{"gene":"SCNN1A","run_date":"2026-06-10T07:46:29","timeline":{"discoveries":[{"year":2005,"finding":"The histone H3 lysine-79 methyltransferase mDot1a binds the ENaCalpha (SCNN1A) promoter and hypermethylates histone H3 K79 at that locus, repressing ENaCalpha transcription; aldosterone downregulates mDot1a, causing H3 K79 hypomethylation and ENaCalpha trans-activation. Methyltransferase-dead mDot1a mutants abolished this repression, confirming catalytic requirement.","method":"RNA interference knockdown, EGFP-tagged overexpression, chromatin immunoprecipitation (ChIP), luciferase reporter assay, active-site mutagenesis of mDot1a methyltransferase domain","journal":"American journal of physiology. Cell physiology","confidence":"High","confidence_rationale":"Tier 1 / Strong — active-site mutagenesis abolishing effect plus ChIP, RNAi knockdown, and reporter assays in two orthogonal directions (KD and OE), single lab but multiple rigorous methods","pmids":["16236820"],"is_preprint":false},{"year":2006,"finding":"AF9 interacts physically with Dot1a both in vitro and in vivo (co-localizing in renal collecting duct cell nuclei) and co-occupies the ENaCalpha promoter; AF9 overexpression causes histone H3 Lys-79 hypermethylation at the ENaCalpha promoter and represses ENaCalpha transcription synergistically with Dot1a, while AF9 knockdown has the opposite effect. Aldosterone negatively regulates AF9 expression, integrating AF9 into the aldosterone-Dot1a-ENaCalpha signaling axis.","method":"Co-immunoprecipitation, GST pulldown (in vitro interaction), chromatin immunoprecipitation, RNA interference, luciferase reporter assay, confocal colocalization, RT-PCR/Western blot for AF9 levels after aldosterone","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP plus in vitro pulldown, ChIP, RNAi, and reporter assays across multiple orthogonal methods; replicates and extends prior Dot1a mechanism paper","pmids":["16636056"],"is_preprint":false},{"year":2003,"finding":"PPARgamma activation in human cortical collecting duct cells upregulates SGK1 (via a PPARgamma response element in the SGK1 promoter), which in turn increases cell-surface ENaCalpha protein levels, thereby enhancing ENaC-mediated sodium reabsorption.","method":"PPARgamma agonist/antagonist treatment, SGK1 mRNA measurement, electrophoretic mobility shift assay (EMSA) for PPARgamma binding to SGK1 promoter, cell-surface ENaCalpha quantification","journal":"FASEB journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — EMSA plus agonist/antagonist pharmacological dissection and surface protein quantification, single lab, no mutagenesis of the response element","pmids":["12923071"],"is_preprint":false},{"year":2011,"finding":"A homozygous missense mutation in SCNN1A (c.727T>C; p.Ser243Pro) in the extracellular loop of alpha-ENaC causes a significant decrease in ENaC channel activity in vitro, exacerbated by high Na+ load, identifying a novel functional domain in the extracellular loop. The reduction is partial, explaining transient/milder phenotype in a mature kidney.","method":"Xenopus laevis oocyte expression system with electrophysiology (two-electrode voltage clamp) to measure ENaC activity of mutant vs wild-type channel","journal":"American journal of physiology. Endocrinology and metabolism","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — in vitro reconstitution in oocytes with functional readout, single lab, single mutation studied","pmids":["21653223"],"is_preprint":false},{"year":2013,"finding":"A splice-site mutation in SCNN1A (c.1439+1G>C) leads to retention of intron 9 due to loss of the 5'-donor splice site, as demonstrated in a minigene construct, establishing the molecular consequence of this PHA1-causing mutation on SCNN1A mRNA processing.","method":"Minigene splicing assay with Sanger sequencing of resulting transcripts","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — direct minigene functional splice assay, single lab, single mutation","pmids":["23762408"],"is_preprint":false},{"year":2023,"finding":"The p.Phe226Cys missense mutation in the extracellular domain of SCNN1A causes ~83% reduction in ENaC channel activity when expressed in Xenopus oocytes, due to both reduced intrinsic open probability and decreased channel protein expression at the plasma membrane, explaining a mild/transient PHA1 phenotype.","method":"Xenopus laevis oocyte expression with two-electrode voltage clamp electrophysiology; Western blot for protein expression of mutant vs wild-type alpha-ENaC","journal":"American journal of physiology. Endocrinology and metabolism","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — in vitro oocyte reconstitution with electrophysiology plus Western blot quantification, single lab, single mutation","pmids":["37134141"],"is_preprint":false},{"year":2015,"finding":"The NF-κB subunits p65/p50, activated downstream of IKKβ, directly bind the ENaCalpha (SCNN1A) gene promoter and are required for aldosterone-induced ENaCalpha transcription in renal cortical collecting duct cells; IKKβ or p65 knockdown reduces aldosterone-induced ENaCalpha mRNA by >60%.","method":"siRNA knockdown of IKKβ and p65, chromatin immunoprecipitation (ChIP), EMSA, luciferase reporter assay in mpkCCDc14 cells","journal":"Journal of nephrology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP plus EMSA plus reporter assay plus RNAi, single lab, multiple orthogonal methods","pmids":["26385798"],"is_preprint":false},{"year":2020,"finding":"MLL2 (KMT2D), a histone H3K4 methyltransferase, associates with the glucocorticoid receptor (GR) in a ligand-independent manner and co-occupies glucocorticoid response elements at the ENaCalpha promoter upon dexamethasone stimulation, promoting chromatin accessibility and GR-mediated ENaCalpha transcription in ARPE-19 cells but not A549 cells.","method":"Co-immunoprecipitation of MLL2 with GR, chromatin immunoprecipitation (ChIP) for MLL2 and GR at GREs, FAIRE-qPCR for chromatin accessibility, siRNA knockdown of MLL2","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, ChIP, FAIRE-qPCR, and RNAi in combination, single lab, cell-type-specific context established","pmids":["32139118"],"is_preprint":false},{"year":2020,"finding":"FOXA1 functions as a pioneer factor enabling cell-type-specific glucocorticoid receptor-driven expression of ENaCalpha (SCNN1A) in ARPE-19 cells; FOXA1 depletion significantly reduces ENaCalpha expression in these cells, whereas ENaCalpha is not a GR target in A549 cells.","method":"RNA-seq, RT-qPCR, siRNA knockdown of FOXA1 in two cell lines","journal":"International journal of immunopathology and pharmacology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — RNAi with transcriptional readout, no ChIP for FOXA1 at ENaCalpha locus reported in abstract, single lab, single method for the key mechanistic claim","pmids":["32838581"],"is_preprint":false},{"year":2001,"finding":"1.56 kb of the mouse Scnn1a 5'-upstream promoter sequence is required for cell-specific expression and corticosteroid-mediated regulation in cortical collecting duct cells, as shown by transient transfection of serial deletion mutants; however, this sequence was insufficient to drive reporter gene expression in transgenic mice.","method":"Serial deletion mutagenesis of Scnn1a promoter, transient transfection reporter assay in CCD cells, transgenic mouse reporter assay","journal":"Biochimica et biophysica acta","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — deletion mutagenesis in cell-based reporter assay plus in vivo transgenic validation (negative), single lab, two orthogonal systems","pmids":["11406278"],"is_preprint":false},{"year":2025,"finding":"Conditional endothelial-specific knockout of ENaC-alpha (enENaC-α KO) in mice increases capillary leak and NOX2 (NADPH oxidase 2) expression in lung endothelium during pneumococcal infection/PLY challenge; direct ENaC activation by TIP peptide reduces NOX2 activity and blunts capillary hyperpermeability, placing ENaC-alpha as a negative regulator of endothelial NOX2-mediated oxidative stress in acute lung injury.","method":"Conditional endothelial Scnn1a knockout mouse model, Evans blue dye permeability assay, electrical cell substrate impedance sensing in monolayers, Western blot for phospho-p47phox, DuoLink colocalization, chemiluminescence for superoxide, pharmacological TIP peptide activation","journal":"American journal of respiratory cell and molecular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — conditional KO with functional permeability readout plus in vitro mechanistic assays, single lab, multiple orthogonal methods","pmids":["39405473"],"is_preprint":false},{"year":2025,"finding":"NDUFS1 deficiency in alveolar epithelial cells reduces ENaCalpha expression through mitochondrial complex I dysfunction and decreased NAD+ production; supplementing NAD+ (via olaparib) restores ENaCalpha abundance and improves alveolar fluid clearance, placing SCNN1A downstream of the NDUFS1-NAD+ axis in lung epithelium.","method":"Proteomic screening, scRNA-seq mining, NDUFS1 KO in alveolar epithelial cells, NAD+ supplementation (olaparib), alveolar fluid clearance measurement, RT-qPCR/Western blot for ENaCalpha","journal":"International journal of medical sciences","confidence":"Low","confidence_rationale":"Tier 3 / Weak — KO with functional readout and pharmacological rescue, single lab, abstract-level detail insufficient to confirm direct mechanistic link vs indirect metabolic effect","pmids":["40860777"],"is_preprint":false},{"year":2025,"finding":"Taurine deficiency in mice (Csad-/- on taurine-free diet) downregulates Scnn1a expression in the uterus during the implantation window, leading to uterine luminal fluid retention and failed embryo implantation; progesterone promotes and estradiol inhibits uterine Csad/taurine synthesis, placing SCNN1A in a steroid hormone-taurine axis controlling uterine fluid resorption.","method":"Csad knockout mouse model on taurine-free diet, RT-qPCR and protein expression for Scnn1a and Aqp8, embryo implantation rate measurement, hormonal manipulation","journal":"Biology of reproduction","confidence":"Low","confidence_rationale":"Tier 3 / Weak — KO model with phenotypic and expression readout, correlative link between taurine and Scnn1a expression, single lab, mechanistic pathway not directly validated","pmids":["39428112"],"is_preprint":false}],"current_model":"SCNN1A encodes the alpha subunit of the epithelial sodium channel (ENaC), which is transcriptionally repressed by the Dot1a-AF9 complex via histone H3 Lys-79 hypermethylation at its promoter in an aldosterone-sensitive manner, and activated through NF-κB (p65/p50) binding to its promoter, MLL2-mediated H3K4 methylation enabling glucocorticoid receptor access, and FOXA1 pioneer factor activity; at the protein level, the alpha subunit forms a channel whose activity is reduced by missense mutations in its extracellular loop (e.g., Ser243Pro, Phe226Cys), and in endothelial cells ENaC-alpha restrains NOX2-mediated oxidative stress and capillary leak during acute lung injury."},"narrative":{"mechanistic_narrative":"SCNN1A encodes the alpha subunit of the epithelial sodium channel (ENaC), which mediates apical sodium reabsorption in aldosterone-responsive epithelia and is controlled by a layered transcriptional program at its promoter [PMID:16236820, PMID:26385798]. In the basal state ENaCalpha transcription is repressed by the histone H3 Lys-79 methyltransferase Dot1a, which binds the promoter and hypermethylates H3K79; aldosterone downregulates Dot1a to relieve this repression and trans-activate the gene [PMID:16236820]. Dot1a acts together with its physical partner AF9, which co-occupies the promoter and represses transcription synergistically, and is itself negatively regulated by aldosterone, integrating AF9 into the aldosterone-Dot1a-ENaCalpha axis [PMID:16636056]. Aldosterone-induced transcription further requires direct binding of the NF-kappaB subunits p65/p50, activated downstream of IKKbeta, to the promoter [PMID:26385798], while glucocorticoid-driven expression depends on cell-type-specific access of the glucocorticoid receptor enabled by MLL2 (KMT2D)-mediated H3K4 methylation and chromatin opening [PMID:32139118]. Steroid signaling also acts through SGK1, induced by PPARgamma, to increase cell-surface ENaCalpha and sodium reabsorption [PMID:12923071]. At the protein level, missense mutations in the alpha-subunit extracellular loop (Ser243Pro, Phe226Cys) reduce channel activity through lowered open probability and decreased surface expression, and a splice-site mutation causing intron 9 retention disrupts mRNA processing, defining molecular bases of pseudohypoaldosteronism type 1 (PHA1) [PMID:21653223, PMID:23762408, PMID:37134141]. Beyond renal epithelia, endothelial ENaC-alpha restrains NOX2-mediated oxidative stress and capillary leak during acute lung injury [PMID:39405473].","teleology":[{"year":2001,"claim":"Before any trans-acting factors were known, it was unclear what genomic region conferred cell-specific and corticosteroid-responsive Scnn1a expression; mapping the promoter localized the regulatory information.","evidence":"Serial promoter deletion reporter assays in cortical collecting duct cells plus transgenic mouse reporters","pmids":["11406278"],"confidence":"Medium","gaps":["1.56 kb sequence insufficient to drive expression in transgenic mice, implying missing distal elements","no specific transcription factors identified","individual response elements not resolved"]},{"year":2003,"claim":"It was unknown how nuclear-receptor signaling beyond aldosterone tuned ENaCalpha surface levels; PPARgamma was shown to act indirectly via SGK1 induction to raise channel abundance at the membrane.","evidence":"PPARgamma agonist/antagonist treatment, EMSA for PPARgamma at the SGK1 promoter, and cell-surface ENaCalpha quantification in human CCD cells","pmids":["12923071"],"confidence":"Medium","gaps":["SGK1 promoter response element not confirmed by mutagenesis","mechanism linking SGK1 to surface ENaCalpha not dissected here","in vivo relevance not tested"]},{"year":2005,"claim":"The chromatin mechanism for basal ENaCalpha repression and its aldosterone-sensitive release was unknown; Dot1a-mediated H3K79 hypermethylation was identified as the repressive mark and aldosterone as a Dot1a downregulator.","evidence":"RNAi knockdown, EGFP-tagged overexpression, ChIP, luciferase reporters, and methyltransferase active-site mutagenesis of mDot1a","pmids":["16236820"],"confidence":"High","gaps":["how aldosterone downregulates Dot1a not defined","demethylase removing the mark not identified","single-locus study"]},{"year":2006,"claim":"Dot1a was not known to require a recruiting partner; AF9 was shown to physically bind Dot1a, co-occupy the promoter, and synergize in repression under aldosterone control.","evidence":"Reciprocal Co-IP, GST pulldown, ChIP, RNAi, luciferase reporters, and confocal colocalization in renal collecting duct cells","pmids":["16636056"],"confidence":"High","gaps":["targeting of AF9 to the promoter sequence not mapped","mechanism of aldosterone-driven AF9 downregulation unresolved","in vivo confirmation absent"]},{"year":2011,"claim":"Whether the alpha-subunit extracellular loop was functionally critical was unclear; a homozygous Ser243Pro mutation was shown to partially reduce channel activity, defining a new functional domain.","evidence":"Xenopus oocyte two-electrode voltage clamp of mutant vs wild-type channel","pmids":["21653223"],"confidence":"Medium","gaps":["single mutation in a heterologous system","in vivo channel behavior not assessed","structural basis of the loop's role not resolved"]},{"year":2013,"claim":"The molecular consequence of a PHA1-associated splice-site variant was undefined; a minigene assay showed loss of the 5' donor site causes intron 9 retention.","evidence":"Minigene splicing assay with Sanger sequencing of transcripts","pmids":["23762408"],"confidence":"Medium","gaps":["effect on protein and channel function not tested","single mutation","patient transcript not directly analyzed"]},{"year":2015,"claim":"The signaling required for aldosterone-induced transcription beyond chromatin marks was incomplete; NF-kappaB p65/p50 downstream of IKKbeta was shown to bind the promoter and be required for induction.","evidence":"siRNA knockdown of IKKbeta and p65, ChIP, EMSA, and luciferase reporters in mpkCCDc14 cells","pmids":["26385798"],"confidence":"Medium","gaps":["integration with the Dot1a-AF9 axis not mechanistically linked","specific NF-kappaB site not mutated","single cell model"]},{"year":2020,"claim":"How glucocorticoid receptor accessed the ENaCalpha promoter in a cell-type-specific manner was unknown; MLL2-mediated H3K4 methylation and FOXA1 pioneer activity were identified as enabling factors.","evidence":"Co-IP of MLL2 with GR, ChIP, FAIRE-qPCR for accessibility, and siRNA of MLL2 and FOXA1 across ARPE-19 versus A549 cells","pmids":["32139118","32838581"],"confidence":"Medium","gaps":["FOXA1 occupancy at the ENaCalpha locus not shown by ChIP","basis of cell-type selectivity incompletely defined","interplay with the aldosterone/NF-kappaB axes untested"]},{"year":2023,"claim":"The mechanism by which extracellular-domain mutations reduce channel function was unclear; Phe226Cys was shown to lower both open probability and plasma-membrane expression.","evidence":"Xenopus oocyte voltage clamp electrophysiology plus Western blot for surface/total protein","pmids":["37134141"],"confidence":"Medium","gaps":["single mutation in a heterologous system","trafficking defect mechanism not defined","no patient tissue validation"]},{"year":2025,"claim":"A role for ENaC-alpha outside transporting epithelia was unknown; endothelial-specific deletion revealed it restrains NOX2-driven oxidative stress and capillary leak in acute lung injury.","evidence":"Conditional endothelial Scnn1a knockout mice with Evans blue permeability, impedance sensing, phospho-p47phox and superoxide assays, and TIP peptide activation","pmids":["39405473"],"confidence":"Medium","gaps":["how ENaC-alpha mechanistically suppresses NOX2 not defined","channel-dependence of the effect not established","single disease model"]},{"year":2025,"claim":"Upstream metabolic and hormonal inputs to SCNN1A in non-renal tissues were unexplored; the NDUFS1-NAD+ axis in lung epithelium and a progesterone/estradiol-taurine axis in uterus were linked to ENaCalpha abundance.","evidence":"NDUFS1 KO with NAD+ rescue and fluid clearance in alveolar cells, and Csad knockout mice with uterine Scnn1a expression and implantation readouts","pmids":["40860777","39428112"],"confidence":"Low","gaps":["direct versus indirect effect on SCNN1A not distinguished","no demonstrated transcriptional mechanism linking these axes to the promoter","correlative expression changes only"]},{"year":null,"claim":"How the repressive Dot1a-AF9 chromatin axis, the activating NF-kappaB and GR/MLL2/FOXA1 programs, and SGK1-driven surface trafficking are coordinated into a single integrated control of ENaCalpha expression and activity remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["no unified model linking the repressive and activating transcriptional inputs","stoichiometry and assembly of the alpha subunit into functional channels not addressed in the corpus","in vivo integration of endothelial and epithelial roles untested"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0005215","term_label":"transporter activity","supporting_discovery_ids":[3,5]},{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[3,5]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[2,5]}],"pathway":[{"term_id":"R-HSA-382551","term_label":"Transport of small molecules","supporting_discovery_ids":[3,5]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[0,1,6,7]},{"term_id":"R-HSA-4839726","term_label":"Chromatin organization","supporting_discovery_ids":[0,1,7]}],"complexes":["epithelial sodium channel (ENaC)"],"partners":["DOT1L","AF9","RELA","NFKB1","NR3C1","KMT2D","FOXA1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P37088","full_name":"Epithelial sodium channel subunit alpha","aliases":["Alpha-NaCH","Amiloride-sensitive sodium channel subunit alpha","Nonvoltage-gated sodium channel 1 subunit alpha","SCNEA","Sodium channel epithelial 1 subunit alpha"],"length_aa":669,"mass_kda":75.7,"function":"This is one of the three pore-forming subunits of the heterotrimeric epithelial sodium channel (ENaC), a critical regulator of sodium balance and fluid homeostasis (PubMed:30251954, PubMed:32729833, PubMed:8023962, PubMed:8278374, PubMed:9792722). ENaC operates in epithelial tissues, where it mediates the electrodiffusion of sodium ions from extracellular fluid through the apical membrane of cells, with water following osmotically (PubMed:24124190, PubMed:28710092, PubMed:8278374). It plays a key role in maintaining sodium homeostasis through electrogenic sodium reabsorption in the kidneys (PubMed:12107247). Additionally, ENaC is essential for airway surface liquid homeostasis, which is crucial for proper mucus clearance (PubMed:24124190, PubMed:28710092) Not functional","subcellular_location":"Apical cell membrane; Cell projection, cilium; Cytoplasmic granule; Cytoplasm; Cytoplasmic vesicle, secretory vesicle, acrosome; Cell projection, cilium, flagellum","url":"https://www.uniprot.org/uniprotkb/P37088/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/SCNN1A","classification":"Not Classified","n_dependent_lines":1,"n_total_lines":1208,"dependency_fraction":0.0008278145695364238},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/SCNN1A","total_profiled":1310},"omim":[{"mim_id":"620125","title":"PSEUDOHYPOALDOSTERONISM, TYPE IB2, AUTOSOMAL RECESSIVE; PHA1B2","url":"https://www.omim.org/entry/620125"},{"mim_id":"618537","title":"ARGININE VASOPRESSIN-INDUCED PROTEIN 1; AVPI1","url":"https://www.omim.org/entry/618537"},{"mim_id":"618126","title":"LIDDLE SYNDROME 3; LIDLS3","url":"https://www.omim.org/entry/618126"},{"mim_id":"614492","title":"PSEUDOHYPOALDOSTERONISM, TYPE IIC; PHA2C","url":"https://www.omim.org/entry/614492"},{"mim_id":"613071","title":"BRONCHIECTASIS WITH OR WITHOUT ELEVATED SWEAT CHLORIDE 3; BESC3","url":"https://www.omim.org/entry/613071"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Uncertain","locations":[{"location":"Vesicles","reliability":"Uncertain"},{"location":"Annulus","reliability":"Uncertain"},{"location":"Acrosome","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"choroid plexus","ntpm":184.9},{"tissue":"salivary gland","ntpm":149.7}],"url":"https://www.proteinatlas.org/search/SCNN1A"},"hgnc":{"alias_symbol":["ENaCalpha"],"prev_symbol":["SCNN1"]},"alphafold":{"accession":"P37088","domains":[{"cath_id":"-","chopping":"56-118_548-574","consensus_level":"medium","plddt":82.872,"start":56,"end":574},{"cath_id":"2.60.470.10","chopping":"122-139_265-538","consensus_level":"medium","plddt":89.0759,"start":122,"end":538}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P37088","model_url":"https://alphafold.ebi.ac.uk/files/AF-P37088-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P37088-F1-predicted_aligned_error_v6.png","plddt_mean":74.56},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=SCNN1A","jax_strain_url":"https://www.jax.org/strain/search?query=SCNN1A"},"sequence":{"accession":"P37088","fasta_url":"https://rest.uniprot.org/uniprotkb/P37088.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P37088/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P37088"}},"corpus_meta":[{"pmid":"16636056","id":"PMC_16636056","title":"Dot1a-AF9 complex mediates histone H3 Lys-79 hypermethylation and repression of ENaCalpha in an aldosterone-sensitive manner.","date":"2006","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/16636056","citation_count":142,"is_preprint":false},{"pmid":"12923071","id":"PMC_12923071","title":"PPARgamma activation enhances cell surface ENaCalpha via up-regulation of SGK1 in human collecting duct cells.","date":"2003","source":"FASEB journal : official publication of the Federation of American Societies for Experimental Biology","url":"https://pubmed.ncbi.nlm.nih.gov/12923071","citation_count":90,"is_preprint":false},{"pmid":"16236820","id":"PMC_16236820","title":"Aldosterone-sensitive repression of ENaCalpha transcription by a histone H3 lysine-79 methyltransferase.","date":"2005","source":"American journal of physiology. 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molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/39405473","citation_count":6,"is_preprint":false},{"pmid":"40787466","id":"PMC_40787466","title":"Integrative single-cell and exosomal multi-omics uncovers SCNN1A and EFNA1 as non-invasive biomarkers and drivers of ovarian cancer metastasis.","date":"2025","source":"Frontiers in immunology","url":"https://pubmed.ncbi.nlm.nih.gov/40787466","citation_count":5,"is_preprint":false},{"pmid":"32139118","id":"PMC_32139118","title":"MLL2 regulates glucocorticoid receptor-mediated transcription of ENACα in human retinal pigment epithelial cells.","date":"2020","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/32139118","citation_count":5,"is_preprint":false},{"pmid":"37134141","id":"PMC_37134141","title":"A mild and transient form of autosomal recessive pseudohypoaldosteronism type 1 caused by a novel mutation in the SCNN1A gene.","date":"2023","source":"American journal of physiology. Endocrinology and metabolism","url":"https://pubmed.ncbi.nlm.nih.gov/37134141","citation_count":4,"is_preprint":false},{"pmid":"23813355","id":"PMC_23813355","title":"A case of SCNN1A splicing mutation presenting as mild systemic pseudohypoaldosteronism type 1.","date":"2013","source":"Journal of pediatric endocrinology & metabolism : JPEM","url":"https://pubmed.ncbi.nlm.nih.gov/23813355","citation_count":4,"is_preprint":false},{"pmid":"32838581","id":"PMC_32838581","title":"Cell-specific expression of ENACα gene by FOXA1 in the glucocorticoid receptor pathway.","date":"2020","source":"International journal of immunopathology and pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/32838581","citation_count":3,"is_preprint":false},{"pmid":"36336351","id":"PMC_36336351","title":"Clinical and genetic characteristics of the patients with hypertension and hypokalemia carrying a novel SCNN1A mutation.","date":"2022","source":"Scandinavian journal of clinical and laboratory investigation","url":"https://pubmed.ncbi.nlm.nih.gov/36336351","citation_count":1,"is_preprint":false},{"pmid":"40860777","id":"PMC_40860777","title":"NDUFS1 upregulates ENaCα by NAD+ to promote alveolar fluid clearance in acute lung injury.","date":"2025","source":"International journal of medical sciences","url":"https://pubmed.ncbi.nlm.nih.gov/40860777","citation_count":1,"is_preprint":false},{"pmid":"38963175","id":"PMC_38963175","title":"Novel homozygous mutation in SCNN1A gene in an Iranian boy with PHA1B.","date":"2024","source":"Journal of pediatric endocrinology & metabolism : JPEM","url":"https://pubmed.ncbi.nlm.nih.gov/38963175","citation_count":1,"is_preprint":false},{"pmid":"39295704","id":"PMC_39295704","title":"Pseudohypoaldosteronism Type 1b in a Saudi Female Infant Due to Homozygous Variant Gene Mutation in SCNN1A: A Case Report.","date":"2024","source":"Cureus","url":"https://pubmed.ncbi.nlm.nih.gov/39295704","citation_count":1,"is_preprint":false},{"pmid":"39428112","id":"PMC_39428112","title":"Taurine is essential for mouse uterine luminal fluid resorption during implantation window via the SCNN1A and AQP8 signaling†.","date":"2025","source":"Biology of reproduction","url":"https://pubmed.ncbi.nlm.nih.gov/39428112","citation_count":1,"is_preprint":false},{"pmid":"39236685","id":"PMC_39236685","title":"Liddle Syndrome with a SCNN1A Mutation: A Case Report and Literature Review.","date":"2024","source":"Kidney & blood pressure research","url":"https://pubmed.ncbi.nlm.nih.gov/39236685","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.11.04.686617","title":"Biased inter-columnar communication and short-term plasticity in mouse barrel cortex","date":"2025-11-06","source":"bioRxiv","url":"https://doi.org/10.1101/2025.11.04.686617","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2024.11.18.623996","title":"The evolutionary path of the epithelial sodium channel δ-subunit in Cetartiodactyla points to a role in sodium sensing","date":"2024-11-19","source":"bioRxiv","url":"https://doi.org/10.1101/2024.11.18.623996","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2024.09.17.613545","title":"Unique Transcriptomic Cell Types of the Granular Retrosplenial Cortex are Preserved Across Mice and Rats Despite Dramatic Changes in Key Marker Genes","date":"2024-09-17","source":"bioRxiv","url":"https://doi.org/10.1101/2024.09.17.613545","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":22670,"output_tokens":3663,"usd":0.061477,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":11318,"output_tokens":3935,"usd":0.077482,"stage2_stop_reason":"end_turn"},"total_usd":0.138959,"stage1_batch_id":"msgbatch_013LWZZT6MWaqB9qyTSeQBmU","stage2_batch_id":"msgbatch_0194ZzQaTGNpruoNJBbEMbt3","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2005,\n      \"finding\": \"The histone H3 lysine-79 methyltransferase mDot1a binds the ENaCalpha (SCNN1A) promoter and hypermethylates histone H3 K79 at that locus, repressing ENaCalpha transcription; aldosterone downregulates mDot1a, causing H3 K79 hypomethylation and ENaCalpha trans-activation. Methyltransferase-dead mDot1a mutants abolished this repression, confirming catalytic requirement.\",\n      \"method\": \"RNA interference knockdown, EGFP-tagged overexpression, chromatin immunoprecipitation (ChIP), luciferase reporter assay, active-site mutagenesis of mDot1a methyltransferase domain\",\n      \"journal\": \"American journal of physiology. Cell physiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — active-site mutagenesis abolishing effect plus ChIP, RNAi knockdown, and reporter assays in two orthogonal directions (KD and OE), single lab but multiple rigorous methods\",\n      \"pmids\": [\"16236820\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"AF9 interacts physically with Dot1a both in vitro and in vivo (co-localizing in renal collecting duct cell nuclei) and co-occupies the ENaCalpha promoter; AF9 overexpression causes histone H3 Lys-79 hypermethylation at the ENaCalpha promoter and represses ENaCalpha transcription synergistically with Dot1a, while AF9 knockdown has the opposite effect. Aldosterone negatively regulates AF9 expression, integrating AF9 into the aldosterone-Dot1a-ENaCalpha signaling axis.\",\n      \"method\": \"Co-immunoprecipitation, GST pulldown (in vitro interaction), chromatin immunoprecipitation, RNA interference, luciferase reporter assay, confocal colocalization, RT-PCR/Western blot for AF9 levels after aldosterone\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP plus in vitro pulldown, ChIP, RNAi, and reporter assays across multiple orthogonal methods; replicates and extends prior Dot1a mechanism paper\",\n      \"pmids\": [\"16636056\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"PPARgamma activation in human cortical collecting duct cells upregulates SGK1 (via a PPARgamma response element in the SGK1 promoter), which in turn increases cell-surface ENaCalpha protein levels, thereby enhancing ENaC-mediated sodium reabsorption.\",\n      \"method\": \"PPARgamma agonist/antagonist treatment, SGK1 mRNA measurement, electrophoretic mobility shift assay (EMSA) for PPARgamma binding to SGK1 promoter, cell-surface ENaCalpha quantification\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — EMSA plus agonist/antagonist pharmacological dissection and surface protein quantification, single lab, no mutagenesis of the response element\",\n      \"pmids\": [\"12923071\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"A homozygous missense mutation in SCNN1A (c.727T>C; p.Ser243Pro) in the extracellular loop of alpha-ENaC causes a significant decrease in ENaC channel activity in vitro, exacerbated by high Na+ load, identifying a novel functional domain in the extracellular loop. The reduction is partial, explaining transient/milder phenotype in a mature kidney.\",\n      \"method\": \"Xenopus laevis oocyte expression system with electrophysiology (two-electrode voltage clamp) to measure ENaC activity of mutant vs wild-type channel\",\n      \"journal\": \"American journal of physiology. Endocrinology and metabolism\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — in vitro reconstitution in oocytes with functional readout, single lab, single mutation studied\",\n      \"pmids\": [\"21653223\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"A splice-site mutation in SCNN1A (c.1439+1G>C) leads to retention of intron 9 due to loss of the 5'-donor splice site, as demonstrated in a minigene construct, establishing the molecular consequence of this PHA1-causing mutation on SCNN1A mRNA processing.\",\n      \"method\": \"Minigene splicing assay with Sanger sequencing of resulting transcripts\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — direct minigene functional splice assay, single lab, single mutation\",\n      \"pmids\": [\"23762408\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"The p.Phe226Cys missense mutation in the extracellular domain of SCNN1A causes ~83% reduction in ENaC channel activity when expressed in Xenopus oocytes, due to both reduced intrinsic open probability and decreased channel protein expression at the plasma membrane, explaining a mild/transient PHA1 phenotype.\",\n      \"method\": \"Xenopus laevis oocyte expression with two-electrode voltage clamp electrophysiology; Western blot for protein expression of mutant vs wild-type alpha-ENaC\",\n      \"journal\": \"American journal of physiology. Endocrinology and metabolism\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — in vitro oocyte reconstitution with electrophysiology plus Western blot quantification, single lab, single mutation\",\n      \"pmids\": [\"37134141\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"The NF-κB subunits p65/p50, activated downstream of IKKβ, directly bind the ENaCalpha (SCNN1A) gene promoter and are required for aldosterone-induced ENaCalpha transcription in renal cortical collecting duct cells; IKKβ or p65 knockdown reduces aldosterone-induced ENaCalpha mRNA by >60%.\",\n      \"method\": \"siRNA knockdown of IKKβ and p65, chromatin immunoprecipitation (ChIP), EMSA, luciferase reporter assay in mpkCCDc14 cells\",\n      \"journal\": \"Journal of nephrology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP plus EMSA plus reporter assay plus RNAi, single lab, multiple orthogonal methods\",\n      \"pmids\": [\"26385798\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"MLL2 (KMT2D), a histone H3K4 methyltransferase, associates with the glucocorticoid receptor (GR) in a ligand-independent manner and co-occupies glucocorticoid response elements at the ENaCalpha promoter upon dexamethasone stimulation, promoting chromatin accessibility and GR-mediated ENaCalpha transcription in ARPE-19 cells but not A549 cells.\",\n      \"method\": \"Co-immunoprecipitation of MLL2 with GR, chromatin immunoprecipitation (ChIP) for MLL2 and GR at GREs, FAIRE-qPCR for chromatin accessibility, siRNA knockdown of MLL2\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, ChIP, FAIRE-qPCR, and RNAi in combination, single lab, cell-type-specific context established\",\n      \"pmids\": [\"32139118\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"FOXA1 functions as a pioneer factor enabling cell-type-specific glucocorticoid receptor-driven expression of ENaCalpha (SCNN1A) in ARPE-19 cells; FOXA1 depletion significantly reduces ENaCalpha expression in these cells, whereas ENaCalpha is not a GR target in A549 cells.\",\n      \"method\": \"RNA-seq, RT-qPCR, siRNA knockdown of FOXA1 in two cell lines\",\n      \"journal\": \"International journal of immunopathology and pharmacology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — RNAi with transcriptional readout, no ChIP for FOXA1 at ENaCalpha locus reported in abstract, single lab, single method for the key mechanistic claim\",\n      \"pmids\": [\"32838581\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"1.56 kb of the mouse Scnn1a 5'-upstream promoter sequence is required for cell-specific expression and corticosteroid-mediated regulation in cortical collecting duct cells, as shown by transient transfection of serial deletion mutants; however, this sequence was insufficient to drive reporter gene expression in transgenic mice.\",\n      \"method\": \"Serial deletion mutagenesis of Scnn1a promoter, transient transfection reporter assay in CCD cells, transgenic mouse reporter assay\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — deletion mutagenesis in cell-based reporter assay plus in vivo transgenic validation (negative), single lab, two orthogonal systems\",\n      \"pmids\": [\"11406278\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Conditional endothelial-specific knockout of ENaC-alpha (enENaC-α KO) in mice increases capillary leak and NOX2 (NADPH oxidase 2) expression in lung endothelium during pneumococcal infection/PLY challenge; direct ENaC activation by TIP peptide reduces NOX2 activity and blunts capillary hyperpermeability, placing ENaC-alpha as a negative regulator of endothelial NOX2-mediated oxidative stress in acute lung injury.\",\n      \"method\": \"Conditional endothelial Scnn1a knockout mouse model, Evans blue dye permeability assay, electrical cell substrate impedance sensing in monolayers, Western blot for phospho-p47phox, DuoLink colocalization, chemiluminescence for superoxide, pharmacological TIP peptide activation\",\n      \"journal\": \"American journal of respiratory cell and molecular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — conditional KO with functional permeability readout plus in vitro mechanistic assays, single lab, multiple orthogonal methods\",\n      \"pmids\": [\"39405473\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"NDUFS1 deficiency in alveolar epithelial cells reduces ENaCalpha expression through mitochondrial complex I dysfunction and decreased NAD+ production; supplementing NAD+ (via olaparib) restores ENaCalpha abundance and improves alveolar fluid clearance, placing SCNN1A downstream of the NDUFS1-NAD+ axis in lung epithelium.\",\n      \"method\": \"Proteomic screening, scRNA-seq mining, NDUFS1 KO in alveolar epithelial cells, NAD+ supplementation (olaparib), alveolar fluid clearance measurement, RT-qPCR/Western blot for ENaCalpha\",\n      \"journal\": \"International journal of medical sciences\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — KO with functional readout and pharmacological rescue, single lab, abstract-level detail insufficient to confirm direct mechanistic link vs indirect metabolic effect\",\n      \"pmids\": [\"40860777\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Taurine deficiency in mice (Csad-/- on taurine-free diet) downregulates Scnn1a expression in the uterus during the implantation window, leading to uterine luminal fluid retention and failed embryo implantation; progesterone promotes and estradiol inhibits uterine Csad/taurine synthesis, placing SCNN1A in a steroid hormone-taurine axis controlling uterine fluid resorption.\",\n      \"method\": \"Csad knockout mouse model on taurine-free diet, RT-qPCR and protein expression for Scnn1a and Aqp8, embryo implantation rate measurement, hormonal manipulation\",\n      \"journal\": \"Biology of reproduction\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — KO model with phenotypic and expression readout, correlative link between taurine and Scnn1a expression, single lab, mechanistic pathway not directly validated\",\n      \"pmids\": [\"39428112\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"SCNN1A encodes the alpha subunit of the epithelial sodium channel (ENaC), which is transcriptionally repressed by the Dot1a-AF9 complex via histone H3 Lys-79 hypermethylation at its promoter in an aldosterone-sensitive manner, and activated through NF-κB (p65/p50) binding to its promoter, MLL2-mediated H3K4 methylation enabling glucocorticoid receptor access, and FOXA1 pioneer factor activity; at the protein level, the alpha subunit forms a channel whose activity is reduced by missense mutations in its extracellular loop (e.g., Ser243Pro, Phe226Cys), and in endothelial cells ENaC-alpha restrains NOX2-mediated oxidative stress and capillary leak during acute lung injury.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"SCNN1A encodes the alpha subunit of the epithelial sodium channel (ENaC), which mediates apical sodium reabsorption in aldosterone-responsive epithelia and is controlled by a layered transcriptional program at its promoter [#0, #6]. In the basal state ENaCalpha transcription is repressed by the histone H3 Lys-79 methyltransferase Dot1a, which binds the promoter and hypermethylates H3K79; aldosterone downregulates Dot1a to relieve this repression and trans-activate the gene [#0]. Dot1a acts together with its physical partner AF9, which co-occupies the promoter and represses transcription synergistically, and is itself negatively regulated by aldosterone, integrating AF9 into the aldosterone-Dot1a-ENaCalpha axis [#1]. Aldosterone-induced transcription further requires direct binding of the NF-kappaB subunits p65/p50, activated downstream of IKKbeta, to the promoter [#6], while glucocorticoid-driven expression depends on cell-type-specific access of the glucocorticoid receptor enabled by MLL2 (KMT2D)-mediated H3K4 methylation and chromatin opening [#7]. Steroid signaling also acts through SGK1, induced by PPARgamma, to increase cell-surface ENaCalpha and sodium reabsorption [#2]. At the protein level, missense mutations in the alpha-subunit extracellular loop (Ser243Pro, Phe226Cys) reduce channel activity through lowered open probability and decreased surface expression, and a splice-site mutation causing intron 9 retention disrupts mRNA processing, defining molecular bases of pseudohypoaldosteronism type 1 (PHA1) [#3, #4, #5]. Beyond renal epithelia, endothelial ENaC-alpha restrains NOX2-mediated oxidative stress and capillary leak during acute lung injury [#10].\",\n  \"teleology\": [\n    {\n      \"year\": 2001,\n      \"claim\": \"Before any trans-acting factors were known, it was unclear what genomic region conferred cell-specific and corticosteroid-responsive Scnn1a expression; mapping the promoter localized the regulatory information.\",\n      \"evidence\": \"Serial promoter deletion reporter assays in cortical collecting duct cells plus transgenic mouse reporters\",\n      \"pmids\": [\"11406278\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"1.56 kb sequence insufficient to drive expression in transgenic mice, implying missing distal elements\", \"no specific transcription factors identified\", \"individual response elements not resolved\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"It was unknown how nuclear-receptor signaling beyond aldosterone tuned ENaCalpha surface levels; PPARgamma was shown to act indirectly via SGK1 induction to raise channel abundance at the membrane.\",\n      \"evidence\": \"PPARgamma agonist/antagonist treatment, EMSA for PPARgamma at the SGK1 promoter, and cell-surface ENaCalpha quantification in human CCD cells\",\n      \"pmids\": [\"12923071\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"SGK1 promoter response element not confirmed by mutagenesis\", \"mechanism linking SGK1 to surface ENaCalpha not dissected here\", \"in vivo relevance not tested\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"The chromatin mechanism for basal ENaCalpha repression and its aldosterone-sensitive release was unknown; Dot1a-mediated H3K79 hypermethylation was identified as the repressive mark and aldosterone as a Dot1a downregulator.\",\n      \"evidence\": \"RNAi knockdown, EGFP-tagged overexpression, ChIP, luciferase reporters, and methyltransferase active-site mutagenesis of mDot1a\",\n      \"pmids\": [\"16236820\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"how aldosterone downregulates Dot1a not defined\", \"demethylase removing the mark not identified\", \"single-locus study\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Dot1a was not known to require a recruiting partner; AF9 was shown to physically bind Dot1a, co-occupy the promoter, and synergize in repression under aldosterone control.\",\n      \"evidence\": \"Reciprocal Co-IP, GST pulldown, ChIP, RNAi, luciferase reporters, and confocal colocalization in renal collecting duct cells\",\n      \"pmids\": [\"16636056\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"targeting of AF9 to the promoter sequence not mapped\", \"mechanism of aldosterone-driven AF9 downregulation unresolved\", \"in vivo confirmation absent\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Whether the alpha-subunit extracellular loop was functionally critical was unclear; a homozygous Ser243Pro mutation was shown to partially reduce channel activity, defining a new functional domain.\",\n      \"evidence\": \"Xenopus oocyte two-electrode voltage clamp of mutant vs wild-type channel\",\n      \"pmids\": [\"21653223\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"single mutation in a heterologous system\", \"in vivo channel behavior not assessed\", \"structural basis of the loop's role not resolved\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"The molecular consequence of a PHA1-associated splice-site variant was undefined; a minigene assay showed loss of the 5' donor site causes intron 9 retention.\",\n      \"evidence\": \"Minigene splicing assay with Sanger sequencing of transcripts\",\n      \"pmids\": [\"23762408\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"effect on protein and channel function not tested\", \"single mutation\", \"patient transcript not directly analyzed\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"The signaling required for aldosterone-induced transcription beyond chromatin marks was incomplete; NF-kappaB p65/p50 downstream of IKKbeta was shown to bind the promoter and be required for induction.\",\n      \"evidence\": \"siRNA knockdown of IKKbeta and p65, ChIP, EMSA, and luciferase reporters in mpkCCDc14 cells\",\n      \"pmids\": [\"26385798\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"integration with the Dot1a-AF9 axis not mechanistically linked\", \"specific NF-kappaB site not mutated\", \"single cell model\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"How glucocorticoid receptor accessed the ENaCalpha promoter in a cell-type-specific manner was unknown; MLL2-mediated H3K4 methylation and FOXA1 pioneer activity were identified as enabling factors.\",\n      \"evidence\": \"Co-IP of MLL2 with GR, ChIP, FAIRE-qPCR for accessibility, and siRNA of MLL2 and FOXA1 across ARPE-19 versus A549 cells\",\n      \"pmids\": [\"32139118\", \"32838581\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"FOXA1 occupancy at the ENaCalpha locus not shown by ChIP\", \"basis of cell-type selectivity incompletely defined\", \"interplay with the aldosterone/NF-kappaB axes untested\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"The mechanism by which extracellular-domain mutations reduce channel function was unclear; Phe226Cys was shown to lower both open probability and plasma-membrane expression.\",\n      \"evidence\": \"Xenopus oocyte voltage clamp electrophysiology plus Western blot for surface/total protein\",\n      \"pmids\": [\"37134141\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"single mutation in a heterologous system\", \"trafficking defect mechanism not defined\", \"no patient tissue validation\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"A role for ENaC-alpha outside transporting epithelia was unknown; endothelial-specific deletion revealed it restrains NOX2-driven oxidative stress and capillary leak in acute lung injury.\",\n      \"evidence\": \"Conditional endothelial Scnn1a knockout mice with Evans blue permeability, impedance sensing, phospho-p47phox and superoxide assays, and TIP peptide activation\",\n      \"pmids\": [\"39405473\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"how ENaC-alpha mechanistically suppresses NOX2 not defined\", \"channel-dependence of the effect not established\", \"single disease model\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Upstream metabolic and hormonal inputs to SCNN1A in non-renal tissues were unexplored; the NDUFS1-NAD+ axis in lung epithelium and a progesterone/estradiol-taurine axis in uterus were linked to ENaCalpha abundance.\",\n      \"evidence\": \"NDUFS1 KO with NAD+ rescue and fluid clearance in alveolar cells, and Csad knockout mice with uterine Scnn1a expression and implantation readouts\",\n      \"pmids\": [\"40860777\", \"39428112\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"direct versus indirect effect on SCNN1A not distinguished\", \"no demonstrated transcriptional mechanism linking these axes to the promoter\", \"correlative expression changes only\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How the repressive Dot1a-AF9 chromatin axis, the activating NF-kappaB and GR/MLL2/FOXA1 programs, and SGK1-driven surface trafficking are coordinated into a single integrated control of ENaCalpha expression and activity remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"no unified model linking the repressive and activating transcriptional inputs\", \"stoichiometry and assembly of the alpha subunit into functional channels not addressed in the corpus\", \"in vivo integration of endothelial and epithelial roles untested\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0005215\", \"supporting_discovery_ids\": [3, 5]},\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [3, 5]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [2, 5]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-382551\", \"supporting_discovery_ids\": [3, 5]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [0, 1, 6, 7]},\n      {\"term_id\": \"R-HSA-4839726\", \"supporting_discovery_ids\": [0, 1, 7]}\n    ],\n    \"complexes\": [\"epithelial sodium channel (ENaC)\"],\n    \"partners\": [\"DOT1L\", \"AF9\", \"RELA\", \"NFKB1\", \"NR3C1\", \"KMT2D\", \"FOXA1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}