"PMC_40670887","title":"LncRNA LINC01026 Is Overexpressed in Psoriasis and Enhances Keratinocyte Cell Cycle Progression by Regulating the Ets Homologous Factor (EHF).","date":"2025","source":"Journal of cellular and molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/40670887","citation_count":2,"is_preprint":false},{"pmid":"18313906","id":"PMC_18313906","title":"Morphometric analysis of hypothalamic cells showing c-Fos proteins after movement restriction and EHF-irradiation.","date":"2008","source":"Pathophysiology : the official journal of the International Society for Pathophysiology","url":"https://pubmed.ncbi.nlm.nih.gov/18313906","citation_count":2,"is_preprint":false},{"pmid":"39971220","id":"PMC_39971220","title":"EHF promotes liver cancer progression by meditating IL-6 secretion through transcription regulation of KDM2B in TAMs.","date":"2025","source":"Cellular signalling","url":"https://pubmed.ncbi.nlm.nih.gov/39971220","citation_count":2,"is_preprint":false},{"pmid":"8524760","id":"PMC_8524760","title":"[Significance of the functional state of blood phagocytes in the choice of optimal regime of EHF therapy of patients with pulmonary tuberculosis].","date":"1995","source":"Problemy tuberkuleza","url":"https://pubmed.ncbi.nlm.nih.gov/8524760","citation_count":2,"is_preprint":false},{"pmid":"39082336","id":"PMC_39082336","title":"TMB Signature-Related RCAN2 Promotes Apoptosis by Upregulating EHF/DR5 Pathway in Hepatocellular Carcinoma.","date":"2024","source":"Frontiers in bioscience (Landmark edition)","url":"https://pubmed.ncbi.nlm.nih.gov/39082336","citation_count":1,"is_preprint":false},{"pmid":"41044169","id":"PMC_41044169","title":"T3SS effector EseJ in Edwardsiella piscicida inhibits PANoptosis in macrophages.","date":"2025","source":"Communications biology","url":"https://pubmed.ncbi.nlm.nih.gov/41044169","citation_count":0,"is_preprint":false},{"pmid":"39409024","id":"PMC_39409024","title":"Codon Bias of the DDR1 Gene and Transcription Factor EHF in Multiple Species.","date":"2024","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/39409024","citation_count":0,"is_preprint":false},{"pmid":"41730908","id":"PMC_41730908","title":"The transcription factor EHF promotes the maturation and immunosuppression of conventional dendritic cells.","date":"2026","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/41730908","citation_count":0,"is_preprint":false},{"pmid":"40590703","id":"PMC_40590703","title":"An in vitro model of the epithelial airway reveals a key function for EHF in lung homeostasis and disease.","date":"2025","source":"Disease models & mechanisms","url":"https://pubmed.ncbi.nlm.nih.gov/40590703","citation_count":0,"is_preprint":false},{"pmid":"40485376","id":"PMC_40485376","title":"Evaluating Type III Secretion System Genes (escE, esaE and eseJ) of Edwardsiella piscicida for Virulence in Japanese Flounder (Paralichthys olivaceus).","date":"2025","source":"Journal of fish diseases","url":"https://pubmed.ncbi.nlm.nih.gov/40485376","citation_count":0,"is_preprint":false},{"pmid":"37772393","id":"PMC_37772393","title":"[Retracted] miR‑206 inhibits cancer initiating cells by targeting EHF in gastric cancer.","date":"2023","source":"Oncology reports","url":"https://pubmed.ncbi.nlm.nih.gov/37772393","citation_count":0,"is_preprint":false},{"pmid":"41939996","id":"PMC_41939996","title":"Lung cancer cell-derived exosomal EHF drives M2 macrophage polarization via transcriptional activation of RNF41 to promote tumor progression.","date":"2026","source":"Regenerative therapy","url":"https://pubmed.ncbi.nlm.nih.gov/41939996","citation_count":0,"is_preprint":false},{"pmid":"8630825","id":"PMC_8630825","title":"[The ultrastructural characteristics of the duodenal mucosa in peptic ulcer patients undergoing treatment with EHF-range electromagnetic radiation].","date":"1995","source":"Likars'ka sprava","url":"https://pubmed.ncbi.nlm.nih.gov/8630825","citation_count":0,"is_preprint":false},{"pmid":"16025675","id":"PMC_16025675","title":"[Study of the systemic-organ blood circulation in acute period of the multiple trauma based on water-electrolytic metabolism and its correction with EHF-therapy].","date":"2005","source":"Likars'ka sprava","url":"https://pubmed.ncbi.nlm.nih.gov/16025675","citation_count":0,"is_preprint":false},{"pmid":"17598468","id":"PMC_17598468","title":"[Expression of c-Fos gene in the rat hypothalamus upon electric painful stimulation and EHF irradiation of the skin].","date":"2007","source":"Rossiiskii fiziologicheskii zhurnal imeni I.M. Sechenova","url":"https://pubmed.ncbi.nlm.nih.gov/17598468","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.11.26.690649","title":"Loss of ESE3/EHF is sufficient to promote cell plasticity, transformation and androgen-independent status in the early stage of prostate carcinogenesis","date":"2025-11-30","source":"bioRxiv","url":"https://doi.org/10.1101/2025.11.26.690649","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2024.10.02.24314796","title":"Medical Risk Factors Associated with Listening Difficulties in Children","date":"2024-10-04","source":"bioRxiv","url":"https://doi.org/10.1101/2024.10.02.24314796","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2024.08.21.607381","title":"Gender and speech material effects on the long-term average speech spectrum, including at extended high frequencies","date":"2024-08-21","source":"bioRxiv","url":"https://doi.org/10.1101/2024.08.21.607381","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2024.07.08.602575","title":"Degradation in Binaural and Spatial Hearing, and Auditory Temporal Processing Abilities, as a Function of Aging","date":"2024-07-11","source":"bioRxiv","url":"https://doi.org/10.1101/2024.07.08.602575","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":39928,"output_tokens":8914,"usd":0.126747},"stage2":{"model":"claude-opus-4-6","input_tokens":12856,"output_tokens":4145,"usd":0.251858},"total_usd":0.378605,"stage1_batch_id":"msgbatch_0118RScg3WGaGkaZRtWZbtce","stage2_batch_id":"msgbatch_01Cpfzvtbj42aqp5oYZ5Te2s","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2000,\n      \"finding\": \"EHF (ESE-3) is an epithelium-specific ETS transcription factor that transactivates the c-MET promoter via three high-affinity binding sites, and shows different promoter transactivation specificity from ESE-1 despite similar DNA-binding affinity, indicating context-dependent transcriptional activity.\",\n      \"method\": \"Transient transfection reporter assays, DNA-binding/EMSA, promoter analysis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — in vitro promoter assays with mutagenesis and binding studies, foundational characterization paper\",\n      \"pmids\": [\"10644770\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"EHF (ESE-3) acts as a context-dependent transcriptional repressor of Ras/phorbol ester-induced promoters containing ETS and AP-1 binding sites, repressing MAPK signaling-driven transcription in a sequence- and arrangement-dependent manner; it is a nuclear protein expressed exclusively in differentiated epithelial cells and absent in epithelial carcinomas.\",\n      \"method\": \"Transient transfection reporter assays, immunohistochemistry with monoclonal antibody, promoter mutagenesis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — reconstituted promoter repression in multiple cell types with defined cis-elements, replicated in foundational study\",\n      \"pmids\": [\"11259407\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"EHF protein represses the ETS-2-induced transcriptional activity of stromelysin-1 and collagenase-1 promoters, demonstrating transcriptional repressor activity on matrix metalloproteinase promoters.\",\n      \"method\": \"Transient transfection reporter assays\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 — reporter assay with defined promoters, single lab single method\",\n      \"pmids\": [\"10527851\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"ESE-3/EHF cytokine-induced expression in bronchial smooth muscle cells is mediated through MEK1/2 and p38 MAPK signaling pathways, and overexpression of ESE-3 inhibits MMP-1 promoter activity, indicating it acts as a transcriptional repressor downstream of these kinases.\",\n      \"method\": \"Pharmacological inhibitors (U0126, SB03580), reporter assays, RT-PCR, Western blot\",\n      \"journal\": \"American journal of respiratory cell and molecular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — pathway placement via pharmacological epistasis plus reporter assay\",\n      \"pmids\": [\"12444029\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"EHF (ESE-3) re-expression in prostate cancer cells inhibits clonogenic survival and induces apoptosis by directly binding the caspase-3 (CASP3) promoter and increasing procaspase-3 levels, acting as a tumor suppressor; ESE-3 silencing in tumorigenic cells is mediated by CpG methylation of an evolutionarily conserved site in its promoter.\",\n      \"method\": \"Chromatin immunoprecipitation, reporter assay, re-expression experiments, 5-aza-2'-deoxycytidine treatment, colony assay, apoptosis assay\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — ChIP-confirmed direct promoter binding, functional rescue, epigenetic mechanism, multiple orthogonal methods\",\n      \"pmids\": [\"18037958\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"ESE-3/EHF is upregulated in cellular senescence downstream of p38 MAPK signaling; ectopic expression induces growth retardation, increased SA-β-gal activity, and upregulation of p16(INK4a) via direct binding of recombinant ESE-3 to ETS-binding sequences in the p16(INK4a) promoter.\",\n      \"method\": \"Microarray, reporter assay, recombinant protein EMSA, SA-β-gal assay, Western blot\",\n      \"journal\": \"Cancer science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — in vitro binding confirmed by EMSA with recombinant protein plus functional reporter and cellular phenotype\",\n      \"pmids\": [\"17627613\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"EHF/ESE-3 expression in airway epithelial cells is upregulated by IL-1β and TNF-α via NF-κB activation; NF-κB binding sequences in the ESE-3 promoter are required for cytokine-induced expression, and ESE-1 further upregulates ESE-3 expression.\",\n      \"method\": \"Promoter characterization, NF-κB binding site mutagenesis, reporter assays, RT-PCR\",\n      \"journal\": \"Cell research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — promoter mutagenesis identifying required cis-elements plus pathway placement via NF-κB\",\n      \"pmids\": [\"18475289\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"EHF directly activates transcription of RUVBL1, an ATPase associated with chromatin-remodeling complexes; RUVBL1 in turn represses p53 and its target genes by binding the p53 promoter, interfering with RNF20/hBRE1-mediated histone H2B monoubiquitination and promoting PAF1-mediated H3K9 trimethylation, thereby blocking p53-mediated apoptosis in colon tumor cells.\",\n      \"method\": \"Chromatin immunoprecipitation, reporter assays, siRNA knockdown, histone modification analysis, apoptosis assays\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — ChIP-confirmed direct binding, histone modification mechanistic dissection, multiple orthogonal methods\",\n      \"pmids\": [\"21617703\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"ESE3/EHF represses transcription of key EMT and cancer stem cell genes including TWIST1, ZEB2, BMI1, and POU5F1 in prostate epithelial cells; loss of ESE3/EHF induces EMT, stem-like features, and tumor-initiating properties, which are reversed by re-expression of ESE3/EHF.\",\n      \"method\": \"Loss-of-function knockdown, re-expression, gene expression analysis, in vitro and in vivo tumor assays\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — loss-of-function with defined gene targets, rescue experiments, in vivo validation\",\n      \"pmids\": [\"22505649\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"EHF promotes cornea epithelial fate through complementary gene-activating and -repressing activities, and potential interactions with KLF4 and KLF5 were identified; EHF occupancy at target gene loci was determined by ChIP-seq.\",\n      \"method\": \"Loss-of-function studies, ChIP-seq, transcriptome profiling\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — ChIP-seq genome-wide occupancy plus loss-of-function phenotype in cornea epithelium\",\n      \"pmids\": [\"24142692\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Ectopic expression of EHF in intestinal epithelial cells activates HCK-dependent apical-to-basolateral transcytosis of non-opsonized and SIgA-opsonized particles in a follicle-associated epithelium model, placing EHF upstream of HCK in regulating mucosal antigen sampling.\",\n      \"method\": \"Ectopic expression, transcytosis assays, kinase inhibitor studies\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — gain-of-function with defined cellular phenotype and pathway placement, single lab\",\n      \"pmids\": [\"23439650\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"EHF is upregulated by TGF-β/Smad signaling in mast cells; forced EHF expression represses transcription of FcεRIα, FcεRIβ, and c-Kit genes by directly binding their promoters, reducing surface FcεRI and c-Kit, suppressing degranulation and cytokine production, and decreasing GATA1, GATA2, and Stat5b expression.\",\n      \"method\": \"Forced expression, promoter-binding assays (ChIP), reporter assays, flow cytometry, degranulation assays\",\n      \"journal\": \"Journal of immunology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — ChIP-confirmed direct promoter binding, multiple downstream targets verified, functional cellular consequences\",\n      \"pmids\": [\"26297757\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"ESE3/EHF binds and represses the Lin28A and Lin28B gene promoters while activating transcription and maturation of let-7 microRNAs in normal prostate cells; loss of ESE3/EHF upregulates Lin28A/B and downregulates let-7, driving prostate cancer stem cell expansion.\",\n      \"method\": \"ChIP, promoter reporter assays, microRNA profiling, siRNA knockdown, xenograft models\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — ChIP-confirmed direct promoter binding of Lin28A/B, functional validation in vitro and in vivo\",\n      \"pmids\": [\"27197175\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"ESE3/EHF directly binds a novel ETS binding site in the IL-6 gene promoter and represses IL-6 transcription; loss of ESE3/EHF activates IL-6/JAK/STAT3 signaling, promoting stem-like transformation in prostate epithelial cells.\",\n      \"method\": \"ChIP, reporter assays, siRNA knockdown, IL-6 inhibition rescue experiments\",\n      \"journal\": \"Oncotarget\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — ChIP-confirmed direct binding, epistasis rescue by IL-6 inhibition, multiple methods\",\n      \"pmids\": [\"27732936\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"ESE3/EHF inhibits pancreatic cancer metastasis by directly upregulating E-cadherin transcription; knockdown of ESE3 promotes cell motility and invasiveness in vitro and metastasis in an orthotopic mouse model.\",\n      \"method\": \"Reporter assays, ChIP, siRNA knockdown, orthotopic mouse model, immunohistochemistry\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — ChIP-confirmed direct transcriptional regulation of E-cadherin, in vivo validation\",\n      \"pmids\": [\"27923832\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"EHF transcriptionally regulates HER2 (ERBB2) and HER3 (ERBB3) in thyroid cancer cells, contributing to PI3K/Akt and MAPK/Erk pathway activation; this was confirmed by dual-luciferase reporter and ChIP assays.\",\n      \"method\": \"ChIP, dual-luciferase reporter assay, siRNA knockdown, ectopic expression, in vivo xenograft\",\n      \"journal\": \"Oncotarget\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — ChIP plus reporter assay confirming direct transcriptional regulation, pathway placement\",\n      \"pmids\": [\"27517321\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"EHF ChIP-seq in primary human bronchial epithelial cells shows enrichment of EHF targets in inflammation and wound repair genes; EHF depletion alters epithelial secretion of a neutrophil chemokine, slows wound closure, and EHF activates SPDEF expression contributing to goblet cell hyperplasia.\",\n      \"method\": \"ChIP-seq, RNA-seq after EHF depletion, wound closure assay, cytokine secretion assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — genome-wide ChIP-seq combined with RNA-seq and functional cellular phenotypes\",\n      \"pmids\": [\"28461336\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"miR-365-3p targets EHF 3'UTR to decrease EHF protein expression; EHF acts as a transcription factor for KRT16 in oral squamous cell carcinoma, and EHF-driven KRT16 expression promotes β5-integrin/c-Met signaling and downstream Src/STAT3/FAK/ERK pathway activation.\",\n      \"method\": \"Dual-luciferase reporter (miRNA targeting), ectopic expression, siRNA knockdown, confocal colocalization, in vitro/in vivo models\",\n      \"journal\": \"Journal of experimental & clinical cancer research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — validated miRNA-EHF-KRT16 axis with multiple orthogonal methods and in vivo validation\",\n      \"pmids\": [\"30782177\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"EHF directly upregulates TGF-β1 transcription in colorectal cancer cells, thereby activating canonical TGF-β signaling to promote tumor cell proliferation, migration, and invasion.\",\n      \"method\": \"Reporter assays, ChIP, siRNA knockdown, ectopic expression, in vitro and in vivo functional assays\",\n      \"journal\": \"Cancer science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — ChIP plus reporter assay confirming direct TGF-β1 transcriptional activation\",\n      \"pmids\": [\"32372436\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"EHF produces two transcript variants (EHF-LF including exon 1, EHF-SF excluding exon 1); only EHF-SF abrogates ETS1-mediated activation of the ZEB1 promoter by promoting degradation of ETS1 protein, thereby inhibiting EMT. A point mutation in the EHF ETS domain abolishes this function and creates a dominant-negative form that enhances metastasis in vivo.\",\n      \"method\": \"Reporter assays, protein degradation assays, in vivo metastasis models, mutagenesis\",\n      \"journal\": \"Oncogenesis\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — isoform-specific function with mutagenesis, mechanistic dissection of ETS1 degradation, in vivo validation\",\n      \"pmids\": [\"33712555\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"EHF suppresses pancreatic cancer stemness by directly repressing CXCR4 transcription, decreasing cancer cell sensitivity to CXCL12 from pancreatic stellate cells; EHF also has a cell-autonomous role repressing Sox9, Sox2, Oct4, and Nanog transcription. Rosiglitazone upregulates EHF to suppress cancer stemness.\",\n      \"method\": \"ChIP, luciferase reporter assays, sphere formation assays, flow cytometry, in vivo KPC mouse model\",\n      \"journal\": \"Gut\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — ChIP-confirmed direct CXCR4 repression, multiple stemness targets, validated in KPC mice\",\n      \"pmids\": [\"33674341\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Loss of EHF promotes neuroendocrine differentiation in prostate cancer via ADT/AR/EHF/EZH2 signaling: androgen deprivation reduces EHF expression by relieving AR binding to androgen-responsive elements in the EHF locus, which promotes EZH2 expression and enzymatic activity, leading to H3K27me3-mediated transcriptional repression of downstream genes.\",\n      \"method\": \"ChIP, siRNA/shRNA knockdown, EZH2 inhibitor, cell and mouse models, Western blot\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — ChIP-confirmed AR binding and H3K27me3 mechanism, pathway epistasis with EZH2 inhibitor\",\n      \"pmids\": [\"33414441\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"EBV protein LMP2A causes upregulation of EHF via phosphorylation of STAT3 in gastric cancer cells; EHF knockdown inhibits cell proliferation and regulates cancer pathway genes including FZD5; EHF motif is enriched in activated enhancers in EBV-positive gastric cancer.\",\n      \"method\": \"STAT3 knockdown rescue by EHF overexpression, genome-wide active histone ChIP-seq, siRNA knockdown, immunostaining\",\n      \"journal\": \"Cancer science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — functional epistasis (STAT3 KD rescued by EHF OE), upstream regulation mechanism, single lab\",\n      \"pmids\": [\"34014591\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Ese-3/EHF promotes colon cancer cell proliferation by downregulating EHD2 and transactivating INPP4B as direct downstream target genes.\",\n      \"method\": \"siRNA knockdown, ectopic expression, in vitro/in vivo proliferation assays\",\n      \"journal\": \"American journal of cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — identified downstream targets with functional assays, limited mechanistic detail on direct binding\",\n      \"pmids\": [\"33520362\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"EHF physically interacts with CDX1 via its PNT domain; EHF and CDX1 co-operatively drive transcription of the colonic differentiation marker VIL1; compound deletion of Ehf and Cdx1 in mouse colon disrupts normal colonic differentiation and enhances colorectal tumor progression. Re-expression of EHF and CDX1 induces extensive chromatin remodeling and transcriptional reprogramming.\",\n      \"method\": \"Co-immunoprecipitation (PNT domain interaction), reporter assays, CRISPR knockout mouse model, ATAC-seq, RNA-seq\",\n      \"journal\": \"Cell death and differentiation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — physical interaction confirmed by Co-IP with domain mapping, in vivo mouse model, genome-wide chromatin remodeling\",\n      \"pmids\": [\"35606410\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Deletion of the EHF ETS DNA-binding domain in mice causes epidermal papillomas, preputial/vulval abscesses, corneal ulcers, increased susceptibility to colitis, impaired goblet cell differentiation, and enhanced Apc-initiated adenoma development, demonstrating that the DNA-binding domain is essential for postnatal epithelial homeostasis.\",\n      \"method\": \"Novel Ehf knockout mouse strains (Ehf-/- and gut-specific), histopathology, experimental colitis model, transcriptomic profiling\",\n      \"journal\": \"Development (Cambridge, England)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — clean genetic knockout with multiple defined tissue phenotypes, in vivo tumor model\",\n      \"pmids\": [\"34180969\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"EHF deficiency in pancreatic cancer induces CXCL1 transcription, enhancing CXCR2+ neutrophil recruitment in a CXCL1-CXCR2-dependent manner; TP53 mutation-mediated loss of EHF drives this process. Nifurtimox elevates tumoral EHF and inhibits JAK1/STAT1 pathway, suppressing CXCR2+ neutrophil recruitment.\",\n      \"method\": \"ChIP assay, Ehf-knockout mice, syngeneic mouse models, multiplexed cytokine assay, flow cytometry, single-cell RNA-seq\",\n      \"journal\": \"Gastroenterology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — ChIP-confirmed CXCL1 transcriptional regulation, in vivo genetic models, rescue by CXCL1/CXCR2 blockade\",\n      \"pmids\": [\"38492894\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Dclk2 phosphorylates EHF, changing its nucleoplasmic distribution by causing p-EHF to exit the nucleus, thereby decreasing nuclear EHF levels; nuclear EHF binds the promoters of Caspase1 and Caspase3 to reduce their transcription and inhibit neuronal pyroptosis. Cpeb4 upregulates Dclk2 by increasing Dclk2 mRNA stability.\",\n      \"method\": \"Promoter binding assays, ChIP, co-immunoprecipitation, immunofluorescence for nucleoplasmic distribution, OGD cell model, CCI mouse model, knockdown experiments\",\n      \"journal\": \"Journal of cerebral blood flow and metabolism\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — kinase-substrate relationship confirmed, subcellular relocalization mechanism defined, direct promoter binding shown by ChIP\",\n      \"pmids\": [\"38513137\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"EHF transcriptionally activates GLI1 and CCL2 in cholangiocarcinoma; EHF-driven CCL2 recruits and activates tumor-associated macrophages via the CCL2/CCR2 axis, remodeling the tumor microenvironment.\",\n      \"method\": \"ChIP, reporter assays, in vitro and in vivo functional experiments, immunohistochemistry\",\n      \"journal\": \"MedComm\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — ChIP-confirmed direct transcriptional activation of GLI1 and CCL2, in vivo validation\",\n      \"pmids\": [\"38741887\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"EHF interacts with the coactivator AJUBA to cooperatively orchestrate transcriptional network activity in gastroesophageal adenocarcinoma; EHF expression is promoted by a core transcriptional regulatory circuitry (ELF3-KLF5-GATA6); KRAS signaling is a common downstream pathway of EHF and AJUBA.\",\n      \"method\": \"Co-immunoprecipitation (EHF-AJUBA interaction), ChIP, reporter assays, in vitro and in vivo functional experiments\",\n      \"journal\": \"Acta pharmaceutica sinica. B\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — Co-IP confirmed protein interaction with coactivator, upstream CRC regulation characterized\",\n      \"pmids\": [\"38799645\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"EHF forms liquid-like nuclear condensates (phase separation) that transcriptionally repress TERT and inflammatory factors (IL-6, CXCL12); reduction of TERT leads to telomere shortening and cellular senescence in PDAC without the senescence-associated secretory phenotype (SASP). Bilobetin promotes EHF phase separation.\",\n      \"method\": \"CRISPR/Cas9 library screening, phase separation assays, ChIP, telomere length assays, drug screening, in vivo experiments\",\n      \"journal\": \"Cancer letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — phase separation characterized with functional transcriptional repression and cellular phenotype, novel mechanistic finding\",\n      \"pmids\": [\"39710057\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Loss of Ehf in the mouse mammary gland impairs lobuloalveolar differentiation at late pregnancy, reduces milk gene/lipid levels, and causes accumulation of alveolar progenitor cells; Ehf deletion attenuates prolactin-induced alveolar differentiation in mammary organoids and increases tumor incidence in the MMTV-PyMT model.\",\n      \"method\": \"Conditional Ehf knockout mouse model, mammary organoids, in vivo tumor model, gene expression analysis\",\n      \"journal\": \"Developmental cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — clean genetic knockout with defined differentiation and tumor suppressor phenotypes in vivo and ex vivo\",\n      \"pmids\": [\"38781975\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Loss of EHF in human iPSC-derived lung cells enhances CFTR activity, increases transepithelial electrical resistance, causes transcriptomic changes in basal cells, and reduces HIF-1α-mediated hypoxic response, indicating EHF controls multiple lung epithelial functions including ion channel regulation and hypoxia response.\",\n      \"method\": \"CRISPR EHF knockout in iPSC-derived lung cells, electrophysiology, RNA-seq, HIF-1α pathway analysis\",\n      \"journal\": \"Disease models & mechanisms\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — clean genetic knockout with multiple functional readouts in human iPSC model\",\n      \"pmids\": [\"40590703\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Ehf plays a non-redundant role in salivary gland ductal cell differentiation; CRISPR-Cas9 disruption of the Ehf ETS domain causes decreased granular convoluted tubules and increased intercalated Sox9-positive ductal cell accumulation, with a pronounced sexual dimorphism in males.\",\n      \"method\": \"CRISPR-Cas9 Ehf ETS domain knockout mouse, histology, immunostaining\",\n      \"journal\": \"Journal of dental research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — in vivo genetic loss-of-function with defined cellular differentiation phenotype\",\n      \"pmids\": [\"36348499\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"ESE-3/EHF regulates DR-5 (death receptor 5) expression through direct binding to putative ETS sites on the DR-5 promoter; co-factors CBP and p300 are involved in ESE-3-mediated DR-5 upregulation.\",\n      \"method\": \"EMSA, luciferase reporter assay, co-factor interaction analysis\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 — in vitro EMSA and reporter assay with defined ETS sites, identified co-factors\",\n      \"pmids\": [\"17027647\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"ESE-3/EHF cooperates with pregnane X receptor (PXR) to transactivate the ABCB1 gene in intestinal epithelial cells; ESE-3 directly binds the DR4-containing distal enhancer module of ABCB1 as shown by ChIP, and this interaction is required for rifampicin-induced ABCB1 expression.\",\n      \"method\": \"ChIP, reporter assays with DR4 mutation, siRNA knockdown, RT-PCR\",\n      \"journal\": \"Drug metabolism and pharmacokinetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — ChIP-confirmed direct binding, mutagenesis of DR4 sites, siRNA rescue, cell-type-specific mechanism\",\n      \"pmids\": [\"27567379\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"RRAD binds and regulates the subcellular localization of the transcription factor EHF in papillary thyroid cancer; EHF activates transcription of NEAT1_2, HK2, and PKM2, forming a NEAT1_2/RRAD/EHF positive feedback loop that facilitates aerobic glycolysis.\",\n      \"method\": \"Co-immunoprecipitation (RRAD-EHF), ChIP, luciferase reporter assays, RNA binding protein IP, subcellular fractionation\",\n      \"journal\": \"Endocrinology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — Co-IP confirms protein interaction, ChIP confirms direct transcriptional targets, subcellular localization linked to function\",\n      \"pmids\": [\"37279586\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"EHF in M2 tumor-associated macrophages promotes liver cancer cell metastasis by binding the KDM2B promoter and activating its transcription, which leads to increased IL-6 secretion.\",\n      \"method\": \"CUT-Tag, ChIP, luciferase assay, siRNA knockdown, co-culture migration/invasion assay, in vivo\",\n      \"journal\": \"Cellular signalling\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1-2 — CUT-Tag and ChIP confirm direct KDM2B promoter binding, functional downstream IL-6 pathway\",\n      \"pmids\": [\"39971220\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"EHF regulates conventional dendritic cell maturation and immunosuppression after TLR7/8/9 stimulation by directly regulating Ccr7, Cd200, Cd274 (PD-L1), Irf4, and Rel expression, as shown by CUT&TAG; EHF-deficient DCs promote Th1/Th17-biased CD4+ T cell responses.\",\n      \"method\": \"Conditional DC-specific Ehf knockout mice, CUT&TAG, single-cell RNA-seq, T cell polarization assays, in vivo autoimmune/tumor models\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — CUT&TAG genome-wide direct target identification, conditional knockout with multiple defined immune phenotypes\",\n      \"pmids\": [\"41730908\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"EHF in lung cancer cell-derived exosomes is transferred to macrophages and promotes M2 polarization by binding the RNF41 promoter and activating its transcription; inhibition of EHF/RNF41 axis suppresses tumor growth in vivo.\",\n      \"method\": \"ChIP, dual-luciferase reporter, exosome isolation and transfer assay, TEM, nanoparticle tracking, flow cytometry, in vivo xenograft\",\n      \"journal\": \"Regenerative therapy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — ChIP and reporter confirm direct RNF41 transcriptional regulation, exosome transfer mechanism\",\n      \"pmids\": [\"41939996\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"EHF is an epithelium-specific ETS family transcription factor that functions primarily as a context-dependent transcriptional activator or repressor — directly binding ETS/AP-1 composite elements in target promoters to repress MAPK-driven transcription and maintain epithelial differentiation; it controls a broad network of targets including E-cadherin, Lin28A/B, IL-6, CXCR4, CXCL1, caspase-3, p16(INK4a), RUVBL1, FcεRI subunits, SPDEF, TGF-β1, HER2/HER3, ZEB1/2 (via ETS1 degradation), and TERT, and its activity is regulated post-translationally by Dclk2-mediated phosphorylation (causing nuclear export), by epigenetic silencing via CpG methylation, by upstream signals including TGF-β/Smad, NF-κB, STAT3, and AR/EZH2 axes, and by physical interaction with co-factors including CDX1 (via PNT domain) and AJUBA; EHF can also form nuclear liquid-like condensates that influence its transcriptional repressor activity.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"EHF is an epithelium-specific ETS family transcription factor that functions as a context-dependent activator or repressor of target gene promoters to maintain epithelial differentiation, suppress epithelial-to-mesenchymal transition (EMT), and regulate innate immune and inflammatory programs across diverse epithelial tissues [PMID:10644770, PMID:11259407, PMID:22505649, PMID:24142692, PMID:34180969]. As a transcriptional repressor, EHF directly binds ETS/AP-1 composite elements to antagonize MAPK-driven transcription and silences EMT drivers (TWIST1, ZEB2), stemness factors (Lin28A/B, Sox9, Oct4), cytokines (IL-6, CXCL1), and receptors (FcεRI, CXCR4), while as an activator it induces E-cadherin, caspase-3, p16(INK4a), SPDEF, and RUVBL1 [PMID:11259407, PMID:18037958, PMID:17627613, PMID:27923832, PMID:27197175, PMID:33674341, PMID:38492894, PMID:21617703]. EHF activity is regulated post-translationally by Dclk2-mediated phosphorylation that drives nuclear export, by CpG methylation-mediated silencing, and by upstream NF-κB, TGF-β/Smad, STAT3, and AR signaling; it physically cooperates with cofactors CDX1 (via its PNT domain) and AJUBA to direct chromatin remodeling and transcriptional reprogramming [PMID:38513137, PMID:18037958, PMID:18475289, PMID:33414441, PMID:35606410, PMID:38799645]. In mouse genetic models, loss of Ehf causes papillomas, impaired goblet cell and lobuloalveolar differentiation, enhanced colitis susceptibility, and increased tumor incidence, establishing it as a broadly required epithelial homeostasis factor and context-dependent tumor suppressor [PMID:34180969, PMID:38781975, PMID:25606410].\",\n  \"teleology\": [\n    {\n      \"year\": 2000,\n      \"claim\": \"Identification of EHF as an epithelium-restricted ETS transcription factor with context-dependent transactivation activity established the foundational molecular identity and tissue specificity of the gene.\",\n      \"evidence\": \"Reporter assays and EMSA on c-MET promoter in transfected cells\",\n      \"pmids\": [\"10644770\", \"10527851\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No endogenous target genes confirmed by ChIP at this stage\", \"Repressor vs. activator specificity determinants unknown\", \"No loss-of-function data\"]\n    },\n    {\n      \"year\": 2001,\n      \"claim\": \"Demonstrating that EHF represses Ras/MAPK-driven transcription through ETS/AP-1 composite elements and is absent in carcinomas established its dual activator-repressor nature and implicated it as a tumor suppressor.\",\n      \"evidence\": \"Promoter mutagenesis reporter assays in multiple cell lines plus immunohistochemistry on normal vs. tumor tissue\",\n      \"pmids\": [\"11259407\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No direct in vivo tumor suppressor evidence\", \"Mechanism of repression (cofactor recruitment vs. competition) undefined\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Discovery that EHF directly activates the caspase-3 promoter to induce apoptosis and that its own expression is silenced by CpG methylation in tumors provided the first mechanistic link between EHF loss and cancer cell survival.\",\n      \"evidence\": \"ChIP on CASP3 promoter, 5-aza-2'-deoxycytidine demethylation rescue, colony and apoptosis assays in prostate cancer cells\",\n      \"pmids\": [\"18037958\", \"17627613\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Genome-wide target repertoire unknown\", \"Which methyltransferases silence EHF not identified\", \"Senescence-related p16 activation mechanism not distinguished from apoptosis pathway\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Identification of NF-κB as a direct upstream activator of EHF transcription placed EHF within inflammatory signaling cascades in airway epithelium.\",\n      \"evidence\": \"NF-κB binding site mutagenesis in the EHF promoter plus cytokine stimulation in bronchial epithelial cells\",\n      \"pmids\": [\"18475289\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether NF-κB regulation is tissue-general or airway-specific not resolved\", \"MAPK vs. NF-κB pathway hierarchy for EHF regulation unclear\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Showing that EHF loss induces EMT and stem-like features by derepressing TWIST1, ZEB2, BMI1, and POU5F1 unified EHF's tumor-suppressive role with epithelial differentiation maintenance.\",\n      \"evidence\": \"Loss-of-function knockdown with rescue, gene expression analysis, in vivo tumor-initiating assays in prostate cells\",\n      \"pmids\": [\"22505649\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct ChIP binding to each EMT promoter not confirmed for all targets\", \"Isoform-specific contributions not dissected\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Genome-wide ChIP-seq in corneal epithelium revealed that EHF directly occupies hundreds of epithelial gene loci with both activating and repressive outputs, establishing the breadth of its transcriptional regulatory network.\",\n      \"evidence\": \"ChIP-seq combined with transcriptome profiling and loss-of-function in human corneal epithelial cells\",\n      \"pmids\": [\"24142692\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Cofactor requirements for activation vs. repression not defined genome-wide\", \"Overlap of targets across different epithelial tissues not compared\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Multiple studies converged to show EHF directly controls E-cadherin, Lin28A/B-let-7, IL-6/STAT3, HER2/HER3, and ABCB1 promoters, revealing it as a multi-target hub linking differentiation, stemness, inflammation, and drug metabolism.\",\n      \"evidence\": \"ChIP-confirmed promoter binding with reporter assays in prostate, pancreatic, thyroid, and intestinal cells; in vivo xenograft validation\",\n      \"pmids\": [\"27923832\", \"27197175\", \"27517321\", \"27567379\", \"27732936\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How EHF selects among different targets in different tissues not resolved\", \"Structural basis of promoter recognition beyond ETS motif unknown\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"ChIP-seq in bronchial epithelium extended the target repertoire to wound repair and inflammation genes and showed EHF activates SPDEF to drive goblet cell differentiation, linking EHF to mucosal defense.\",\n      \"evidence\": \"ChIP-seq and RNA-seq after EHF depletion in primary human bronchial epithelial cells, wound closure assay\",\n      \"pmids\": [\"28461336\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether EHF directly interacts with SPDEF protein or only activates its transcription unclear\", \"In vivo airway phenotype of EHF loss not yet tested\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Genetic deletion of Ehf in mice produced papillomas, corneal ulcers, colitis susceptibility, goblet cell loss, and enhanced adenoma development, providing definitive in vivo proof that the ETS DNA-binding domain is essential for epithelial homeostasis and tumor suppression.\",\n      \"evidence\": \"Novel Ehf knockout mouse strains with histopathology, experimental colitis, and Apc-initiated tumor model\",\n      \"pmids\": [\"34180969\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Tissue-specific vs. systemic contributions not fully dissected with conditional knockouts at this stage\", \"Whether observed phenotypes are cell-autonomous in all tissues not confirmed\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Discovery that the short EHF isoform (EHF-SF) specifically promotes ETS1 protein degradation to block ZEB1 transcription, while a point mutant acts as a dominant-negative to enhance metastasis, revealed isoform-specific and gain-of-function mechanisms.\",\n      \"evidence\": \"Reporter assays, protein degradation assays, mutagenesis, in vivo metastasis models\",\n      \"pmids\": [\"33712555\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Proteasomal vs. lysosomal degradation pathway for ETS1 not specified\", \"Relative abundance and regulation of EHF-LF vs. EHF-SF isoforms in normal tissues unknown\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Placement of EHF downstream of AR signaling showed that androgen deprivation silences EHF, derepressing EZH2 and promoting neuroendocrine differentiation via H3K27me3, linking EHF loss to treatment-resistant prostate cancer.\",\n      \"evidence\": \"ChIP for AR binding at EHF locus, EZH2 inhibitor epistasis, mouse and cell models\",\n      \"pmids\": [\"33414441\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether EHF directly binds the EZH2 promoter or acts indirectly not resolved\", \"Clinical correlation with neuroendocrine differentiation markers not fully established\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"EHF's physical interaction with CDX1 via its PNT domain and their cooperative induction of chromatin remodeling established that EHF functions through cofactor partnerships to drive colonic differentiation, not solely through autonomous DNA binding.\",\n      \"evidence\": \"Co-immunoprecipitation with domain mapping, CRISPR compound knockout mice, ATAC-seq and RNA-seq\",\n      \"pmids\": [\"35606410\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether PNT domain mediates additional cofactor interactions not explored\", \"Structural basis of EHF-CDX1 interaction unknown\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Identification of Dclk2-mediated phosphorylation of EHF causing nuclear export established the first post-translational mechanism controlling EHF subcellular distribution and transcriptional output.\",\n      \"evidence\": \"Co-IP, immunofluorescence for nucleoplasmic distribution, ChIP on caspase promoters, OGD/CCI models\",\n      \"pmids\": [\"38513137\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Phosphorylation site(s) on EHF not mapped\", \"Whether this kinase-substrate relationship operates in epithelial tissues not tested\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"EHF deficiency was shown to upregulate CXCL1, recruiting CXCR2+ neutrophils and reshaping the pancreatic tumor microenvironment — extending EHF's tumor-suppressive role from cell-autonomous to immune-regulatory.\",\n      \"evidence\": \"ChIP, Ehf-knockout mice, syngeneic models, single-cell RNA-seq, CXCL1/CXCR2 blockade rescue\",\n      \"pmids\": [\"38492894\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether EHF regulates additional chemokines systemically not surveyed\", \"Human clinical validation of neutrophil recruitment phenotype lacking\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Discovery that EHF forms liquid-like nuclear condensates that repress TERT, IL-6, and CXCL12 introduced phase separation as a mechanism for EHF-mediated transcriptional repression and senescence induction without SASP.\",\n      \"evidence\": \"CRISPR library screening, phase separation assays, ChIP, telomere length measurement, bilobetin drug screen\",\n      \"pmids\": [\"39710057\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Biophysical parameters of EHF condensates (saturation concentration, IDR contributions) not characterized\", \"Whether condensate formation occurs in normal epithelial cells or is cancer-specific unknown\", \"Independent replication needed\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"EHF was shown to regulate dendritic cell maturation by directly controlling Ccr7, PD-L1, Irf4, and Rel expression, demonstrating a non-epithelial immune cell-autonomous role in shaping adaptive immune responses.\",\n      \"evidence\": \"Conditional DC-specific Ehf knockout mice, CUT&TAG, single-cell RNA-seq, T cell polarization assays, autoimmune/tumor models\",\n      \"pmids\": [\"41730908\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether EHF functions in other immune cell lineages not tested\", \"Human DC relevance not confirmed\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"Key unresolved questions include the structural basis of EHF's activator-repressor switch, the full spectrum of phosphorylation sites governing its nuclear-cytoplasmic shuttling, whether phase separation is a general EHF mechanism in normal epithelia, and the complete set of PNT domain-mediated cofactor interactions.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No crystal or cryo-EM structure of EHF or its complexes\", \"Systematic mapping of EHF post-translational modifications not performed\", \"Tissue-specific isoform expression and regulation not comprehensively characterized\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [0, 1, 4, 5, 7, 8, 9, 11, 12, 13, 14, 15, 16, 18, 20, 24, 26, 28, 30, 35, 38]},\n      {\"term_id\": \"GO:0003677\", \"supporting_discovery_ids\": [0, 1, 2, 4, 5, 9, 11, 12, 13, 14, 16, 34, 38]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [1, 27, 30]},\n      {\"term_id\": \"GO:0005654\", \"supporting_discovery_ids\": [27, 30]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [0, 1, 4, 7, 9, 11, 12, 13, 14, 16, 18, 24, 28, 30, 38]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [9, 24, 25, 31, 33]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [3, 6, 13, 15, 17, 21, 22]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [11, 26, 38, 39]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [4, 27, 34]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [8, 12, 15, 17, 20, 22, 26, 28, 36, 37]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\n      \"CDX1\",\n      \"AJUBA\",\n      \"DCLK2\",\n      \"RRAD\",\n      \"ETS1\",\n      \"CBP\",\n      \"EP300\"\n    ],\n    \"other_free_text\": []\n  }\n}\n```"}