{"gene":"CLDN4","run_date":"2026-06-09T22:57:18","timeline":{"discoveries":[{"year":2006,"finding":"The CLDN4 promoter contains two Sp1 binding sites both required for promoter activity; cells overexpressing CLDN4 exhibit low DNA methylation and high histone H3 acetylation of the critical Sp1-containing promoter region, whereas CLDN4-negative cells show the reverse; treatment with demethylating and/or acetylating agents induces CLDN4 expression in CLDN4-negative cells.","method":"Promoter deletion analysis, site-directed mutagenesis of Sp1 sites, ChIP for histone H3 acetylation, bisulfite sequencing for DNA methylation, pharmacological treatment with demethylating/acetylating agents","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — promoter mutagenesis plus multiple orthogonal epigenetic methods (methylation, acetylation ChIP, pharmacological reversal) in one study","pmids":["16714763"],"is_preprint":false},{"year":2010,"finding":"CLDN4 repression in normal ovarian epithelial cells is associated with bivalent histone modifications (H3K4me3 + H3K27me3) and DNA hypermethylation; during ovarian tumorigenesis, derepression correlates with loss of H3K27me3 and H4K20me3; combined DNA demethylation and histone acetylation treatment robustly reverses CLDN4 repression, whereas loss of H3K27me3 alone is insufficient.","method":"ChIP for H3K4me3, H3K27me3, H4K20me3; bisulfite sequencing; pharmacological treatment (demethylating + acetylating agents); gene expression analysis","journal":"Carcinogenesis","confidence":"High","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal chromatin and methylation methods, functional pharmacological reversal, single lab","pmids":["20053926"],"is_preprint":false},{"year":2010,"finding":"C-CPE (C-terminus of Clostridium perfringens enterotoxin) binds CLDN4 as a specific receptor, decreases CLDN4 protein expression, and relocates CLDN4 from cell-cell contact regions to the cytoplasm in ovarian cancer cells; C-CPE sensitizes EOC cells to Taxol and Carboplatin in a CLDN4-dependent manner and activates the ubiquitin-proteasome pathway.","method":"Quantitative RT-PCR, immunofluorescence, Western blot, 3D culture and monolayer culture assays, xenograft mouse model, oligonucleotide microarray","journal":"Clinical cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vitro and in vivo functional assays with CLDN4-dependent rescue, single lab, multiple orthogonal readouts","pmids":["21123456"],"is_preprint":false},{"year":2007,"finding":"STAT2 DNA-binding activity transcriptionally activates CLDN4 as an interferon-stimulated gene; siRNA knockdown of CLDN4 in fibroblasts reduces IFN-alpha/beta-induced antiproliferative and antiviral responses, consistent with cells expressing a DNA-binding mutant of STAT2.","method":"siRNA knockdown, gene microarray, site-directed mutagenesis of STAT2, biological IFN response assays (antiproliferative, antiviral)","journal":"Journal of interferon & cytokine research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — siRNA knockdown with two orthogonal functional readouts, corroborated by STAT2 mutagenesis, single lab","pmids":["17651017"],"is_preprint":false},{"year":2019,"finding":"PAK4 phosphorylates CEBPB on Thr-235, which then binds the -1093 to -991 bp region of the CLDN4 promoter to transcriptionally upregulate CLDN4 expression, promoting breast cancer cell migration and invasion; restoration of CLDN4 in PAK4-knockdown cells reverses inhibition of migration and invasion.","method":"PAK4 siRNA knockdown, CEBPB promoter ChIP/binding assay, luciferase reporter for CLDN4 promoter, rescue overexpression of CLDN4, migration/invasion assays","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP/reporter plus rescue experiment, single lab, multiple orthogonal methods","pmids":["30808546"],"is_preprint":false},{"year":2020,"finding":"ZNF703 directly binds the CLDN4 promoter and transactivates CLDN4 expression to promote EMT and HCC metastasis; downregulation of CLDN4 attenuates ZNF703-mediated metastasis, and CLDN4 upregulation reverses the reduced metastasis seen upon ZNF703 knockdown.","method":"ChIP for ZNF703 binding to CLDN4 promoter, loss- and gain-of-function experiments, in vitro and in vivo metastasis assays, epistasis rescue","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP and rescue epistasis, in vitro and in vivo, single lab","pmids":["32269215"],"is_preprint":false},{"year":2022,"finding":"TGF-β signaling upregulates CLDN4 expression in GBM cells and promotes nuclear translocation of CLDN4; CLDN4 modulates the TNF-α/NF-κB signaling pathway; inhibition of CLDN4 suppresses mesenchymal transition, invasion, and migration, and TGF-β inhibitor ITD-1 downregulates CLDN4 and inhibits invasion.","method":"CLDN4 knockdown/overexpression, subcellular fractionation/imaging for nuclear translocation, TGF-β pathway inhibitor treatment, in vitro invasion/migration assays, in vivo tumor growth assays","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — nuclear translocation validated with fractionation, epistasis via inhibitor treatment plus functional assays, single lab","pmids":["35418179"],"is_preprint":false},{"year":2023,"finding":"RBM15, an m6A RNA methyltransferase, suppresses CLDN4 expression through m6A-mediated epigenetic inhibition in hepatic cells; RBM15 overexpression increases insulin resistance, and this effect is mediated through m6A regulation of CLDN4.","method":"MeRIP sequencing, mRNA-seq, RBM15 overexpression/knockdown, glucose uptake tests, Western blot, immunohistochemistry","journal":"Molecular medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — MeRIP-seq identifies m6A mark on CLDN4 mRNA, functional glucose uptake assay, single lab","pmids":["36803098"],"is_preprint":false},{"year":2023,"finding":"IGF2BP3 interacts with CLDN4 mRNA and augments its stability in an m6A-dependent manner; CLDN4 upregulation by IGF2BP3 activates NF-κB signaling in gallbladder cancer cells; restoration of CLDN4 reverses the inhibitory effect of IGF2BP3 knockdown on gallbladder cancer progression.","method":"RNA immunoprecipitation (RIP), m6A methylation assays, IGF2BP3 knockdown/overexpression, NF-κB pathway analysis, rescue CLDN4 overexpression, in vitro/in vivo tumor assays","journal":"Translational oncology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — RIP plus m6A assay and rescue epistasis, in vitro and in vivo, single lab","pmids":["37643553"],"is_preprint":false},{"year":2023,"finding":"FOXA1 transcriptionally activates CLDN4 by binding its promoter; CLDN4 overexpression induces activation of the PI3K/AKT pathway; celastrol inhibits GC progression by downregulating FOXA1, thereby reducing CLDN4 and impeding PI3K/AKT.","method":"Luciferase reporter assay for FOXA1 binding to CLDN4 promoter, FOXA1/CLDN4 overexpression/knockdown, Western blot for PI3K/AKT phosphorylation, MTT and Transwell assays","journal":"Toxicology research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — luciferase reporter plus functional pathway assays, single lab","pmids":["37397926"],"is_preprint":false},{"year":2022,"finding":"CLDN4 knockdown in AML cells inhibits mRNA expression of PIK3R3 and MAP2K2, suppressing AKT and ERK1/2 activation; partial rescue of cell viability by AKT activator SC79 confirms AKT as a downstream effector; CLDN4 promotes AML cell growth and suppresses apoptosis.","method":"CLDN4 siRNA knockdown, Western blot for pAKT and pERK1/2, SC79 (AKT activator) rescue experiment, cell viability and apoptosis assays","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pathway activation confirmed by pharmacological rescue, single lab","pmids":["35760010"],"is_preprint":false},{"year":2022,"finding":"In bladder urothelial carcinoma cells, demethylation-induced excess CLDN4 that is not integrated into tight junctions (TJ-unintegrated CLDN4 monomer) binds integrin β1, increases FAK phosphorylation, and promotes stemness, drug resistance, and metastatic ability; CLDN4 knockdown reduces FAK phosphorylation.","method":"Co-immunoprecipitation/pulldown of CLDN4 with integrin β1, Western blot for FAK phosphorylation, CLDN4 knockdown, demethylating agent (AZA) treatment, fractionation to identify TJ-unintegrated CLDN4","journal":"International journal of molecular sciences","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — Co-IP binding assay, FAK phosphorylation readout with knockdown rescue, single lab","pmids":["35742959"],"is_preprint":false},{"year":2025,"finding":"CLDN4 is palmitoylated at cysteine residues C104 and C107; this palmitoylation regulates ubiquitination at lysine K103, inhibits clathrin-mediated endocytosis, and sustains CLDN4 anchoring within lipid rafts; lipid raft-anchored CLDN4 drives mobilization of contactin-1 to lipid rafts and activates Notch signaling, promoting hepatic-to-biliary lineage transition and lenvatinib resistance in HCC.","method":"Palmitoylation site mutagenesis (C104/C107), ubiquitination assays, lipid raft fractionation, clathrin-mediated endocytosis assays, Co-IP for contactin-1, Notch pathway reporter assays, drug resistance assays","journal":"Cell reports. Medicine","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — site-directed mutagenesis of palmitoylation sites combined with ubiquitination assays, lipid raft fractionation, and functional Notch pathway readouts in one study","pmids":["40592346"],"is_preprint":false},{"year":2025,"finding":"CLDN4 knockout in SCLC cells promotes cell proliferation by accelerating cell cycle progression; CLDN4 knockout upregulates SAA1 which partly mediates the proliferation-promoting effect; CLDN4 expression is directly regulated by SP1, with DNA methylation also contributing to transcriptional regulation.","method":"CRISPR/CRISPR-based CLDN4 knockout, RNA-seq, SP1 binding assays, DNA methylation analysis, SAA1 knockdown rescue, cell cycle and proliferation assays","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — knockout with RNA-seq pathway identification and SP1 binding, single lab","pmids":["41016339"],"is_preprint":false},{"year":2025,"finding":"CLDN4 overexpression in corpus cavernosum smooth muscle cells activates the JNK signaling pathway, increases fibrotic protein expression, and impairs erectile function in vivo; hypoxia increases CLDN4 expression, and Cldn4 overexpression in rat corpus cavernosum increases local fibrosis and impairs erectile function.","method":"Lentiviral Cldn4 overexpression in vitro and in vivo (rat corpus cavernosum), Western blot for JNK pathway and fibrotic proteins, functional erectile assays, immunofluorescence","journal":"The journal of sexual medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — gain-of-function in vitro and in vivo with pathway readout, single lab","pmids":["38477100"],"is_preprint":false},{"year":2025,"finding":"CLDN4 knockdown in cerulein-stimulated pancreatic acinar cells reduces ROS, iron accumulation, inflammatory cytokines (TNF-α, IL-6, IL-17), restores GPX4 levels, and reduces ACSL4 expression, suppressing ferroptosis; CLDN4 knockdown decreases JAK2/STAT3 pathway activation, and combined CLDN4 knockdown with JAK2 inhibitor AG490 provides additive protective effects in AP models.","method":"shRNA CLDN4 knockdown, RNA-seq, Western blot for JAK2/STAT3, GPX4, ACSL4, ELISA for cytokines, flow cytometry for ROS, cerulein AP mouse model, pharmacological JAK2 inhibition (AG490) epistasis","journal":"Functional & integrative genomics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function with pharmacological epistasis (AG490), in vitro and in vivo, single lab","pmids":["40892111"],"is_preprint":false},{"year":2020,"finding":"SPTBN2 interacts with CLDN4 to promote endometrial cancer cell migration and invasion via the PI3K/AKT pathway; CLDN4 overexpression partially reverses the decrease in migration/invasion caused by SPTBN2 knockdown, and CLDN4 is itself upregulated in EEC and promotes metastasis.","method":"Co-IP/interaction assay between SPTBN2 and CLDN4, SPTBN2/CLDN4 knockdown and overexpression, PI3K/AKT pathway western blot, migration/invasion assays, rescue epistasis","journal":"Cell death discovery","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single Co-IP with functional rescue, single lab, single paper","pmids":["34887379"],"is_preprint":false},{"year":2025,"finding":"AP-1 transcription factors (JUNB, FOSB, FOS) are required for CLDN4 induction in transitional AT2 cells following viral lung injury; CLDN4+ AT2 cells represent a KRT8-high transitional substate with chromatin enriched for AP-1 motifs; AP-1 promotes AT2 cell dispersion and senescence signaling toward fibroblasts.","method":"Mouse genetics (AP-1 member conditional knockout), multiomics (joint transcriptomic-epigenomic profiling), immunostaining, chromatin accessibility (ATAC-seq) for AP-1 motif enrichment, in vivo viral injury model","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — mouse genetic knockouts with multiomics, single lab preprint","pmids":[],"is_preprint":true}],"current_model":"CLDN4 is a tight junction protein whose expression is transcriptionally regulated by Sp1, FOXA1, CEBPB (phosphorylated by PAK4), ZNF703, ELF3, and AP-1 factors, and epigenetically controlled by DNA methylation and repressive histone marks (H3K27me3, H4K20me3); STAT2 DNA-binding activity induces CLDN4 as an interferon-stimulated gene mediating antiviral and antiproliferative IFN responses; post-translationally, CLDN4 palmitoylation at C104/C107 regulates its ubiquitination at K103, prevents clathrin-mediated endocytosis, anchors it in lipid rafts where it mobilizes contactin-1 and activates Notch signaling; when not integrated into tight junctions, CLDN4 binds integrin β1 to activate FAK and downstream PI3K/AKT; additionally CLDN4 can translocate to the nucleus in response to TGF-β to modulate TNF-α/NF-κB signaling, and promotes ferroptosis-related inflammation via the JAK2/STAT3 pathway, while its mRNA stability is enhanced by the m6A readers IGF2BP3 and RBM15."},"narrative":{"mechanistic_narrative":"CLDN4 is a tight junction protein whose expression is tightly controlled by transcriptional and epigenetic mechanisms and whose non-junctional pools act as signaling effectors in epithelial transformation and tissue injury [PMID:16714763, PMID:35742959]. Its promoter is driven by two essential Sp1 sites and is silenced in normal cells by DNA hypermethylation, repressive histone marks (H3K27me3, H4K20me3) and bivalent chromatin, with combined demethylation and histone acetylation robustly derepressing the gene [PMID:16714763, PMID:20053926, PMID:41016339]. A range of sequence-specific factors converge on the CLDN4 promoter to activate it in disease contexts: PAK4-phosphorylated CEBPB, FOXA1, ZNF703, AP-1 factors, and STAT2, the latter establishing CLDN4 as an interferon-stimulated gene that mediates antiviral and antiproliferative IFN responses [PMID:17651017, PMID:30808546, PMID:32269215, PMID:37397926]. Post-transcriptionally, CLDN4 mRNA fate is set by m6A: IGF2BP3 stabilizes the transcript while RBM15-dependent m6A methylation suppresses it [PMID:36803098, PMID:37643553]. A key mechanistic theme is that CLDN4 not integrated into tight junctions acts as a signaling hub: such monomeric CLDN4 binds integrin β1 to drive FAK phosphorylation and PI3K/AKT activation, promoting stemness, drug resistance and metastasis [PMID:35742959, PMID:37397926, PMID:34887379]. Palmitoylation at C104/C107 controls CLDN4 fate by regulating ubiquitination at K103, blocking clathrin-mediated endocytosis and anchoring CLDN4 in lipid rafts, where it mobilizes contactin-1 and activates Notch signaling to drive lineage transition and lenvatinib resistance [PMID:40592346]. CLDN4 also feeds NF-κB signaling, undergoes TGF-β-induced nuclear translocation, drives JAK2/STAT3-dependent ferroptotic inflammation, and engages MAPK pathways (ERK1/2, JNK) across cancer, pancreatitis and fibrosis models [PMID:35418179, PMID:37643553, PMID:35760010, PMID:40892111, PMID:38477100].","teleology":[{"year":2006,"claim":"Established the core transcriptional control of CLDN4, showing its promoter depends on Sp1 sites whose activity is gated by DNA methylation and histone acetylation status.","evidence":"Promoter deletion/mutagenesis of Sp1 sites, ChIP for H3 acetylation, bisulfite sequencing and pharmacological reversal in cell lines","pmids":["16714763"],"confidence":"High","gaps":["Did not address upstream signals that recruit Sp1 or remodel the locus","Tested in a limited set of cell lines"]},{"year":2007,"claim":"Defined CLDN4 as an interferon-stimulated gene, linking it to STAT2-driven antiviral and antiproliferative IFN responses beyond a purely structural role.","evidence":"siRNA knockdown with antiproliferative/antiviral readouts plus STAT2 DNA-binding mutagenesis in fibroblasts","pmids":["17651017"],"confidence":"Medium","gaps":["Direct STAT2 occupancy at the CLDN4 promoter not shown","Molecular role of CLDN4 within the IFN response uncharacterized"]},{"year":2010,"claim":"Resolved the chromatin logic of CLDN4 silencing and derepression, showing bivalent and repressive histone marks plus DNA methylation cooperate, with loss of H3K27me3 alone being insufficient.","evidence":"ChIP for H3K4me3/H3K27me3/H4K20me3, bisulfite sequencing and pharmacological reversal in ovarian epithelial/tumor cells; C-CPE receptor binding and proteasomal degradation in EOC cells","pmids":["20053926","21123456"],"confidence":"High","gaps":["Writers/erasers responsible for the mark switch not identified","C-CPE degradation mechanism not connected to physiological turnover"]},{"year":2020,"claim":"Identified specific transcription factors (PAK4-CEBPB, ZNF703) and a binding partner (SPTBN2) that activate CLDN4 to drive migration, invasion and metastasis, framing CLDN4 as a downstream pro-metastatic effector.","evidence":"Promoter ChIP/reporter, Co-IP, and CLDN4 rescue epistasis with migration/invasion and in vivo metastasis assays in breast, HCC and endometrial cancer models","pmids":["30808546","32269215","34887379"],"confidence":"Medium","gaps":["SPTBN2-CLDN4 interaction rests on a single Co-IP without reciprocal validation","How CLDN4 mechanistically promotes motility downstream of these factors not fully resolved"]},{"year":2022,"claim":"Showed that non-junctional CLDN4 acts as a signaling node, binding integrin β1 to activate FAK and feeding PI3K/AKT and ERK pathways that promote stemness, drug resistance and survival.","evidence":"Co-IP of CLDN4 with integrin β1, FAK phosphorylation with knockdown, demethylation-induced TJ-unintegrated CLDN4 fractionation in bladder carcinoma; AKT/ERK pathway readouts with SC79 rescue in AML","pmids":["35742959","35760010"],"confidence":"Medium","gaps":["Structural basis of CLDN4-integrin β1 binding unknown","Whether monomeric versus junctional CLDN4 is the relevant species in all contexts not established"]},{"year":2022,"claim":"Demonstrated TGF-β-induced nuclear translocation of CLDN4 and its modulation of TNF-α/NF-κB signaling, indicating a non-membrane function in mesenchymal transition.","evidence":"Subcellular fractionation/imaging, TGF-β inhibitor treatment and invasion/migration assays in GBM cells","pmids":["35418179"],"confidence":"Medium","gaps":["Nuclear binding partners and direct transcriptional targets of CLDN4 not identified","Mechanism of nuclear import unknown"]},{"year":2023,"claim":"Established m6A as a post-transcriptional control layer for CLDN4, with opposing reader/writer effects: IGF2BP3 stabilizing the mRNA and RBM15 suppressing it.","evidence":"RIP/MeRIP and m6A assays with knockdown/overexpression, NF-κB analysis and CLDN4 rescue in gallbladder cancer; MeRIP-seq with glucose uptake assays in hepatic insulin resistance","pmids":["37643553","36803098"],"confidence":"Medium","gaps":["Specific m6A sites on CLDN4 mRNA not mapped to function","Reconciliation of stabilizing versus suppressive m6A outcomes across tissues not addressed"]},{"year":2023,"claim":"Linked FOXA1-driven CLDN4 expression to PI3K/AKT activation, reinforcing CLDN4 as a transcriptionally inducible driver of oncogenic signaling.","evidence":"Luciferase reporter for FOXA1 binding, overexpression/knockdown and PI3K/AKT phosphorylation with functional assays in gastric cancer","pmids":["37397926"],"confidence":"Medium","gaps":["Direct molecular link between CLDN4 and PI3K/AKT components not defined","Promoter occupancy by FOXA1 inferred from reporter rather than ChIP"]},{"year":2025,"claim":"Defined a palmitoylation-ubiquitination switch that controls CLDN4 trafficking, anchoring it in lipid rafts to mobilize contactin-1 and activate Notch signaling, mechanistically explaining its role in lineage transition and drug resistance.","evidence":"C104/C107 palmitoylation-site mutagenesis, ubiquitination and lipid raft assays, endocytosis assays, Co-IP for contactin-1 and Notch reporters in HCC","pmids":["40592346"],"confidence":"High","gaps":["Palmitoyltransferase and deubiquitinase enzymes not identified","Generality of the raft/Notch axis beyond HCC unknown"]},{"year":2025,"claim":"Extended CLDN4 function to inflammatory ferroptosis and tissue fibrosis, implicating JAK2/STAT3 and JNK pathways in non-cancer pathology, and to cell cycle suppression via SAA1.","evidence":"shRNA knockdown with AG490 epistasis in cerulein pancreatitis; Cldn4 overexpression with JNK readout in rat corpus cavernosum; CRISPR knockout with RNA-seq/SAA1 rescue in SCLC","pmids":["40892111","38477100","41016339"],"confidence":"Medium","gaps":["Whether CLDN4 directly activates JAK2/STAT3 or JNK or acts indirectly is unresolved","Context-dependent pro- versus anti-proliferative roles not reconciled"]},{"year":null,"claim":"How junctional versus non-junctional CLDN4 pools are partitioned and how a single tight junction protein integrates such diverse downstream pathways (integrin/FAK, Notch, NF-κB, JAK2/STAT3, JNK) into context-specific outcomes remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unifying structural model distinguishing signaling-competent CLDN4 conformers","Direct biochemical link between CLDN4 and most downstream kinases unestablished","Physiological (non-disease) function in normal epithelia underexplored in the corpus"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[0,11]},{"term_id":"GO:0098631","term_label":"cell adhesion mediator activity","supporting_discovery_ids":[11]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[11,12]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[2]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[6]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[11,12]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[0,1]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[3]},{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[7,8]}],"complexes":["tight junction","lipid raft"],"partners":["ITGB1","CNTN1","SPTBN2","IGF2BP3"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"O14493","full_name":"Claudin-4","aliases":["Clostridium perfringens enterotoxin receptor","CPE-R","CPE-receptor","Williams-Beuren syndrome chromosomal region 8 protein"],"length_aa":209,"mass_kda":22.1,"function":"Can associate with other claudins to regulate tight junction structural and functional strand dynamics (PubMed:35773259, PubMed:36008380). May coassemble with CLDN8 into tight junction strands containing anion-selective channels that convey paracellular chloride permeability in renal collecting ducts (By similarity) (PubMed:36008380). May integrate into CLDN3 strands to modulate localized tight junction barrier properties (PubMed:35773259, PubMed:36008380). May disrupt strand assembly of channel-forming CLDN2 and CLDN15 and inhibit cation conductance (PubMed:35773259, PubMed:36008380). Cannot form tight junction strands on its own (PubMed:35773259, PubMed:36008380)","subcellular_location":"Cell junction, tight junction; Cell membrane","url":"https://www.uniprot.org/uniprotkb/O14493/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/CLDN4","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/CLDN4","total_profiled":1310},"omim":[{"mim_id":"611232","title":"CLAUDIN 12; CLDN12","url":"https://www.omim.org/entry/611232"},{"mim_id":"611231","title":"CLAUDIN 8; CLDN8","url":"https://www.omim.org/entry/611231"},{"mim_id":"609131","title":"CLAUDIN 7; CLDN7","url":"https://www.omim.org/entry/609131"},{"mim_id":"608576","title":"GRAINYHEAD-LIKE 2; GRHL2","url":"https://www.omim.org/entry/608576"},{"mim_id":"603718","title":"CLAUDIN 1; CLDN1","url":"https://www.omim.org/entry/603718"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Plasma membrane","reliability":"Supported"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"esophagus","ntpm":317.9},{"tissue":"intestine","ntpm":265.3},{"tissue":"urinary bladder","ntpm":282.7}],"url":"https://www.proteinatlas.org/search/CLDN4"},"hgnc":{"alias_symbol":["CPE-R","WBSCR8","hCPE-R"],"prev_symbol":["CPETR","CPETR1"]},"alphafold":{"accession":"O14493","domains":[{"cath_id":"1.20.140.150","chopping":"3-29_73-182","consensus_level":"high","plddt":90.9558,"start":3,"end":182}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/O14493","model_url":"https://alphafold.ebi.ac.uk/files/AF-O14493-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-O14493-F1-predicted_aligned_error_v6.png","plddt_mean":84.56},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=CLDN4","jax_strain_url":"https://www.jax.org/strain/search?query=CLDN4"},"sequence":{"accession":"O14493","fasta_url":"https://rest.uniprot.org/uniprotkb/O14493.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/O14493/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/O14493"}},"corpus_meta":[{"pmid":"28819095","id":"PMC_28819095","title":"Non-coding RNAs participate in the regulatory network of CLDN4 via ceRNA mediated miRNA evasion.","date":"2017","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/28819095","citation_count":229,"is_preprint":false},{"pmid":"16714763","id":"PMC_16714763","title":"Crucial roles of Sp1 and epigenetic modifications in the regulation of the CLDN4 promoter in ovarian cancer cells.","date":"2006","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/16714763","citation_count":121,"is_preprint":false},{"pmid":"20053926","id":"PMC_20053926","title":"Derepression of CLDN3 and CLDN4 during ovarian tumorigenesis is associated with loss of repressive histone modifications.","date":"2010","source":"Carcinogenesis","url":"https://pubmed.ncbi.nlm.nih.gov/20053926","citation_count":65,"is_preprint":false},{"pmid":"21123456","id":"PMC_21123456","title":"C terminus of Clostridium perfringens enterotoxin downregulates CLDN4 and sensitizes ovarian cancer cells to Taxol and Carboplatin.","date":"2010","source":"Clinical cancer research : an official journal of the American Association for Cancer Research","url":"https://pubmed.ncbi.nlm.nih.gov/21123456","citation_count":50,"is_preprint":false},{"pmid":"35418179","id":"PMC_35418179","title":"TGF-β induces GBM mesenchymal transition through upregulation of CLDN4 and nuclear translocation to activate TNF-α/NF-κB signal pathway.","date":"2022","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/35418179","citation_count":49,"is_preprint":false},{"pmid":"30808546","id":"PMC_30808546","title":"A novel PAK4-CEBPB-CLDN4 axis involving in breast cancer cell migration and invasion.","date":"2019","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/30808546","citation_count":39,"is_preprint":false},{"pmid":"32779991","id":"PMC_32779991","title":"ELFN1-AS1 accelerates cell proliferation, invasion and migration via regulating miR-497-3p/CLDN4 axis in ovarian cancer.","date":"2020","source":"Bioengineered","url":"https://pubmed.ncbi.nlm.nih.gov/32779991","citation_count":38,"is_preprint":false},{"pmid":"9878248","id":"PMC_9878248","title":"Genes for the CPE receptor (CPETR1) and the human homolog of RVP1 (CPETR2) are localized within the Williams-Beuren syndrome deletion.","date":"1998","source":"Genomics","url":"https://pubmed.ncbi.nlm.nih.gov/9878248","citation_count":37,"is_preprint":false},{"pmid":"32269215","id":"PMC_32269215","title":"Zinc finger protein 703 induces EMT and sorafenib resistance in hepatocellular carcinoma by transactivating CLDN4 expression.","date":"2020","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/32269215","citation_count":35,"is_preprint":false},{"pmid":"36803098","id":"PMC_36803098","title":"RBM15 suppresses hepatic insulin sensitivity of offspring of gestational diabetes mellitus mice via m6A-mediated regulation of CLDN4.","date":"2023","source":"Molecular medicine (Cambridge, Mass.)","url":"https://pubmed.ncbi.nlm.nih.gov/36803098","citation_count":35,"is_preprint":false},{"pmid":"16670314","id":"PMC_16670314","title":"Advanced intercross line mapping of Eae5 reveals Ncf-1 and CLDN4 as candidate genes for experimental autoimmune encephalomyelitis.","date":"2006","source":"Journal of immunology (Baltimore, Md. : 1950)","url":"https://pubmed.ncbi.nlm.nih.gov/16670314","citation_count":35,"is_preprint":false},{"pmid":"37397926","id":"PMC_37397926","title":"Celastrol inhibits gastric cancer cell proliferation, migration, and invasion via the FOXA1/CLDN4 axis.","date":"2023","source":"Toxicology research","url":"https://pubmed.ncbi.nlm.nih.gov/37397926","citation_count":33,"is_preprint":false},{"pmid":"31856376","id":"PMC_31856376","title":"CLDN4 silencing promotes proliferation and reduces chemotherapy sensitivity of gastric cancer cells through activation of the PI3K/Akt signalling 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Medicine","url":"https://pubmed.ncbi.nlm.nih.gov/40592346","citation_count":8,"is_preprint":false},{"pmid":"17651017","id":"PMC_17651017","title":"Interferon-inducible Stat2 activation of JUND and CLDN4: mediators of IFN responses.","date":"2007","source":"Journal of interferon & cytokine research : the official journal of the International Society for Interferon and Cytokine Research","url":"https://pubmed.ncbi.nlm.nih.gov/17651017","citation_count":7,"is_preprint":false},{"pmid":"35760010","id":"PMC_35760010","title":"CLDN4 promotes growth of acute myeloid leukemia cells via regulating AKT and ERK1/2 signaling.","date":"2022","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/35760010","citation_count":4,"is_preprint":false},{"pmid":"40892111","id":"PMC_40892111","title":"CLDN4 promotes ferroptosis and inflammation involving JAK2/STAT3 pathway in acute pancreatitis.","date":"2025","source":"Functional & integrative genomics","url":"https://pubmed.ncbi.nlm.nih.gov/40892111","citation_count":2,"is_preprint":false},{"pmid":"38477100","id":"PMC_38477100","title":"Cldn4 overexpression promotes penile cavernous smooth muscle cell fibrotic response via the JNK signaling pathway.","date":"2024","source":"The journal of sexual medicine","url":"https://pubmed.ncbi.nlm.nih.gov/38477100","citation_count":2,"is_preprint":false},{"pmid":"41252335","id":"PMC_41252335","title":"NK Cell-Derived Small Extracellular Vesicles Armed With CLDN4-Targeting Peptides Potentiate Radiotherapy in Gastric Cancer.","date":"2025","source":"Journal of extracellular vesicles","url":"https://pubmed.ncbi.nlm.nih.gov/41252335","citation_count":1,"is_preprint":false},{"pmid":"41016339","id":"PMC_41016339","title":"CLDN4 regulates cell proliferation in small cell lung cancer cells via SAA1 inhibition.","date":"2025","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/41016339","citation_count":1,"is_preprint":false},{"pmid":"39442989","id":"PMC_39442989","title":"[Risk model construction and immune cell infiltration analysis of the CLDN4 gene in ovarian cancer cells].","date":"2024","source":"Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology","url":"https://pubmed.ncbi.nlm.nih.gov/39442989","citation_count":0,"is_preprint":false},{"pmid":"40074190","id":"PMC_40074190","title":"MicroRNA-regulated flounder CLDN4 functions in anti-bacterial immunity.","date":"2025","source":"Fish & shellfish immunology","url":"https://pubmed.ncbi.nlm.nih.gov/40074190","citation_count":0,"is_preprint":false},{"pmid":"41456351","id":"PMC_41456351","title":"Fasudil alleviates lipopolysaccharides-triggered damage to BEAS-2B cells and human lung organoids by inducing CLDN4 expression.","date":"2025","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/41456351","citation_count":0,"is_preprint":false},{"pmid":"42159105","id":"PMC_42159105","title":"Shenfu Injection Alleviates Pancreatitis-Induced Damage and Oxidative Stress by Upregulating CLDN4 to Protect Endothelial Cells.","date":"2026","source":"Clinical laboratory","url":"https://pubmed.ncbi.nlm.nih.gov/42159105","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.10.25.684549","title":"AP-1 mediated chromatin changes govern alveolar type 2 cell transition in lung injury-repair","date":"2025-10-26","source":"bioRxiv","url":"https://doi.org/10.1101/2025.10.25.684549","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.04.17.649313","title":"A Computational Framework for Analysis of cfDNA Fragmentation Profiles","date":"2025-04-18","source":"bioRxiv","url":"https://doi.org/10.1101/2025.04.17.649313","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":20254,"output_tokens":4951,"usd":0.067514,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":13147,"output_tokens":4160,"usd":0.084867,"stage2_stop_reason":"end_turn"},"total_usd":0.152381,"stage1_batch_id":"msgbatch_01ACAxJQabDsht39JXeWG22d","stage2_batch_id":"msgbatch_01QEU22A9U5sbPTgsMzTmvSf","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2006,\n      \"finding\": \"The CLDN4 promoter contains two Sp1 binding sites both required for promoter activity; cells overexpressing CLDN4 exhibit low DNA methylation and high histone H3 acetylation of the critical Sp1-containing promoter region, whereas CLDN4-negative cells show the reverse; treatment with demethylating and/or acetylating agents induces CLDN4 expression in CLDN4-negative cells.\",\n      \"method\": \"Promoter deletion analysis, site-directed mutagenesis of Sp1 sites, ChIP for histone H3 acetylation, bisulfite sequencing for DNA methylation, pharmacological treatment with demethylating/acetylating agents\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — promoter mutagenesis plus multiple orthogonal epigenetic methods (methylation, acetylation ChIP, pharmacological reversal) in one study\",\n      \"pmids\": [\"16714763\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"CLDN4 repression in normal ovarian epithelial cells is associated with bivalent histone modifications (H3K4me3 + H3K27me3) and DNA hypermethylation; during ovarian tumorigenesis, derepression correlates with loss of H3K27me3 and H4K20me3; combined DNA demethylation and histone acetylation treatment robustly reverses CLDN4 repression, whereas loss of H3K27me3 alone is insufficient.\",\n      \"method\": \"ChIP for H3K4me3, H3K27me3, H4K20me3; bisulfite sequencing; pharmacological treatment (demethylating + acetylating agents); gene expression analysis\",\n      \"journal\": \"Carcinogenesis\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal chromatin and methylation methods, functional pharmacological reversal, single lab\",\n      \"pmids\": [\"20053926\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"C-CPE (C-terminus of Clostridium perfringens enterotoxin) binds CLDN4 as a specific receptor, decreases CLDN4 protein expression, and relocates CLDN4 from cell-cell contact regions to the cytoplasm in ovarian cancer cells; C-CPE sensitizes EOC cells to Taxol and Carboplatin in a CLDN4-dependent manner and activates the ubiquitin-proteasome pathway.\",\n      \"method\": \"Quantitative RT-PCR, immunofluorescence, Western blot, 3D culture and monolayer culture assays, xenograft mouse model, oligonucleotide microarray\",\n      \"journal\": \"Clinical cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro and in vivo functional assays with CLDN4-dependent rescue, single lab, multiple orthogonal readouts\",\n      \"pmids\": [\"21123456\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"STAT2 DNA-binding activity transcriptionally activates CLDN4 as an interferon-stimulated gene; siRNA knockdown of CLDN4 in fibroblasts reduces IFN-alpha/beta-induced antiproliferative and antiviral responses, consistent with cells expressing a DNA-binding mutant of STAT2.\",\n      \"method\": \"siRNA knockdown, gene microarray, site-directed mutagenesis of STAT2, biological IFN response assays (antiproliferative, antiviral)\",\n      \"journal\": \"Journal of interferon & cytokine research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — siRNA knockdown with two orthogonal functional readouts, corroborated by STAT2 mutagenesis, single lab\",\n      \"pmids\": [\"17651017\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"PAK4 phosphorylates CEBPB on Thr-235, which then binds the -1093 to -991 bp region of the CLDN4 promoter to transcriptionally upregulate CLDN4 expression, promoting breast cancer cell migration and invasion; restoration of CLDN4 in PAK4-knockdown cells reverses inhibition of migration and invasion.\",\n      \"method\": \"PAK4 siRNA knockdown, CEBPB promoter ChIP/binding assay, luciferase reporter for CLDN4 promoter, rescue overexpression of CLDN4, migration/invasion assays\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP/reporter plus rescue experiment, single lab, multiple orthogonal methods\",\n      \"pmids\": [\"30808546\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"ZNF703 directly binds the CLDN4 promoter and transactivates CLDN4 expression to promote EMT and HCC metastasis; downregulation of CLDN4 attenuates ZNF703-mediated metastasis, and CLDN4 upregulation reverses the reduced metastasis seen upon ZNF703 knockdown.\",\n      \"method\": \"ChIP for ZNF703 binding to CLDN4 promoter, loss- and gain-of-function experiments, in vitro and in vivo metastasis assays, epistasis rescue\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP and rescue epistasis, in vitro and in vivo, single lab\",\n      \"pmids\": [\"32269215\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"TGF-β signaling upregulates CLDN4 expression in GBM cells and promotes nuclear translocation of CLDN4; CLDN4 modulates the TNF-α/NF-κB signaling pathway; inhibition of CLDN4 suppresses mesenchymal transition, invasion, and migration, and TGF-β inhibitor ITD-1 downregulates CLDN4 and inhibits invasion.\",\n      \"method\": \"CLDN4 knockdown/overexpression, subcellular fractionation/imaging for nuclear translocation, TGF-β pathway inhibitor treatment, in vitro invasion/migration assays, in vivo tumor growth assays\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — nuclear translocation validated with fractionation, epistasis via inhibitor treatment plus functional assays, single lab\",\n      \"pmids\": [\"35418179\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"RBM15, an m6A RNA methyltransferase, suppresses CLDN4 expression through m6A-mediated epigenetic inhibition in hepatic cells; RBM15 overexpression increases insulin resistance, and this effect is mediated through m6A regulation of CLDN4.\",\n      \"method\": \"MeRIP sequencing, mRNA-seq, RBM15 overexpression/knockdown, glucose uptake tests, Western blot, immunohistochemistry\",\n      \"journal\": \"Molecular medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — MeRIP-seq identifies m6A mark on CLDN4 mRNA, functional glucose uptake assay, single lab\",\n      \"pmids\": [\"36803098\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"IGF2BP3 interacts with CLDN4 mRNA and augments its stability in an m6A-dependent manner; CLDN4 upregulation by IGF2BP3 activates NF-κB signaling in gallbladder cancer cells; restoration of CLDN4 reverses the inhibitory effect of IGF2BP3 knockdown on gallbladder cancer progression.\",\n      \"method\": \"RNA immunoprecipitation (RIP), m6A methylation assays, IGF2BP3 knockdown/overexpression, NF-κB pathway analysis, rescue CLDN4 overexpression, in vitro/in vivo tumor assays\",\n      \"journal\": \"Translational oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — RIP plus m6A assay and rescue epistasis, in vitro and in vivo, single lab\",\n      \"pmids\": [\"37643553\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"FOXA1 transcriptionally activates CLDN4 by binding its promoter; CLDN4 overexpression induces activation of the PI3K/AKT pathway; celastrol inhibits GC progression by downregulating FOXA1, thereby reducing CLDN4 and impeding PI3K/AKT.\",\n      \"method\": \"Luciferase reporter assay for FOXA1 binding to CLDN4 promoter, FOXA1/CLDN4 overexpression/knockdown, Western blot for PI3K/AKT phosphorylation, MTT and Transwell assays\",\n      \"journal\": \"Toxicology research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — luciferase reporter plus functional pathway assays, single lab\",\n      \"pmids\": [\"37397926\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"CLDN4 knockdown in AML cells inhibits mRNA expression of PIK3R3 and MAP2K2, suppressing AKT and ERK1/2 activation; partial rescue of cell viability by AKT activator SC79 confirms AKT as a downstream effector; CLDN4 promotes AML cell growth and suppresses apoptosis.\",\n      \"method\": \"CLDN4 siRNA knockdown, Western blot for pAKT and pERK1/2, SC79 (AKT activator) rescue experiment, cell viability and apoptosis assays\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pathway activation confirmed by pharmacological rescue, single lab\",\n      \"pmids\": [\"35760010\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"In bladder urothelial carcinoma cells, demethylation-induced excess CLDN4 that is not integrated into tight junctions (TJ-unintegrated CLDN4 monomer) binds integrin β1, increases FAK phosphorylation, and promotes stemness, drug resistance, and metastatic ability; CLDN4 knockdown reduces FAK phosphorylation.\",\n      \"method\": \"Co-immunoprecipitation/pulldown of CLDN4 with integrin β1, Western blot for FAK phosphorylation, CLDN4 knockdown, demethylating agent (AZA) treatment, fractionation to identify TJ-unintegrated CLDN4\",\n      \"journal\": \"International journal of molecular sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — Co-IP binding assay, FAK phosphorylation readout with knockdown rescue, single lab\",\n      \"pmids\": [\"35742959\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"CLDN4 is palmitoylated at cysteine residues C104 and C107; this palmitoylation regulates ubiquitination at lysine K103, inhibits clathrin-mediated endocytosis, and sustains CLDN4 anchoring within lipid rafts; lipid raft-anchored CLDN4 drives mobilization of contactin-1 to lipid rafts and activates Notch signaling, promoting hepatic-to-biliary lineage transition and lenvatinib resistance in HCC.\",\n      \"method\": \"Palmitoylation site mutagenesis (C104/C107), ubiquitination assays, lipid raft fractionation, clathrin-mediated endocytosis assays, Co-IP for contactin-1, Notch pathway reporter assays, drug resistance assays\",\n      \"journal\": \"Cell reports. Medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — site-directed mutagenesis of palmitoylation sites combined with ubiquitination assays, lipid raft fractionation, and functional Notch pathway readouts in one study\",\n      \"pmids\": [\"40592346\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"CLDN4 knockout in SCLC cells promotes cell proliferation by accelerating cell cycle progression; CLDN4 knockout upregulates SAA1 which partly mediates the proliferation-promoting effect; CLDN4 expression is directly regulated by SP1, with DNA methylation also contributing to transcriptional regulation.\",\n      \"method\": \"CRISPR/CRISPR-based CLDN4 knockout, RNA-seq, SP1 binding assays, DNA methylation analysis, SAA1 knockdown rescue, cell cycle and proliferation assays\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — knockout with RNA-seq pathway identification and SP1 binding, single lab\",\n      \"pmids\": [\"41016339\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"CLDN4 overexpression in corpus cavernosum smooth muscle cells activates the JNK signaling pathway, increases fibrotic protein expression, and impairs erectile function in vivo; hypoxia increases CLDN4 expression, and Cldn4 overexpression in rat corpus cavernosum increases local fibrosis and impairs erectile function.\",\n      \"method\": \"Lentiviral Cldn4 overexpression in vitro and in vivo (rat corpus cavernosum), Western blot for JNK pathway and fibrotic proteins, functional erectile assays, immunofluorescence\",\n      \"journal\": \"The journal of sexual medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — gain-of-function in vitro and in vivo with pathway readout, single lab\",\n      \"pmids\": [\"38477100\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"CLDN4 knockdown in cerulein-stimulated pancreatic acinar cells reduces ROS, iron accumulation, inflammatory cytokines (TNF-α, IL-6, IL-17), restores GPX4 levels, and reduces ACSL4 expression, suppressing ferroptosis; CLDN4 knockdown decreases JAK2/STAT3 pathway activation, and combined CLDN4 knockdown with JAK2 inhibitor AG490 provides additive protective effects in AP models.\",\n      \"method\": \"shRNA CLDN4 knockdown, RNA-seq, Western blot for JAK2/STAT3, GPX4, ACSL4, ELISA for cytokines, flow cytometry for ROS, cerulein AP mouse model, pharmacological JAK2 inhibition (AG490) epistasis\",\n      \"journal\": \"Functional & integrative genomics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function with pharmacological epistasis (AG490), in vitro and in vivo, single lab\",\n      \"pmids\": [\"40892111\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"SPTBN2 interacts with CLDN4 to promote endometrial cancer cell migration and invasion via the PI3K/AKT pathway; CLDN4 overexpression partially reverses the decrease in migration/invasion caused by SPTBN2 knockdown, and CLDN4 is itself upregulated in EEC and promotes metastasis.\",\n      \"method\": \"Co-IP/interaction assay between SPTBN2 and CLDN4, SPTBN2/CLDN4 knockdown and overexpression, PI3K/AKT pathway western blot, migration/invasion assays, rescue epistasis\",\n      \"journal\": \"Cell death discovery\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single Co-IP with functional rescue, single lab, single paper\",\n      \"pmids\": [\"34887379\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"AP-1 transcription factors (JUNB, FOSB, FOS) are required for CLDN4 induction in transitional AT2 cells following viral lung injury; CLDN4+ AT2 cells represent a KRT8-high transitional substate with chromatin enriched for AP-1 motifs; AP-1 promotes AT2 cell dispersion and senescence signaling toward fibroblasts.\",\n      \"method\": \"Mouse genetics (AP-1 member conditional knockout), multiomics (joint transcriptomic-epigenomic profiling), immunostaining, chromatin accessibility (ATAC-seq) for AP-1 motif enrichment, in vivo viral injury model\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — mouse genetic knockouts with multiomics, single lab preprint\",\n      \"pmids\": [],\n      \"is_preprint\": true\n    }\n  ],\n  \"current_model\": \"CLDN4 is a tight junction protein whose expression is transcriptionally regulated by Sp1, FOXA1, CEBPB (phosphorylated by PAK4), ZNF703, ELF3, and AP-1 factors, and epigenetically controlled by DNA methylation and repressive histone marks (H3K27me3, H4K20me3); STAT2 DNA-binding activity induces CLDN4 as an interferon-stimulated gene mediating antiviral and antiproliferative IFN responses; post-translationally, CLDN4 palmitoylation at C104/C107 regulates its ubiquitination at K103, prevents clathrin-mediated endocytosis, anchors it in lipid rafts where it mobilizes contactin-1 and activates Notch signaling; when not integrated into tight junctions, CLDN4 binds integrin β1 to activate FAK and downstream PI3K/AKT; additionally CLDN4 can translocate to the nucleus in response to TGF-β to modulate TNF-α/NF-κB signaling, and promotes ferroptosis-related inflammation via the JAK2/STAT3 pathway, while its mRNA stability is enhanced by the m6A readers IGF2BP3 and RBM15.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"CLDN4 is a tight junction protein whose expression is tightly controlled by transcriptional and epigenetic mechanisms and whose non-junctional pools act as signaling effectors in epithelial transformation and tissue injury [#0, #11]. Its promoter is driven by two essential Sp1 sites and is silenced in normal cells by DNA hypermethylation, repressive histone marks (H3K27me3, H4K20me3) and bivalent chromatin, with combined demethylation and histone acetylation robustly derepressing the gene [#0, #1, #13]. A range of sequence-specific factors converge on the CLDN4 promoter to activate it in disease contexts: PAK4-phosphorylated CEBPB, FOXA1, ZNF703, AP-1 factors, and STAT2, the latter establishing CLDN4 as an interferon-stimulated gene that mediates antiviral and antiproliferative IFN responses [#3, #4, #5, #9, #17]. Post-transcriptionally, CLDN4 mRNA fate is set by m6A: IGF2BP3 stabilizes the transcript while RBM15-dependent m6A methylation suppresses it [#7, #8]. A key mechanistic theme is that CLDN4 not integrated into tight junctions acts as a signaling hub: such monomeric CLDN4 binds integrin \\u03b21 to drive FAK phosphorylation and PI3K/AKT activation, promoting stemness, drug resistance and metastasis [#11, #9, #16]. Palmitoylation at C104/C107 controls CLDN4 fate by regulating ubiquitination at K103, blocking clathrin-mediated endocytosis and anchoring CLDN4 in lipid rafts, where it mobilizes contactin-1 and activates Notch signaling to drive lineage transition and lenvatinib resistance [#12]. CLDN4 also feeds NF-\\u03baB signaling, undergoes TGF-\\u03b2-induced nuclear translocation, drives JAK2/STAT3-dependent ferroptotic inflammation, and engages MAPK pathways (ERK1/2, JNK) across cancer, pancreatitis and fibrosis models [#6, #8, #10, #15, #14].\"\n  ,\n  \"teleology\": [\n    {\n      \"year\": 2006,\n      \"claim\": \"Established the core transcriptional control of CLDN4, showing its promoter depends on Sp1 sites whose activity is gated by DNA methylation and histone acetylation status.\",\n      \"evidence\": \"Promoter deletion/mutagenesis of Sp1 sites, ChIP for H3 acetylation, bisulfite sequencing and pharmacological reversal in cell lines\",\n      \"pmids\": [\"16714763\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not address upstream signals that recruit Sp1 or remodel the locus\", \"Tested in a limited set of cell lines\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Defined CLDN4 as an interferon-stimulated gene, linking it to STAT2-driven antiviral and antiproliferative IFN responses beyond a purely structural role.\",\n      \"evidence\": \"siRNA knockdown with antiproliferative/antiviral readouts plus STAT2 DNA-binding mutagenesis in fibroblasts\",\n      \"pmids\": [\"17651017\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct STAT2 occupancy at the CLDN4 promoter not shown\", \"Molecular role of CLDN4 within the IFN response uncharacterized\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Resolved the chromatin logic of CLDN4 silencing and derepression, showing bivalent and repressive histone marks plus DNA methylation cooperate, with loss of H3K27me3 alone being insufficient.\",\n      \"evidence\": \"ChIP for H3K4me3/H3K27me3/H4K20me3, bisulfite sequencing and pharmacological reversal in ovarian epithelial/tumor cells; C-CPE receptor binding and proteasomal degradation in EOC cells\",\n      \"pmids\": [\"20053926\", \"21123456\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Writers/erasers responsible for the mark switch not identified\", \"C-CPE degradation mechanism not connected to physiological turnover\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Identified specific transcription factors (PAK4-CEBPB, ZNF703) and a binding partner (SPTBN2) that activate CLDN4 to drive migration, invasion and metastasis, framing CLDN4 as a downstream pro-metastatic effector.\",\n      \"evidence\": \"Promoter ChIP/reporter, Co-IP, and CLDN4 rescue epistasis with migration/invasion and in vivo metastasis assays in breast, HCC and endometrial cancer models\",\n      \"pmids\": [\"30808546\", \"32269215\", \"34887379\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"SPTBN2-CLDN4 interaction rests on a single Co-IP without reciprocal validation\", \"How CLDN4 mechanistically promotes motility downstream of these factors not fully resolved\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Showed that non-junctional CLDN4 acts as a signaling node, binding integrin \\u03b21 to activate FAK and feeding PI3K/AKT and ERK pathways that promote stemness, drug resistance and survival.\",\n      \"evidence\": \"Co-IP of CLDN4 with integrin \\u03b21, FAK phosphorylation with knockdown, demethylation-induced TJ-unintegrated CLDN4 fractionation in bladder carcinoma; AKT/ERK pathway readouts with SC79 rescue in AML\",\n      \"pmids\": [\"35742959\", \"35760010\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of CLDN4-integrin \\u03b21 binding unknown\", \"Whether monomeric versus junctional CLDN4 is the relevant species in all contexts not established\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Demonstrated TGF-\\u03b2-induced nuclear translocation of CLDN4 and its modulation of TNF-\\u03b1/NF-\\u03baB signaling, indicating a non-membrane function in mesenchymal transition.\",\n      \"evidence\": \"Subcellular fractionation/imaging, TGF-\\u03b2 inhibitor treatment and invasion/migration assays in GBM cells\",\n      \"pmids\": [\"35418179\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Nuclear binding partners and direct transcriptional targets of CLDN4 not identified\", \"Mechanism of nuclear import unknown\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Established m6A as a post-transcriptional control layer for CLDN4, with opposing reader/writer effects: IGF2BP3 stabilizing the mRNA and RBM15 suppressing it.\",\n      \"evidence\": \"RIP/MeRIP and m6A assays with knockdown/overexpression, NF-\\u03baB analysis and CLDN4 rescue in gallbladder cancer; MeRIP-seq with glucose uptake assays in hepatic insulin resistance\",\n      \"pmids\": [\"37643553\", \"36803098\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Specific m6A sites on CLDN4 mRNA not mapped to function\", \"Reconciliation of stabilizing versus suppressive m6A outcomes across tissues not addressed\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Linked FOXA1-driven CLDN4 expression to PI3K/AKT activation, reinforcing CLDN4 as a transcriptionally inducible driver of oncogenic signaling.\",\n      \"evidence\": \"Luciferase reporter for FOXA1 binding, overexpression/knockdown and PI3K/AKT phosphorylation with functional assays in gastric cancer\",\n      \"pmids\": [\"37397926\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct molecular link between CLDN4 and PI3K/AKT components not defined\", \"Promoter occupancy by FOXA1 inferred from reporter rather than ChIP\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Defined a palmitoylation-ubiquitination switch that controls CLDN4 trafficking, anchoring it in lipid rafts to mobilize contactin-1 and activate Notch signaling, mechanistically explaining its role in lineage transition and drug resistance.\",\n      \"evidence\": \"C104/C107 palmitoylation-site mutagenesis, ubiquitination and lipid raft assays, endocytosis assays, Co-IP for contactin-1 and Notch reporters in HCC\",\n      \"pmids\": [\"40592346\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Palmitoyltransferase and deubiquitinase enzymes not identified\", \"Generality of the raft/Notch axis beyond HCC unknown\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Extended CLDN4 function to inflammatory ferroptosis and tissue fibrosis, implicating JAK2/STAT3 and JNK pathways in non-cancer pathology, and to cell cycle suppression via SAA1.\",\n      \"evidence\": \"shRNA knockdown with AG490 epistasis in cerulein pancreatitis; Cldn4 overexpression with JNK readout in rat corpus cavernosum; CRISPR knockout with RNA-seq/SAA1 rescue in SCLC\",\n      \"pmids\": [\"40892111\", \"38477100\", \"41016339\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether CLDN4 directly activates JAK2/STAT3 or JNK or acts indirectly is unresolved\", \"Context-dependent pro- versus anti-proliferative roles not reconciled\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How junctional versus non-junctional CLDN4 pools are partitioned and how a single tight junction protein integrates such diverse downstream pathways (integrin/FAK, Notch, NF-\\u03baB, JAK2/STAT3, JNK) into context-specific outcomes remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unifying structural model distinguishing signaling-competent CLDN4 conformers\", \"Direct biochemical link between CLDN4 and most downstream kinases unestablished\", \"Physiological (non-disease) function in normal epithelia underexplored in the corpus\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [0, 11]},\n      {\"term_id\": \"GO:0098631\", \"supporting_discovery_ids\": [11]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [11, 12]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [2]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [6]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [11, 12]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [3]},\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [7, 8]}\n    ],\n    \"complexes\": [\"tight junction\", \"lipid raft\"],\n    \"partners\": [\"ITGB1\", \"CNTN1\", \"SPTBN2\", \"IGF2BP3\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"tie","faith_supported":7,"faith_total":7,"faith_pct":100.0}}