{"gene":"EOLA1","run_date":"2026-06-09T23:54:43","timeline":{"discoveries":[{"year":2004,"finding":"EOLA1 physically interacts with metallothionein 2A (MT2A), identified by yeast two-hybrid screening of a human liver cDNA library using EOLA1 cDNA as bait, and confirmed by co-immunoprecipitation; stable transfection of EOLA1 stimulates ECV304 cell proliferation.","method":"Yeast two-hybrid screening + co-immunoprecipitation; stable transfection + cell proliferation assay","journal":"Biochemical and biophysical research communications / Chinese journal of medical genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal co-IP confirmed the Y2H interaction; proliferation phenotype shown by stable transfection; two papers from same lab using orthogonal methods","pmids":["15541360","16215939"],"is_preprint":false},{"year":2005,"finding":"Inducible overexpression of EOLA1 in ECV304 cells significantly enhances cell proliferation, establishing a functional role for EOLA1 protein in promoting endothelial cell proliferation.","method":"Inducible overexpression vector (pOPRSVI-EOLA1) transfection; cell counting growth curve","journal":"Chinese journal of burns","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, single overexpression method, no mechanistic pathway placement","pmids":["16185414"],"is_preprint":false},{"year":2007,"finding":"Knockdown of EOLA1 expression by siRNA in ECV304 cells slows cell proliferation, confirming EOLA1 is required for normal endothelial cell proliferation.","method":"siRNA knockdown + cell proliferation counting","journal":"Chinese journal of medical genetics","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, single RNAi method, phenotype only","pmids":["17557240"],"is_preprint":false},{"year":2010,"finding":"EOLA1 protein localizes to both the nucleus and cytoplasmic matrix of ECV304 endothelial cells, consistent with a role as a signal transduction factor.","method":"EGFP-EOLA1 fusion protein expression, laser scanning confocal microscopy, and immunoelectron microscopy","journal":"Chinese journal of burns","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — two orthogonal direct localization methods (confocal + immunoelectron microscopy), single lab","pmids":["21223654"],"is_preprint":false},{"year":2014,"finding":"EOLA1 acts as a negative regulator of LPS-induced IL-6 production and apoptosis in HUVECs by regulating MT2A; LPS induces EOLA1 expression in a time-dependent manner, deletion of EOLA1 promotes LPS-induced IL-6 production and apoptosis, and MT2A is activated by LPS and plays a key role in LPS-induced IL-6 expression downstream of EOLA1.","method":"siRNA knockdown of EOLA1 and MT2A, LPS stimulation, IL-6 measurement, apoptosis assay in HUVECs","journal":"Molecular and cellular biochemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function (siRNA) with defined cytokine and apoptosis readouts; pathway placement of EOLA1 upstream of MT2A; single lab","pmids":["24916366"],"is_preprint":false},{"year":2015,"finding":"EOLA1 negatively regulates LPS-induced VCAM-1 expression in ECV304 cells via its association with MT2A; knockdown of EOLA1 enhances LPS-induced VCAM-1 production whereas overexpression reduces it; MT2A knockdown also reduces LPS-induced VCAM-1 production. EOLA1 subcellular localization is broadly diffuse in the cells.","method":"siRNA knockdown and overexpression of EOLA1; siRNA knockdown of MT2A; VCAM-1 quantification after LPS treatment; subcellular localization assessment","journal":"International journal of inflammation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal gain- and loss-of-function with defined molecular readout; pathway placement through MT2A; single lab","pmids":["26881174"],"is_preprint":false},{"year":2019,"finding":"Crystal structure of human EOLA1 was determined at 1.71 Å resolution, revealing a characteristic β-barrel fold surrounded by α-helices structurally similar to an ASC-1 homology (ASCH) domain. EOLA1 retains a conserved 'GxKxxExR' motif in its cavity, and the cavity contains aromatic and polar residues implying nucleotide binding capacity. A positively charged cleft is present, similar to YTH domain proteins and a ribonuclease-active ASCH protein, suggesting potential RNA binding activity.","method":"X-ray crystallography (1.71 Å resolution crystal structure)","journal":"Molecules","confidence":"High","confidence_rationale":"Tier 1 / Moderate — high-resolution crystal structure with detailed structural analysis; single lab but rigorous structural method","pmids":["31569543"],"is_preprint":false},{"year":2025,"finding":"Crystal structures of mouse EOLA1 were determined, revealing differences in substrate preferences compared to homologous ASCH domain proteins (EcYqfB and human TRIP4-ASCH domain). The E. coli ASCH domain protein YqfB has amidohydrolase activity converting N4-acetylcytidine (ac4C) to cytidine, placing EOLA1 in the ASCH structural family but showing distinct substrate selectivity.","method":"X-ray crystallography of mouse EOLA1 and comparative structural analysis with EcYqfB and TRIP4-ASCH","journal":"Structure","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — crystal structure of mouse EOLA1 determined, but functional characterization of EOLA1's own catalytic activity was not directly shown; primary data on activity was for EcYqfB","pmids":["40939588"],"is_preprint":false}],"current_model":"EOLA1 is a nuclear and cytoplasmic endothelial protein with an ASCH-domain β-barrel fold that physically interacts with metallothionein 2A (MT2A) and acts as a negative regulator of LPS-induced inflammatory responses (IL-6 production, VCAM-1 expression) and apoptosis in endothelial cells; its crystal structure reveals a conserved nucleotide-binding cavity and positively charged RNA-binding cleft, suggesting it may control gene expression through RNA binding, while its interaction with MT2A mediates its anti-inflammatory signaling and its expression promotes endothelial cell proliferation."},"narrative":{"mechanistic_narrative":"EOLA1 is an endothelial protein that functions as a negative regulator of LPS-induced inflammatory signaling and apoptosis while promoting endothelial cell proliferation [PMID:24916366, PMID:15541360, PMID:16215939]. It physically interacts with metallothionein 2A (MT2A), and this association is central to its anti-inflammatory output: loss of EOLA1 enhances LPS-induced IL-6 production and apoptosis in HUVECs, with MT2A acting downstream as a key effector of IL-6 expression [PMID:15541360, PMID:16215939, PMID:24916366]. EOLA1 likewise restrains LPS-induced VCAM-1 expression through its MT2A partnership, with knockdown raising and overexpression lowering VCAM-1 [PMID:26881174]. EOLA1 localizes to both nucleus and cytoplasm, consistent with a signal-transduction role [PMID:21223654]. Structurally, EOLA1 adopts an ASC-1 homology (ASCH) β-barrel fold with a conserved 'GxKxxExR' cavity implying nucleotide binding and a positively charged cleft resembling that of RNA-binding ASCH and YTH domain proteins [PMID:31569543]; comparative crystallography of the mouse ortholog places it in the ASCH family but with substrate selectivity distinct from the ac4C amidohydrolase EcYqfB and human TRIP4-ASCH [PMID:40939588]. A catalytic or RNA-binding activity intrinsic to EOLA1 has not been directly demonstrated in the available corpus.","teleology":[{"year":2004,"claim":"Established the first molecular partner of EOLA1 and its first cellular phenotype, framing it as an endothelial protein that interacts with MT2A and drives proliferation.","evidence":"Yeast two-hybrid screen of a human liver cDNA library with reciprocal co-IP confirmation; stable transfection proliferation assay in ECV304 cells","pmids":["15541360","16215939"],"confidence":"Medium","gaps":["Interaction mapped only by Y2H/co-IP, no interface or domain mapping","Functional consequence of the MT2A interaction not yet defined","Proliferation effect tied to a single cell line"]},{"year":2005,"claim":"Tested whether elevated EOLA1 protein is sufficient to drive proliferation, reinforcing a pro-proliferative role.","evidence":"Inducible overexpression (pOPRSVI-EOLA1) and growth-curve counting in ECV304 cells","pmids":["16185414"],"confidence":"Low","gaps":["Single overexpression method without mechanistic pathway placement","No molecular readout linking EOLA1 to a proliferation pathway"]},{"year":2007,"claim":"Addressed whether EOLA1 is required for proliferation, complementing the gain-of-function data with loss-of-function evidence.","evidence":"siRNA knockdown plus proliferation counting in ECV304 cells","pmids":["17557240"],"confidence":"Low","gaps":["Phenotype only, no mechanism","Single RNAi method, no rescue control"]},{"year":2010,"claim":"Determined where EOLA1 acts in the cell, supporting a dual nuclear/cytoplasmic signal-transduction role.","evidence":"EGFP-EOLA1 fusion imaged by laser scanning confocal and immunoelectron microscopy in ECV304 cells","pmids":["21223654"],"confidence":"Medium","gaps":["Localization shown via overexpressed fusion, not endogenous protein","Functional significance of nuclear vs cytoplasmic pools not dissected"]},{"year":2014,"claim":"Placed EOLA1 in an inflammatory signaling pathway, showing it acts upstream of MT2A to restrain LPS-induced IL-6 and apoptosis.","evidence":"siRNA knockdown of EOLA1 and MT2A, LPS stimulation, IL-6 and apoptosis readouts in HUVECs","pmids":["24916366"],"confidence":"Medium","gaps":["Molecular mechanism by which EOLA1 regulates MT2A unresolved","Single lab and cell system","How LPS induces EOLA1 expression not defined"]},{"year":2015,"claim":"Extended EOLA1's anti-inflammatory role to adhesion molecule control, showing MT2A-dependent suppression of LPS-induced VCAM-1.","evidence":"Reciprocal siRNA knockdown and overexpression of EOLA1, MT2A knockdown, VCAM-1 quantification after LPS in ECV304 cells","pmids":["26881174"],"confidence":"Medium","gaps":["Direct biochemical link between EOLA1-MT2A binding and VCAM-1 transcription not established","Diffuse localization not correlated with signaling state"]},{"year":2019,"claim":"Provided the first structural framework, assigning EOLA1 to the ASCH β-barrel fold with cavity features implying nucleotide and RNA binding.","evidence":"1.71 Å X-ray crystal structure of human EOLA1 with structural comparison to YTH and ASCH proteins","pmids":["31569543"],"confidence":"High","gaps":["No experimental demonstration of nucleotide or RNA binding","No ligand-bound structure","Structural inference not linked to the MT2A/inflammatory function"]},{"year":2025,"claim":"Compared EOLA1 against catalytically active ASCH proteins, refining family placement and indicating distinct substrate selectivity.","evidence":"X-ray crystallography of mouse EOLA1 with comparative analysis against EcYqfB and TRIP4-ASCH","pmids":["40939588"],"confidence":"Medium","gaps":["EOLA1's own catalytic activity not directly measured","Activity data shown only for EcYqfB, not EOLA1","No identified physiological substrate"]},{"year":null,"claim":"Whether EOLA1 possesses intrinsic catalytic or RNA-binding activity and how this connects to its MT2A-dependent anti-inflammatory function remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No demonstrated enzymatic or RNA-binding activity for EOLA1 itself","Mechanistic bridge between the structural fold and the inflammatory phenotype absent","No in vivo or disease-level validation"]}],"mechanism_profile":{"molecular_activity":[],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[3]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[3]}],"pathway":[{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[4,5]}],"complexes":[],"partners":["MT2A"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q8TE69","full_name":"Protein EOLA1","aliases":["Endothelial-overexpressed lipopolysaccharide-associated factor 1","Endothelium and lymphocyte associated ASCH domain 1"],"length_aa":158,"mass_kda":17.9,"function":"May play a role in cell protection during the inflammatory response. In epithelial cells, negatively regulates IL6 production and apoptosis through the regulation of MT2A expression (PubMed:24916366)","subcellular_location":"","url":"https://www.uniprot.org/uniprotkb/Q8TE69/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/EOLA1","classification":"Not Classified","n_dependent_lines":1,"n_total_lines":1047,"dependency_fraction":0.0009551098376313276},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/EOLA1","total_profiled":1310},"omim":[{"mim_id":"300954","title":"ENDOTHELIUM- AND LYMPHOCYTE-ASSOCIATED ASCH DOMAIN 1; EOLA1","url":"https://www.omim.org/entry/300954"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/EOLA1"},"hgnc":{"alias_symbol":[],"prev_symbol":["CXorf40","CXorf40A"]},"alphafold":{"accession":"Q8TE69","domains":[{"cath_id":"2.30.130.30","chopping":"3-152","consensus_level":"high","plddt":97.835,"start":3,"end":152}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8TE69","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q8TE69-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q8TE69-F1-predicted_aligned_error_v6.png","plddt_mean":96.38},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=EOLA1","jax_strain_url":"https://www.jax.org/strain/search?query=EOLA1"},"sequence":{"accession":"Q8TE69","fasta_url":"https://rest.uniprot.org/uniprotkb/Q8TE69.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q8TE69/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8TE69"}},"corpus_meta":[{"pmid":"24916366","id":"PMC_24916366","title":"EOLA1 protects lipopolysaccharide induced IL-6 production and apoptosis by regulation of MT2A in human umbilical vein endothelial cells.","date":"2014","source":"Molecular and cellular biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/24916366","citation_count":20,"is_preprint":false},{"pmid":"15541360","id":"PMC_15541360","title":"Identification and characterization of a novel gene EOLA1 stimulating ECV304 cell proliferation.","date":"2004","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/15541360","citation_count":13,"is_preprint":false},{"pmid":"26881174","id":"PMC_26881174","title":"EOLA1 Inhibits Lipopolysaccharide-Induced Vascular Cell Adhesion Molecule-1 Expression by Association with MT2A in ECV304 Cells.","date":"2015","source":"International journal of inflammation","url":"https://pubmed.ncbi.nlm.nih.gov/26881174","citation_count":6,"is_preprint":false},{"pmid":"31007603","id":"PMC_31007603","title":"The Reduced Expression of EOLA1 May Be Related to Refractory Diabetic Foot Ulcer.","date":"2019","source":"Mediators of inflammation","url":"https://pubmed.ncbi.nlm.nih.gov/31007603","citation_count":5,"is_preprint":false},{"pmid":"17557240","id":"PMC_17557240","title":"[The effect of inhibiting EOLA1 expression in ECV304 cells].","date":"2007","source":"Zhonghua yi xue yi chuan xue za zhi = Zhonghua yixue yichuanxue zazhi = Chinese journal of medical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/17557240","citation_count":3,"is_preprint":false},{"pmid":"36257379","id":"PMC_36257379","title":"Diagnosis of patients with mucopolysaccharidosis type II via RNA sequencing.","date":"2022","source":"Clinica chimica acta; international journal of clinical chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/36257379","citation_count":2,"is_preprint":false},{"pmid":"18534126","id":"PMC_18534126","title":"Changes in the expression of endothelial-overexpressed lipopolysaccharide-associated factor 1 in grafts during acute rejection following liver transplantation in rats.","date":"2008","source":"The Journal of international medical research","url":"https://pubmed.ncbi.nlm.nih.gov/18534126","citation_count":2,"is_preprint":false},{"pmid":"16215939","id":"PMC_16215939","title":"[Identification and characterization of a novel gene EOLA1 stimulating ECV304 cell proliferation].","date":"2005","source":"Zhonghua yi xue yi chuan xue za zhi = Zhonghua yixue yichuanxue zazhi = Chinese journal of medical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/16215939","citation_count":1,"is_preprint":false},{"pmid":"21223656","id":"PMC_21223656","title":"[Preparation of polyclonal antibody of human endothelial-overexpressed lipopolysaccharide-associated factor 1].","date":"2010","source":"Zhonghua shao shang za zhi = Zhonghua shaoshang zazhi = Chinese journal of burns","url":"https://pubmed.ncbi.nlm.nih.gov/21223656","citation_count":1,"is_preprint":false},{"pmid":"40939588","id":"PMC_40939588","title":"Structural analysis of ASCH domain-containing proteins and their implications for nucleotide processing.","date":"2025","source":"Structure (London, England : 1993)","url":"https://pubmed.ncbi.nlm.nih.gov/40939588","citation_count":1,"is_preprint":false},{"pmid":"21223654","id":"PMC_21223654","title":"[Subcellular localization of human endothelial-overexpressed lipopolysaccharide-associated factor 1 protein].","date":"2010","source":"Zhonghua shao shang za zhi = Zhonghua shaoshang zazhi = Chinese journal of burns","url":"https://pubmed.ncbi.nlm.nih.gov/21223654","citation_count":1,"is_preprint":false},{"pmid":"16383041","id":"PMC_16383041","title":"[Purification of human endothelial overexpressed lipopolysaccharide-associated factor 1 protein].","date":"2005","source":"Zhonghua shao shang za zhi = Zhonghua shaoshang zazhi = Chinese journal of burns","url":"https://pubmed.ncbi.nlm.nih.gov/16383041","citation_count":1,"is_preprint":false},{"pmid":"31569543","id":"PMC_31569543","title":"Crystal Structure of Human EOLA1 Implies Its Possibility of RNA Binding.","date":"2019","source":"Molecules (Basel, Switzerland)","url":"https://pubmed.ncbi.nlm.nih.gov/31569543","citation_count":0,"is_preprint":false},{"pmid":"33062250","id":"PMC_33062250","title":"Corrigendum to \"EOLA1 Inhibits Lipopolysaccharide-Induced Vascular Cell Adhesion Molecule-1 Expression by Association with MT2A in ECV304 Cells\".","date":"2020","source":"International journal of inflammation","url":"https://pubmed.ncbi.nlm.nih.gov/33062250","citation_count":0,"is_preprint":false},{"pmid":"16185414","id":"PMC_16185414","title":"[Establishment of human endothelial-overexpressed lipopolysaccharide-associated factor 1 compelling expression model and its effects on the proliferation of ECV304 cells].","date":"2005","source":"Zhonghua shao shang za zhi = Zhonghua shaoshang zazhi = Chinese journal of burns","url":"https://pubmed.ncbi.nlm.nih.gov/16185414","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":8840,"output_tokens":1958,"usd":0.027945,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":8917,"output_tokens":2604,"usd":0.054843,"stage2_stop_reason":"end_turn"},"total_usd":0.082788,"stage1_batch_id":"msgbatch_01AsR7g8dGTEiEw9U83bJeWn","stage2_batch_id":"msgbatch_01YV5ZyxtFNUZEjxRcpd9jzb","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2004,\n      \"finding\": \"EOLA1 physically interacts with metallothionein 2A (MT2A), identified by yeast two-hybrid screening of a human liver cDNA library using EOLA1 cDNA as bait, and confirmed by co-immunoprecipitation; stable transfection of EOLA1 stimulates ECV304 cell proliferation.\",\n      \"method\": \"Yeast two-hybrid screening + co-immunoprecipitation; stable transfection + cell proliferation assay\",\n      \"journal\": \"Biochemical and biophysical research communications / Chinese journal of medical genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal co-IP confirmed the Y2H interaction; proliferation phenotype shown by stable transfection; two papers from same lab using orthogonal methods\",\n      \"pmids\": [\"15541360\", \"16215939\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Inducible overexpression of EOLA1 in ECV304 cells significantly enhances cell proliferation, establishing a functional role for EOLA1 protein in promoting endothelial cell proliferation.\",\n      \"method\": \"Inducible overexpression vector (pOPRSVI-EOLA1) transfection; cell counting growth curve\",\n      \"journal\": \"Chinese journal of burns\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, single overexpression method, no mechanistic pathway placement\",\n      \"pmids\": [\"16185414\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Knockdown of EOLA1 expression by siRNA in ECV304 cells slows cell proliferation, confirming EOLA1 is required for normal endothelial cell proliferation.\",\n      \"method\": \"siRNA knockdown + cell proliferation counting\",\n      \"journal\": \"Chinese journal of medical genetics\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, single RNAi method, phenotype only\",\n      \"pmids\": [\"17557240\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"EOLA1 protein localizes to both the nucleus and cytoplasmic matrix of ECV304 endothelial cells, consistent with a role as a signal transduction factor.\",\n      \"method\": \"EGFP-EOLA1 fusion protein expression, laser scanning confocal microscopy, and immunoelectron microscopy\",\n      \"journal\": \"Chinese journal of burns\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — two orthogonal direct localization methods (confocal + immunoelectron microscopy), single lab\",\n      \"pmids\": [\"21223654\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"EOLA1 acts as a negative regulator of LPS-induced IL-6 production and apoptosis in HUVECs by regulating MT2A; LPS induces EOLA1 expression in a time-dependent manner, deletion of EOLA1 promotes LPS-induced IL-6 production and apoptosis, and MT2A is activated by LPS and plays a key role in LPS-induced IL-6 expression downstream of EOLA1.\",\n      \"method\": \"siRNA knockdown of EOLA1 and MT2A, LPS stimulation, IL-6 measurement, apoptosis assay in HUVECs\",\n      \"journal\": \"Molecular and cellular biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function (siRNA) with defined cytokine and apoptosis readouts; pathway placement of EOLA1 upstream of MT2A; single lab\",\n      \"pmids\": [\"24916366\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"EOLA1 negatively regulates LPS-induced VCAM-1 expression in ECV304 cells via its association with MT2A; knockdown of EOLA1 enhances LPS-induced VCAM-1 production whereas overexpression reduces it; MT2A knockdown also reduces LPS-induced VCAM-1 production. EOLA1 subcellular localization is broadly diffuse in the cells.\",\n      \"method\": \"siRNA knockdown and overexpression of EOLA1; siRNA knockdown of MT2A; VCAM-1 quantification after LPS treatment; subcellular localization assessment\",\n      \"journal\": \"International journal of inflammation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal gain- and loss-of-function with defined molecular readout; pathway placement through MT2A; single lab\",\n      \"pmids\": [\"26881174\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Crystal structure of human EOLA1 was determined at 1.71 Å resolution, revealing a characteristic β-barrel fold surrounded by α-helices structurally similar to an ASC-1 homology (ASCH) domain. EOLA1 retains a conserved 'GxKxxExR' motif in its cavity, and the cavity contains aromatic and polar residues implying nucleotide binding capacity. A positively charged cleft is present, similar to YTH domain proteins and a ribonuclease-active ASCH protein, suggesting potential RNA binding activity.\",\n      \"method\": \"X-ray crystallography (1.71 Å resolution crystal structure)\",\n      \"journal\": \"Molecules\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — high-resolution crystal structure with detailed structural analysis; single lab but rigorous structural method\",\n      \"pmids\": [\"31569543\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Crystal structures of mouse EOLA1 were determined, revealing differences in substrate preferences compared to homologous ASCH domain proteins (EcYqfB and human TRIP4-ASCH domain). The E. coli ASCH domain protein YqfB has amidohydrolase activity converting N4-acetylcytidine (ac4C) to cytidine, placing EOLA1 in the ASCH structural family but showing distinct substrate selectivity.\",\n      \"method\": \"X-ray crystallography of mouse EOLA1 and comparative structural analysis with EcYqfB and TRIP4-ASCH\",\n      \"journal\": \"Structure\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — crystal structure of mouse EOLA1 determined, but functional characterization of EOLA1's own catalytic activity was not directly shown; primary data on activity was for EcYqfB\",\n      \"pmids\": [\"40939588\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"EOLA1 is a nuclear and cytoplasmic endothelial protein with an ASCH-domain β-barrel fold that physically interacts with metallothionein 2A (MT2A) and acts as a negative regulator of LPS-induced inflammatory responses (IL-6 production, VCAM-1 expression) and apoptosis in endothelial cells; its crystal structure reveals a conserved nucleotide-binding cavity and positively charged RNA-binding cleft, suggesting it may control gene expression through RNA binding, while its interaction with MT2A mediates its anti-inflammatory signaling and its expression promotes endothelial cell proliferation.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"EOLA1 is an endothelial protein that functions as a negative regulator of LPS-induced inflammatory signaling and apoptosis while promoting endothelial cell proliferation [#4, #0]. It physically interacts with metallothionein 2A (MT2A), and this association is central to its anti-inflammatory output: loss of EOLA1 enhances LPS-induced IL-6 production and apoptosis in HUVECs, with MT2A acting downstream as a key effector of IL-6 expression [#0, #4]. EOLA1 likewise restrains LPS-induced VCAM-1 expression through its MT2A partnership, with knockdown raising and overexpression lowering VCAM-1 [#5]. EOLA1 localizes to both nucleus and cytoplasm, consistent with a signal-transduction role [#3]. Structurally, EOLA1 adopts an ASC-1 homology (ASCH) β-barrel fold with a conserved 'GxKxxExR' cavity implying nucleotide binding and a positively charged cleft resembling that of RNA-binding ASCH and YTH domain proteins [#6]; comparative crystallography of the mouse ortholog places it in the ASCH family but with substrate selectivity distinct from the ac4C amidohydrolase EcYqfB and human TRIP4-ASCH [#7]. A catalytic or RNA-binding activity intrinsic to EOLA1 has not been directly demonstrated in the available corpus.\",\n  \"teleology\": [\n    {\n      \"year\": 2004,\n      \"claim\": \"Established the first molecular partner of EOLA1 and its first cellular phenotype, framing it as an endothelial protein that interacts with MT2A and drives proliferation.\",\n      \"evidence\": \"Yeast two-hybrid screen of a human liver cDNA library with reciprocal co-IP confirmation; stable transfection proliferation assay in ECV304 cells\",\n      \"pmids\": [\"15541360\", \"16215939\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Interaction mapped only by Y2H/co-IP, no interface or domain mapping\", \"Functional consequence of the MT2A interaction not yet defined\", \"Proliferation effect tied to a single cell line\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Tested whether elevated EOLA1 protein is sufficient to drive proliferation, reinforcing a pro-proliferative role.\",\n      \"evidence\": \"Inducible overexpression (pOPRSVI-EOLA1) and growth-curve counting in ECV304 cells\",\n      \"pmids\": [\"16185414\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single overexpression method without mechanistic pathway placement\", \"No molecular readout linking EOLA1 to a proliferation pathway\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Addressed whether EOLA1 is required for proliferation, complementing the gain-of-function data with loss-of-function evidence.\",\n      \"evidence\": \"siRNA knockdown plus proliferation counting in ECV304 cells\",\n      \"pmids\": [\"17557240\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Phenotype only, no mechanism\", \"Single RNAi method, no rescue control\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Determined where EOLA1 acts in the cell, supporting a dual nuclear/cytoplasmic signal-transduction role.\",\n      \"evidence\": \"EGFP-EOLA1 fusion imaged by laser scanning confocal and immunoelectron microscopy in ECV304 cells\",\n      \"pmids\": [\"21223654\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Localization shown via overexpressed fusion, not endogenous protein\", \"Functional significance of nuclear vs cytoplasmic pools not dissected\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Placed EOLA1 in an inflammatory signaling pathway, showing it acts upstream of MT2A to restrain LPS-induced IL-6 and apoptosis.\",\n      \"evidence\": \"siRNA knockdown of EOLA1 and MT2A, LPS stimulation, IL-6 and apoptosis readouts in HUVECs\",\n      \"pmids\": [\"24916366\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular mechanism by which EOLA1 regulates MT2A unresolved\", \"Single lab and cell system\", \"How LPS induces EOLA1 expression not defined\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Extended EOLA1's anti-inflammatory role to adhesion molecule control, showing MT2A-dependent suppression of LPS-induced VCAM-1.\",\n      \"evidence\": \"Reciprocal siRNA knockdown and overexpression of EOLA1, MT2A knockdown, VCAM-1 quantification after LPS in ECV304 cells\",\n      \"pmids\": [\"26881174\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct biochemical link between EOLA1-MT2A binding and VCAM-1 transcription not established\", \"Diffuse localization not correlated with signaling state\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Provided the first structural framework, assigning EOLA1 to the ASCH β-barrel fold with cavity features implying nucleotide and RNA binding.\",\n      \"evidence\": \"1.71 Å X-ray crystal structure of human EOLA1 with structural comparison to YTH and ASCH proteins\",\n      \"pmids\": [\"31569543\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No experimental demonstration of nucleotide or RNA binding\", \"No ligand-bound structure\", \"Structural inference not linked to the MT2A/inflammatory function\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Compared EOLA1 against catalytically active ASCH proteins, refining family placement and indicating distinct substrate selectivity.\",\n      \"evidence\": \"X-ray crystallography of mouse EOLA1 with comparative analysis against EcYqfB and TRIP4-ASCH\",\n      \"pmids\": [\"40939588\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"EOLA1's own catalytic activity not directly measured\", \"Activity data shown only for EcYqfB, not EOLA1\", \"No identified physiological substrate\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"Whether EOLA1 possesses intrinsic catalytic or RNA-binding activity and how this connects to its MT2A-dependent anti-inflammatory function remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No demonstrated enzymatic or RNA-binding activity for EOLA1 itself\", \"Mechanistic bridge between the structural fold and the inflammatory phenotype absent\", \"No in vivo or disease-level validation\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [3]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [3]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [4, 5]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"MT2A\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}