{"gene":"ATP1B1","run_date":"2026-06-09T22:02:44","timeline":{"discoveries":[{"year":2021,"finding":"ATP1B1 expression is induced by DNA and RNA virus infections. Upon viral infection, ATP1B1 interacts with TRAF3 and TRAF6 and potentiates their ubiquitination, leading to increased phosphorylation of downstream molecules TAK1 and TBK1, thereby enhancing IFN and proinflammatory cytokine production and inhibiting viral replication. Knockdown of ATP1B1 by shRNA had the opposite effects.","method":"shRNA knockdown, overexpression, co-immunoprecipitation, ubiquitination assay, phosphorylation assay, viral replication assay","journal":"Journal of immunology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal co-IP and functional knockdown with defined molecular readouts in a single lab, multiple orthogonal methods","pmids":["34011520"],"is_preprint":false},{"year":2025,"finding":"PRRSV nsp6 competitively interacts with ATP1B1 at residue Leu3 and impairs the ATP1B1-TRAF6 complex. ATP1B1 stabilizes TRAF6 protein levels by downregulating K48-linked ubiquitination of TRAF6, thereby triggering NF-κB signaling and inflammatory response. PRRSV nsp6 disrupts this complex, leading to TRAF6 proteasomal degradation and compromised antiviral innate immunity.","method":"Co-immunoprecipitation, ubiquitination assay, site-directed mutagenesis (nsp6 L3S), recombinant virus rescue, overexpression/knockdown","journal":"Veterinary microbiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP with mutagenesis and functional viral rescue in a single lab, multiple orthogonal methods","pmids":["41252779"],"is_preprint":false},{"year":2018,"finding":"ATP1B1 physically interacts with DCF1 (dendritic cell factor 1), with asparagine residue 266 of ATP1B1 required for this interaction. DCF1 knockout in mice results in upregulation of ATP1B1 in the hippocampus. The DCF1-ATP1B1 interaction in astrocytes impairs their structural plasticity and influences glutamate release via the P38 signaling pathway.","method":"Immunoprecipitation-mass spectrometry, co-immunoprecipitation, cell fluorescence co-localization, site-directed mutagenesis (Asn266), Dcf1 knockout mouse model","journal":"Experimental neurology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP with mutagenesis and genetic knockout model, single lab with multiple orthogonal methods","pmids":["29337145"],"is_preprint":false},{"year":2011,"finding":"HCMV UL136 protein interacts with ATP1B1 (the β1 subunit of Na+/K+-ATPase), identified by yeast two-hybrid screening and confirmed by in vitro pull-down assay and immunofluorescent co-localization at cell membranes.","method":"Yeast two-hybrid screening, pull-down assay, immunofluorescent co-localization","journal":"Brazilian journal of medical and biological research","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, yeast two-hybrid plus pull-down without further functional mechanistic characterization","pmids":["22030864"],"is_preprint":false},{"year":2013,"finding":"A multi-allelic T-rich sequence (TRS) in the 3'UTR of ATP1B1, located downstream of the proximal polyadenylation signal (A2), regulates alternative polyadenylation. In vitro, the T12GT3GT6 allele increases polyadenylation at the A2 site compared to the T23 allele. In human kidneys, the T12GT3GT6 allele is associated with higher relative abundance of A2-polyadenylated ATP1B1 mRNA.","method":"In vitro polyadenylation assay, RT-PCR quantification of polyadenylation isoforms in human kidney samples, genetic association study","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 1–2 / Moderate — in vitro polyadenylation assay combined with human tissue validation, single lab","pmids":["24098465"],"is_preprint":false},{"year":2014,"finding":"The ATP1B1 promoter is epigenetically silenced by hypermethylation in renal cell carcinoma. Knockdown of the VHL tumor suppressor gene in RCC cell lines results in enhanced ATP1B1 promoter hypermethylation accompanied by reduced NaK-β1 protein expression. Treatment with the demethylating agent 5-Aza-2'-deoxycytidine rescued ATP1B1 mRNA and protein expression.","method":"Bisulfite sequencing/methylation analysis of patient tissues and cell lines, VHL shRNA knockdown, 5-Aza-2'-deoxycytidine treatment, RT-PCR, Western blot","journal":"Epigenetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal methods (methylation assay, genetic knockdown, pharmacological rescue) in a single lab","pmids":["24452105"],"is_preprint":false},{"year":2024,"finding":"Co-immunoprecipitation mass spectrometry identified 159 proteins interacting with ATP1B1 in A549 alveolar epithelial cells. Key candidate interactors confirmed by parallel reaction monitoring include HSP90AB1, EIF4A1, TUBB4B, HSPA8, STAT1, and PLEC, suggesting ATP1B1 participates in protein translation, posttranslational processing, and function regulation in alveolar epithelial cells.","method":"Co-immunoprecipitation mass spectrometry, protein-protein interaction network analysis, parallel reaction monitoring (PRM) validation","journal":"Heliyon","confidence":"Low","confidence_rationale":"Tier 3 / Weak — Co-IP/MS interactome with PRM validation but no functional mechanistic follow-up for individual interactions; single lab","pmids":["38912441"],"is_preprint":false},{"year":2025,"finding":"A mRNA-delivered peptide degrader selectively eliminates the oncogenic fusion ATP1B1::PRKACA (found in fibrolamellar carcinoma and related tumors) without affecting native PRKACA, with lethality to FLC tumor cells in a preclinical model. Degradation specificity was attributed to structural properties of the fusion protein.","method":"mRNA-delivered peptide degrader, preclinical tumor cell model, siRNA combination approach","journal":"bioRxiv","confidence":"Low","confidence_rationale":"Tier 2 / Weak — functional preclinical model with mechanistic claim about fusion protein structure, but preprint and single study","pmids":[],"is_preprint":true},{"year":2008,"finding":"Overexpression of ATP1B1 in gastric adenocarcinoma SGC-7901 cells via pEGFP-ATP1B1 transfection increased ATP1B1 mRNA expression and ATPase activity, and inhibited cell proliferation compared to controls.","method":"Eukaryotic expression plasmid transfection, real-time PCR, ATPase activity assay, MTT proliferation assay","journal":"Journal of Sichuan University Medical science edition","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single overexpression experiment with cellular phenotype readout, single lab, no pathway placement","pmids":["18630675"],"is_preprint":false}],"current_model":"ATP1B1 encodes the β1 subunit of the Na+/K+-ATPase and functions as an antiviral innate immune regulator by interacting with and promoting ubiquitination of TRAF3 and TRAF6, thereby activating TAK1/TBK1 signaling and IFN/cytokine production; its expression is regulated by promoter methylation (silenced in renal carcinoma via VHL loss) and by a 3'UTR polymorphism controlling alternative polyadenylation; it also physically interacts with DCF1 to modulate astrocyte structural plasticity via P38 signaling, and is hijacked by viral proteins (HCMV UL136, PRRSV nsp6) that disrupt its TRAF6-stabilizing function to evade innate immunity."},"narrative":{"mechanistic_narrative":"ATP1B1 encodes the β1 subunit of the Na+/K+-ATPase and additionally operates as a positive regulator of antiviral innate immunity. Its expression is induced by both DNA and RNA virus infection, whereupon it binds TRAF3 and TRAF6 and potentiates their ubiquitination to drive TAK1 and TBK1 phosphorylation, amplifying type I interferon and proinflammatory cytokine output and restricting viral replication [PMID:34011520]. A core arm of this activity is stabilization of TRAF6: ATP1B1 suppresses K48-linked ubiquitination of TRAF6 to sustain NF-κB-dependent inflammatory signaling, a function targeted by the viral protein PRRSV nsp6, which competes for ATP1B1 at residue Leu3 to disrupt the ATP1B1–TRAF6 complex and promote TRAF6 proteasomal degradation [PMID:41252779]. Beyond immunity, ATP1B1 physically interacts with DCF1 through asparagine 266, and this interaction in astrocytes modulates structural plasticity and glutamate release via P38 signaling [PMID:29337145]. ATP1B1 expression is controlled at multiple levels, including promoter hypermethylation that silences it in renal cell carcinoma downstream of VHL loss [PMID:24452105] and a 3'UTR T-rich polymorphism that governs alternative polyadenylation in human kidney [PMID:24098465]. Overexpression raises ATPase activity and suppresses proliferation of gastric carcinoma cells [PMID:18630675], but a unified link between its pump function and its signaling roles has not been characterized in the available corpus.","teleology":[{"year":2008,"claim":"Established an early functional readout linking ATP1B1 levels to ATPase activity and growth control, before any signaling role was known.","evidence":"Plasmid overexpression with ATPase activity and MTT proliferation assays in gastric carcinoma cells","pmids":["18630675"],"confidence":"Low","gaps":["Single overexpression experiment with no pathway placement","Does not distinguish pump activity from other effects on proliferation"]},{"year":2011,"claim":"Identified ATP1B1 as a host target of a viral protein, hinting at relevance to infection biology before its immune function was defined.","evidence":"Yeast two-hybrid screen with pull-down and co-localization for HCMV UL136 interaction","pmids":["22030864"],"confidence":"Low","gaps":["No functional consequence of the UL136 interaction tested","Interaction not validated by reciprocal endogenous methods"]},{"year":2013,"claim":"Showed ATP1B1 expression is regulated post-transcriptionally, defining how a 3'UTR polymorphism tunes mRNA isoform abundance.","evidence":"In vitro polyadenylation assays and RT-PCR isoform quantification in human kidney with genetic association","pmids":["24098465"],"confidence":"Medium","gaps":["Functional consequence of altered polyadenylation on protein output not established","No link to any physiological or disease phenotype"]},{"year":2014,"claim":"Defined an epigenetic control mechanism, placing ATP1B1 silencing downstream of VHL loss in renal carcinoma.","evidence":"Methylation analysis, VHL shRNA knockdown, and demethylating-agent rescue in RCC cell lines and tissues","pmids":["24452105"],"confidence":"Medium","gaps":["Mechanism linking VHL loss to promoter methylation not resolved","Functional role of ATP1B1 loss in tumor progression not tested"]},{"year":2018,"claim":"Revealed a neuronal/glial role by mapping the DCF1 interaction interface and connecting it to astrocyte plasticity.","evidence":"IP-MS, co-IP, Asn266 mutagenesis, and Dcf1 knockout mouse with P38 pathway readouts","pmids":["29337145"],"confidence":"Medium","gaps":["How the DCF1 interaction couples to P38 mechanistically not resolved","Relationship to pump function or immune function unclear"]},{"year":2021,"claim":"Defined ATP1B1's core antiviral signaling function, answering whether it actively shapes innate immunity rather than acting only as a pump subunit.","evidence":"shRNA knockdown, overexpression, reciprocal co-IP, ubiquitination and phosphorylation assays, and viral replication assays","pmids":["34011520"],"confidence":"Medium","gaps":["Whether enzymatic pump activity is required for signaling not addressed","Direct vs. scaffold role in TRAF ubiquitination not dissected","Single lab"]},{"year":2024,"claim":"Broadened the interactome to suggest involvement in translation and posttranslational processing in alveolar epithelial cells.","evidence":"Co-IP mass spectrometry with PRM validation of candidate interactors in A549 cells","pmids":["38912441"],"confidence":"Low","gaps":["No functional follow-up for individual interactions","Interactions not validated as direct or functionally consequential"]},{"year":2025,"claim":"Resolved the molecular basis of viral evasion by showing nsp6 competes at Leu3 to destabilize TRAF6, mechanistically extending the antiviral model.","evidence":"Co-IP, ubiquitination assay, nsp6 L3S mutagenesis, and recombinant virus rescue","pmids":["41252779"],"confidence":"Medium","gaps":["Structural detail of the Leu3 interface not resolved","Generalizability beyond PRRSV not tested"]},{"year":2025,"claim":"Explored therapeutic targeting of the oncogenic ATP1B1::PRKACA fusion via selective peptide degradation.","evidence":"mRNA-delivered peptide degrader in a preclinical FLC tumor cell model (preprint)","pmids":[],"confidence":"Low","gaps":["Preprint, single study, not peer-reviewed","Structural basis of degradation specificity asserted but not validated","In vivo efficacy not established"]},{"year":null,"claim":"It remains unknown how ATP1B1's canonical Na+/K+-ATPase pump function relates mechanistically to its TRAF-dependent immune signaling and its DCF1-dependent neuronal roles.","evidence":"No discovery in the corpus integrates the pump and signaling functions","pmids":[],"confidence":"Low","gaps":["No reconstitution distinguishing scaffold from catalytic contributions","No structural model of the TRAF3/TRAF6 interface"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,1]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[0,1,2]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[3]}],"pathway":[{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[0,1]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[0,1,2]}],"complexes":[],"partners":["TRAF3","TRAF6","DCF1","STAT1","HSP90AB1","HSPA8"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P05026","full_name":"Sodium/potassium-transporting ATPase subunit beta-1","aliases":["Sodium/potassium-dependent ATPase subunit beta-1"],"length_aa":303,"mass_kda":35.1,"function":"This is the non-catalytic component of the active enzyme, which catalyzes the hydrolysis of ATP coupled with the exchange of Na(+) and K(+) ions across the plasma membrane. The beta subunit regulates, through assembly of alpha/beta heterodimers, the number of sodium pumps transported to the plasma membrane (PubMed:19694409). Plays a role in innate immunity by enhancing virus-triggered induction of interferons (IFNs) and interferon stimulated genes (ISGs). Mechanistically, enhances the ubiquitination of TRAF3 and TRAF6 as well as the phosphorylation of TAK1 and TBK1 (PubMed:34011520) Involved in cell adhesion and establishing epithelial cell polarity","subcellular_location":"Cell membrane; Apical cell membrane; Cell membrane, sarcolemma","url":"https://www.uniprot.org/uniprotkb/P05026/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/ATP1B1","classification":"Not Classified","n_dependent_lines":8,"n_total_lines":1208,"dependency_fraction":0.006622516556291391},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"ATP1A1","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/ATP1B1","total_profiled":1310},"omim":[{"mim_id":"621029","title":"RING FINGER PROTEIN 183; RNF183","url":"https://www.omim.org/entry/621029"},{"mim_id":"613465","title":"NME/NM23 FAMILY, MEMBER 7; NME7","url":"https://www.omim.org/entry/613465"},{"mim_id":"612873","title":"Na+/K+ TRANSPORTING ATPase-INTERACTING 4; NKAIN4","url":"https://www.omim.org/entry/612873"},{"mim_id":"612871","title":"Na+/K+ TRANSPORTING ATPase-INTERACTING 1; NKAIN1","url":"https://www.omim.org/entry/612871"},{"mim_id":"611642","title":"HEPATOCYTE CELL ADHESION MOLECULE; HEPACAM","url":"https://www.omim.org/entry/611642"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"kidney","ntpm":1376.9}],"url":"https://www.proteinatlas.org/search/ATP1B1"},"hgnc":{"alias_symbol":[],"prev_symbol":["ATP1B"]},"alphafold":{"accession":"P05026","domains":[{"cath_id":"2.60.40.1660","chopping":"76-300","consensus_level":"high","plddt":90.6621,"start":76,"end":300},{"cath_id":"1.20.5","chopping":"16-59","consensus_level":"high","plddt":91.5945,"start":16,"end":59}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P05026","model_url":"https://alphafold.ebi.ac.uk/files/AF-P05026-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P05026-F1-predicted_aligned_error_v6.png","plddt_mean":89.56},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=ATP1B1","jax_strain_url":"https://www.jax.org/strain/search?query=ATP1B1"},"sequence":{"accession":"P05026","fasta_url":"https://rest.uniprot.org/uniprotkb/P05026.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P05026/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P05026"}},"corpus_meta":[{"pmid":"21881522","id":"PMC_21881522","title":"Association of ATP1B1, RGS5 and SELE polymorphisms with hypertension and blood pressure in African-Americans.","date":"2011","source":"Journal of hypertension","url":"https://pubmed.ncbi.nlm.nih.gov/21881522","citation_count":29,"is_preprint":false},{"pmid":"24452105","id":"PMC_24452105","title":"Epigenetic silencing of Na,K-ATPase β 1 subunit gene ATP1B1 by methylation in clear cell renal cell carcinoma.","date":"2014","source":"Epigenetics","url":"https://pubmed.ncbi.nlm.nih.gov/24452105","citation_count":29,"is_preprint":false},{"pmid":"11275679","id":"PMC_11275679","title":"Differential transcription of ion transporters, NHE1, ATP1B1, NKCC1 in human peripheral blood lymphocytes activated to proliferation.","date":"2001","source":"Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and 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sciences","url":"https://pubmed.ncbi.nlm.nih.gov/36012584","citation_count":13,"is_preprint":false},{"pmid":"24098465","id":"PMC_24098465","title":"A polymorphic 3'UTR element in ATP1B1 regulates alternative polyadenylation and is associated with blood pressure.","date":"2013","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/24098465","citation_count":13,"is_preprint":false},{"pmid":"35901982","id":"PMC_35901982","title":"Rapamycin inhibits the progression of human acute myeloid leukemia by regulating the circ_0094100/miR-217/ATP1B1 axis.","date":"2022","source":"Experimental hematology","url":"https://pubmed.ncbi.nlm.nih.gov/35901982","citation_count":12,"is_preprint":false},{"pmid":"34011520","id":"PMC_34011520","title":"Inducible ATP1B1 Upregulates Antiviral Innate Immune Responses by the Ubiquitination of TRAF3 and TRAF6.","date":"2021","source":"Journal of immunology (Baltimore, Md. : 1950)","url":"https://pubmed.ncbi.nlm.nih.gov/34011520","citation_count":10,"is_preprint":false},{"pmid":"22030864","id":"PMC_22030864","title":"Interaction between human cytomegalovirus UL136 protein and ATP1B1 protein.","date":"2011","source":"Brazilian journal of medical and biological research = Revista brasileira de pesquisas medicas e biologicas","url":"https://pubmed.ncbi.nlm.nih.gov/22030864","citation_count":9,"is_preprint":false},{"pmid":"1690387","id":"PMC_1690387","title":"MspI and PvuII polymorphisms in the Na,K-ATPase beta subunit gene ATP1B1.","date":"1990","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/1690387","citation_count":9,"is_preprint":false},{"pmid":"36736852","id":"PMC_36736852","title":"Intergenic variants, rs1200114 and rs1200108 are genetically associated along with a decreased ATP1B1 expression in Fuchs Endothelial Corneal Dystrophy.","date":"2023","source":"Experimental eye 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assay.","date":"2024","source":"Heliyon","url":"https://pubmed.ncbi.nlm.nih.gov/38912441","citation_count":2,"is_preprint":false},{"pmid":"41252779","id":"PMC_41252779","title":"Porcine reproductive and respiratory syndrome virus nsp6 hijacks ATP1B1 antagonizing TRAF6 mediated antiviral innate immunity.","date":"2025","source":"Veterinary microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/41252779","citation_count":1,"is_preprint":false},{"pmid":"29337145","id":"PMC_29337145","title":"Interaction of DCF1 with ATP1B1 induces impairment in astrocyte structural plasticity via the P38 signaling pathway.","date":"2018","source":"Experimental neurology","url":"https://pubmed.ncbi.nlm.nih.gov/29337145","citation_count":1,"is_preprint":false},{"pmid":"11534181","id":"PMC_11534181","title":"[Study of \"group\" expression of mRNA of the ion transporters ATP1B1, NHE1 and NKCC1, beta-actin, glycerophosphosphate dehydrogenase, proteins regulating proliferation and apoptosis of p53, Bcl-2, IL-2 and hSGK kinase at the prereplicative stage of human lymphocyte activation].","date":"2001","source":"Tsitologiia","url":"https://pubmed.ncbi.nlm.nih.gov/11534181","citation_count":1,"is_preprint":false},{"pmid":"40558505","id":"PMC_40558505","title":"Atp1b2 Knock-In Mice Exhibit a Cone-Rod Dystrophy-Like Phenotype.","date":"2025","source":"Cells","url":"https://pubmed.ncbi.nlm.nih.gov/40558505","citation_count":0,"is_preprint":false},{"pmid":"18630675","id":"PMC_18630675","title":"[Construction of eukaryotic expression plasmid pEGFP-ATP1B1 and its effect on gastric adenocarcinoma cell SGC-7901].","date":"2008","source":"Sichuan da xue xue bao. Yi xue ban = Journal of Sichuan University. Medical science edition","url":"https://pubmed.ncbi.nlm.nih.gov/18630675","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.04.24.650501","title":"Specific degrader for fusion oncokinase kills tumors and is augmented by bimodal degrader-siRNA","date":"2025-04-25","source":"bioRxiv","url":"https://doi.org/10.1101/2025.04.24.650501","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.09.03.673966","title":"High affinity cross-context cellular assays reveal novel protein-protein interactions of peripheral myelin protein of 22 kDa","date":"2025-09-04","source":"bioRxiv","url":"https://doi.org/10.1101/2025.09.03.673966","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":12221,"output_tokens":2403,"usd":0.036354,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9511,"output_tokens":2831,"usd":0.059165,"stage2_stop_reason":"end_turn"},"total_usd":0.095519,"stage1_batch_id":"msgbatch_0179Z2pxRYBsXi2xADneafRw","stage2_batch_id":"msgbatch_01LaY9ZaVdgjWArcZo9eqQE5","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2021,\n      \"finding\": \"ATP1B1 expression is induced by DNA and RNA virus infections. Upon viral infection, ATP1B1 interacts with TRAF3 and TRAF6 and potentiates their ubiquitination, leading to increased phosphorylation of downstream molecules TAK1 and TBK1, thereby enhancing IFN and proinflammatory cytokine production and inhibiting viral replication. Knockdown of ATP1B1 by shRNA had the opposite effects.\",\n      \"method\": \"shRNA knockdown, overexpression, co-immunoprecipitation, ubiquitination assay, phosphorylation assay, viral replication assay\",\n      \"journal\": \"Journal of immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal co-IP and functional knockdown with defined molecular readouts in a single lab, multiple orthogonal methods\",\n      \"pmids\": [\"34011520\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"PRRSV nsp6 competitively interacts with ATP1B1 at residue Leu3 and impairs the ATP1B1-TRAF6 complex. ATP1B1 stabilizes TRAF6 protein levels by downregulating K48-linked ubiquitination of TRAF6, thereby triggering NF-κB signaling and inflammatory response. PRRSV nsp6 disrupts this complex, leading to TRAF6 proteasomal degradation and compromised antiviral innate immunity.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, site-directed mutagenesis (nsp6 L3S), recombinant virus rescue, overexpression/knockdown\",\n      \"journal\": \"Veterinary microbiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP with mutagenesis and functional viral rescue in a single lab, multiple orthogonal methods\",\n      \"pmids\": [\"41252779\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"ATP1B1 physically interacts with DCF1 (dendritic cell factor 1), with asparagine residue 266 of ATP1B1 required for this interaction. DCF1 knockout in mice results in upregulation of ATP1B1 in the hippocampus. The DCF1-ATP1B1 interaction in astrocytes impairs their structural plasticity and influences glutamate release via the P38 signaling pathway.\",\n      \"method\": \"Immunoprecipitation-mass spectrometry, co-immunoprecipitation, cell fluorescence co-localization, site-directed mutagenesis (Asn266), Dcf1 knockout mouse model\",\n      \"journal\": \"Experimental neurology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP with mutagenesis and genetic knockout model, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"29337145\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"HCMV UL136 protein interacts with ATP1B1 (the β1 subunit of Na+/K+-ATPase), identified by yeast two-hybrid screening and confirmed by in vitro pull-down assay and immunofluorescent co-localization at cell membranes.\",\n      \"method\": \"Yeast two-hybrid screening, pull-down assay, immunofluorescent co-localization\",\n      \"journal\": \"Brazilian journal of medical and biological research\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, yeast two-hybrid plus pull-down without further functional mechanistic characterization\",\n      \"pmids\": [\"22030864\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"A multi-allelic T-rich sequence (TRS) in the 3'UTR of ATP1B1, located downstream of the proximal polyadenylation signal (A2), regulates alternative polyadenylation. In vitro, the T12GT3GT6 allele increases polyadenylation at the A2 site compared to the T23 allele. In human kidneys, the T12GT3GT6 allele is associated with higher relative abundance of A2-polyadenylated ATP1B1 mRNA.\",\n      \"method\": \"In vitro polyadenylation assay, RT-PCR quantification of polyadenylation isoforms in human kidney samples, genetic association study\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — in vitro polyadenylation assay combined with human tissue validation, single lab\",\n      \"pmids\": [\"24098465\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"The ATP1B1 promoter is epigenetically silenced by hypermethylation in renal cell carcinoma. Knockdown of the VHL tumor suppressor gene in RCC cell lines results in enhanced ATP1B1 promoter hypermethylation accompanied by reduced NaK-β1 protein expression. Treatment with the demethylating agent 5-Aza-2'-deoxycytidine rescued ATP1B1 mRNA and protein expression.\",\n      \"method\": \"Bisulfite sequencing/methylation analysis of patient tissues and cell lines, VHL shRNA knockdown, 5-Aza-2'-deoxycytidine treatment, RT-PCR, Western blot\",\n      \"journal\": \"Epigenetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal methods (methylation assay, genetic knockdown, pharmacological rescue) in a single lab\",\n      \"pmids\": [\"24452105\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Co-immunoprecipitation mass spectrometry identified 159 proteins interacting with ATP1B1 in A549 alveolar epithelial cells. Key candidate interactors confirmed by parallel reaction monitoring include HSP90AB1, EIF4A1, TUBB4B, HSPA8, STAT1, and PLEC, suggesting ATP1B1 participates in protein translation, posttranslational processing, and function regulation in alveolar epithelial cells.\",\n      \"method\": \"Co-immunoprecipitation mass spectrometry, protein-protein interaction network analysis, parallel reaction monitoring (PRM) validation\",\n      \"journal\": \"Heliyon\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — Co-IP/MS interactome with PRM validation but no functional mechanistic follow-up for individual interactions; single lab\",\n      \"pmids\": [\"38912441\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"A mRNA-delivered peptide degrader selectively eliminates the oncogenic fusion ATP1B1::PRKACA (found in fibrolamellar carcinoma and related tumors) without affecting native PRKACA, with lethality to FLC tumor cells in a preclinical model. Degradation specificity was attributed to structural properties of the fusion protein.\",\n      \"method\": \"mRNA-delivered peptide degrader, preclinical tumor cell model, siRNA combination approach\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 2 / Weak — functional preclinical model with mechanistic claim about fusion protein structure, but preprint and single study\",\n      \"pmids\": [],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Overexpression of ATP1B1 in gastric adenocarcinoma SGC-7901 cells via pEGFP-ATP1B1 transfection increased ATP1B1 mRNA expression and ATPase activity, and inhibited cell proliferation compared to controls.\",\n      \"method\": \"Eukaryotic expression plasmid transfection, real-time PCR, ATPase activity assay, MTT proliferation assay\",\n      \"journal\": \"Journal of Sichuan University Medical science edition\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single overexpression experiment with cellular phenotype readout, single lab, no pathway placement\",\n      \"pmids\": [\"18630675\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ATP1B1 encodes the β1 subunit of the Na+/K+-ATPase and functions as an antiviral innate immune regulator by interacting with and promoting ubiquitination of TRAF3 and TRAF6, thereby activating TAK1/TBK1 signaling and IFN/cytokine production; its expression is regulated by promoter methylation (silenced in renal carcinoma via VHL loss) and by a 3'UTR polymorphism controlling alternative polyadenylation; it also physically interacts with DCF1 to modulate astrocyte structural plasticity via P38 signaling, and is hijacked by viral proteins (HCMV UL136, PRRSV nsp6) that disrupt its TRAF6-stabilizing function to evade innate immunity.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"ATP1B1 encodes the β1 subunit of the Na+/K+-ATPase and additionally operates as a positive regulator of antiviral innate immunity. Its expression is induced by both DNA and RNA virus infection, whereupon it binds TRAF3 and TRAF6 and potentiates their ubiquitination to drive TAK1 and TBK1 phosphorylation, amplifying type I interferon and proinflammatory cytokine output and restricting viral replication [#0]. A core arm of this activity is stabilization of TRAF6: ATP1B1 suppresses K48-linked ubiquitination of TRAF6 to sustain NF-κB-dependent inflammatory signaling, a function targeted by the viral protein PRRSV nsp6, which competes for ATP1B1 at residue Leu3 to disrupt the ATP1B1–TRAF6 complex and promote TRAF6 proteasomal degradation [#1]. Beyond immunity, ATP1B1 physically interacts with DCF1 through asparagine 266, and this interaction in astrocytes modulates structural plasticity and glutamate release via P38 signaling [#2]. ATP1B1 expression is controlled at multiple levels, including promoter hypermethylation that silences it in renal cell carcinoma downstream of VHL loss [#5] and a 3'UTR T-rich polymorphism that governs alternative polyadenylation in human kidney [#4]. Overexpression raises ATPase activity and suppresses proliferation of gastric carcinoma cells [#8], but a unified link between its pump function and its signaling roles has not been characterized in the available corpus.\",\n  \"teleology\": [\n    {\n      \"year\": 2008,\n      \"claim\": \"Established an early functional readout linking ATP1B1 levels to ATPase activity and growth control, before any signaling role was known.\",\n      \"evidence\": \"Plasmid overexpression with ATPase activity and MTT proliferation assays in gastric carcinoma cells\",\n      \"pmids\": [\"18630675\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single overexpression experiment with no pathway placement\", \"Does not distinguish pump activity from other effects on proliferation\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Identified ATP1B1 as a host target of a viral protein, hinting at relevance to infection biology before its immune function was defined.\",\n      \"evidence\": \"Yeast two-hybrid screen with pull-down and co-localization for HCMV UL136 interaction\",\n      \"pmids\": [\"22030864\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No functional consequence of the UL136 interaction tested\", \"Interaction not validated by reciprocal endogenous methods\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Showed ATP1B1 expression is regulated post-transcriptionally, defining how a 3'UTR polymorphism tunes mRNA isoform abundance.\",\n      \"evidence\": \"In vitro polyadenylation assays and RT-PCR isoform quantification in human kidney with genetic association\",\n      \"pmids\": [\"24098465\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional consequence of altered polyadenylation on protein output not established\", \"No link to any physiological or disease phenotype\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Defined an epigenetic control mechanism, placing ATP1B1 silencing downstream of VHL loss in renal carcinoma.\",\n      \"evidence\": \"Methylation analysis, VHL shRNA knockdown, and demethylating-agent rescue in RCC cell lines and tissues\",\n      \"pmids\": [\"24452105\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism linking VHL loss to promoter methylation not resolved\", \"Functional role of ATP1B1 loss in tumor progression not tested\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Revealed a neuronal/glial role by mapping the DCF1 interaction interface and connecting it to astrocyte plasticity.\",\n      \"evidence\": \"IP-MS, co-IP, Asn266 mutagenesis, and Dcf1 knockout mouse with P38 pathway readouts\",\n      \"pmids\": [\"29337145\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"How the DCF1 interaction couples to P38 mechanistically not resolved\", \"Relationship to pump function or immune function unclear\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Defined ATP1B1's core antiviral signaling function, answering whether it actively shapes innate immunity rather than acting only as a pump subunit.\",\n      \"evidence\": \"shRNA knockdown, overexpression, reciprocal co-IP, ubiquitination and phosphorylation assays, and viral replication assays\",\n      \"pmids\": [\"34011520\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether enzymatic pump activity is required for signaling not addressed\", \"Direct vs. scaffold role in TRAF ubiquitination not dissected\", \"Single lab\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Broadened the interactome to suggest involvement in translation and posttranslational processing in alveolar epithelial cells.\",\n      \"evidence\": \"Co-IP mass spectrometry with PRM validation of candidate interactors in A549 cells\",\n      \"pmids\": [\"38912441\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No functional follow-up for individual interactions\", \"Interactions not validated as direct or functionally consequential\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Resolved the molecular basis of viral evasion by showing nsp6 competes at Leu3 to destabilize TRAF6, mechanistically extending the antiviral model.\",\n      \"evidence\": \"Co-IP, ubiquitination assay, nsp6 L3S mutagenesis, and recombinant virus rescue\",\n      \"pmids\": [\"41252779\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural detail of the Leu3 interface not resolved\", \"Generalizability beyond PRRSV not tested\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Explored therapeutic targeting of the oncogenic ATP1B1::PRKACA fusion via selective peptide degradation.\",\n      \"evidence\": \"mRNA-delivered peptide degrader in a preclinical FLC tumor cell model (preprint)\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Preprint, single study, not peer-reviewed\", \"Structural basis of degradation specificity asserted but not validated\", \"In vivo efficacy not established\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unknown how ATP1B1's canonical Na+/K+-ATPase pump function relates mechanistically to its TRAF-dependent immune signaling and its DCF1-dependent neuronal roles.\",\n      \"evidence\": \"No discovery in the corpus integrates the pump and signaling functions\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No reconstitution distinguishing scaffold from catalytic contributions\", \"No structural model of the TRAF3/TRAF6 interface\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [0, 1, 2]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [3]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [0, 1, 2]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"TRAF3\", \"TRAF6\", \"DCF1\", \"STAT1\", \"HSP90AB1\", \"HSPA8\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":4,"faith_total":4,"faith_pct":100.0}}