{"gene":"EYA3","run_date":"2026-06-09T23:54:43","timeline":{"discoveries":[{"year":2018,"finding":"The Ser/Thr phosphatase activity of Eya3 is not intrinsic but arises from its direct interaction with the PP2A-B55α holoenzyme. Eya3 alters PP2A regulation of c-Myc by enabling PP2A-B55α to dephosphorylate pT58 on c-Myc, increasing c-Myc stability—in direct contrast to PP2A-B56α-mediated dephosphorylation of pS62 which destabilizes c-Myc. Eya3 and PP2A-B55α promote metastasis in a xenograft model of breast cancer.","method":"Co-immunoprecipitation, in vitro phosphatase assays, mutagenesis, xenograft model","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — multiple orthogonal methods including in vitro assays, Co-IP, and in vivo xenograft; subsequently replicated and structurally validated by later studies","pmids":["29535359"],"is_preprint":false},{"year":2025,"finding":"Cryo-EM and NMR structures of PP2A:B55 with Eya3 reveal that Eya3 binds B55α through an extended peptide in the N-terminal domain. Eya3 and PP2A substrates share a common set of interaction pockets on B55α but with distinct binding mechanisms. The core B55 recruitment motif in Eya3 is conserved across the Eya family. NMR-based dephosphorylation assays demonstrated that B55 recruitment by Eya3 directs selective dephosphorylation of specific phosphosites.","method":"Cryo-electron microscopy, NMR spectroscopy, NMR-based dephosphorylation assays","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structure plus NMR functional validation in a single rigorous study, independently corroborated by a second structural study (PMID:40414499)","pmids":["40247147"],"is_preprint":false},{"year":2025,"finding":"Cryo-EM structures of PP2A-B55α bound with Eya3 show that Eya3 binds B55α through an extended peptide in the Eya3 N-terminal domain. A competitive inhibitory peptide (B55i) disrupts the B55α–Eya3 interaction in vitro; when expressed in TNBC cells, B55i increases Myc pT58 and decreases Myc protein levels, confirming functional relevance of the Eya3–PP2A-B55α interaction for Myc stability.","method":"Cryo-electron microscopy, in vitro binding/inhibition assays, cell-based Myc phosphorylation/protein level assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — cryo-EM structure plus functional cell-based validation; single lab but multiple orthogonal methods","pmids":["40414499","39975004"],"is_preprint":false},{"year":2018,"finding":"Eya3 utilizes its Thr phosphatase activity (via PP2A-B55α) to dephosphorylate Myc at pT58, producing stabilized Myc, which is required for Eya3-mediated upregulation of PD-L1. Eya3 loss decreases tumor growth in immune-competent mice, increases CD8+ T cell infiltration, and CD8+ T cell depletion reverses the effects of Eya3 knockdown.","method":"Knockdown experiments, phosphatase assays, CD8+ T cell depletion, in vivo tumor growth assays, rescue experiments (PD-L1 re-expression)","journal":"The Journal of clinical investigation","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods including in vitro phosphatase assays, in vivo immune depletion, and functional rescue experiments","pmids":["29757193"],"is_preprint":false},{"year":2025,"finding":"EYA3 up-regulates NF-κB signaling to enhance CCL2 expression, which suppresses cytotoxic NK cell activation and infiltration into the pre-metastatic niche (PMN), thereby promoting TNBC metastasis. Restoration of NF-κB signaling downstream of Eya3 knockdown rescues metastasis without restoring primary tumor growth, isolating EYA3/NF-κB effects to the metastatic site.","method":"Knockdown experiments, NF-κB pathway rescue, in vitro NK cell activation assays, in vivo PMN analysis","journal":"Science advances","confidence":"High","confidence_rationale":"Tier 2 / Strong — epistatic rescue experiments combined with in vitro and in vivo validation across multiple orthogonal methods; published peer-reviewed and preprint versions","pmids":["40333987","39211066"],"is_preprint":false},{"year":2018,"finding":"EYA3 tyrosine phosphatase activity promotes the survival of pulmonary arterial smooth muscle cells under DNA-damaging conditions. Transgenic mice with an inactivating mutation in the EYA3 tyrosine phosphatase domain are significantly protected from vascular remodeling. Pharmacological inhibition of EYA3 tyrosine phosphatase substantially reverses vascular remodeling in a rat model of pulmonary hypertension.","method":"Transgenic mouse with phosphatase-dead EYA3 mutation, pharmacological inhibition, rat disease model","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — active-site mutagenesis in vivo, pharmacological inhibition, and disease model with clear phenotypic readout","pmids":["31515519"],"is_preprint":false},{"year":2018,"finding":"WDR1 is a substrate of EYA3 tyrosine phosphatase but not EYA1. Src kinase phosphorylates tyrosine residues in EYA3, and EYA3 can autodephosphorylate these residues. Src phosphorylation of EYA3 controls its subcellular localization (nuclear and cytoskeletal). EYA3-mediated dephosphorylation of WDR1 induces major changes in cellular actin cytoskeleton organization.","method":"Phosphotyrosine peptide microarray, in vitro phosphatase assays, subcellular fractionation/localization, actin cytoskeleton imaging","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vitro phosphatase assay with substrate identified by peptide microarray, functional cytoskeletal readout; single lab","pmids":["29440662"],"is_preprint":false},{"year":2019,"finding":"Src kinase phosphorylates EYA3 at 13 tyrosine residues with different phosphorylation and autodephosphorylation kinetics. Residues Y77, Y96, and Y237 are involved in cell proliferation; mutation of these three residues abolishes the pro-proliferative effect of EYA3 overexpression. EYA3 controls its own phosphorylation state through autodephosphorylation.","method":"Native and bottom-up mass spectrometry, site-directed mutagenesis, cell cycle analysis in HEK293T cells","journal":"International journal of molecular sciences","confidence":"Medium","confidence_rationale":"Tier 1-2 / Moderate — mutagenesis plus mass spectrometry mapping and functional cell cycle assay; single lab","pmids":["31847183"],"is_preprint":false},{"year":2012,"finding":"EYA3 is regulated by the EWS/FLI1 fusion protein via repression of miR-708, which targets the EYA3 3'-UTR, rather than through direct promoter binding. EYA3 knockdown in Ewing sarcoma cells sensitizes them to DNA-damaging chemotherapeutics and impairs DNA damage repair, implicating EYA3 as a mediator of chemoresistance.","method":"miRNA luciferase reporter assay, EYA3 knockdown, DNA damage/repair assays, chemotherapy sensitivity assays","journal":"Molecular cancer research : MCR","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reporter assay establishing miR-708 targeting of EYA3 3'-UTR, knockdown with functional DNA repair readout; single lab","pmids":["22723308"],"is_preprint":false},{"year":2021,"finding":"EYA3 tyrosine phosphatase activity regulates VEGFA levels in Ewing sarcoma tumors and promotes DNA damage repair and cell survival. Pharmacological inhibition of EYA3 tyrosine phosphatase (benzarone) inhibits tumor growth and angiogenesis. Elevated EYA3 substrate H2AX-pY142 upon EYA3 loss confirms H2AX-pY142 as an EYA3 substrate in vivo.","method":"Genetic knockdown and pharmacological inhibition, xenograft tumor model, patient-derived xenograft, H2AX-pY142 substrate measurement","journal":"Molecular cancer therapeutics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo pharmacological and genetic approaches with substrate engagement readout; single lab","pmids":["33649104"],"is_preprint":false},{"year":2010,"finding":"Eya3 and its partner Six1 synergistically activate TSHβ expression in the pars tuberalis upon long-day photoperiod stimulation, and this activation is further enhanced by Tef and Hlf. Eya3 expression is acutely induced by late-night light stimulation and precedes TSHβ induction, placing Eya3 upstream of TSHβ in the photoperiodic pathway.","method":"Genome-wide expression analysis, transcriptional reporter assays, acute light stimulation experiments in CBA/N mice","journal":"Current biology : CB","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — transcriptional reporter assay establishing synergistic activation, genome-wide expression analysis for pathway placement; replicated in sheep (PMID:20434341)","pmids":["21129973","20434341"],"is_preprint":false},{"year":2008,"finding":"Ski directly interacts with Six1 and Eya3 via the Dachshund homology domain of Ski to form a complex that binds the MEF3 site on the Myog regulatory region and activates myogenin (Myog) transcription during muscle terminal differentiation. This interaction is required for Ski-mediated promotion of myoblast differentiation.","method":"Chromatin immunoprecipitation (ChIP), transcriptional reporter assays, retroviral overexpression/knockdown, co-immunoprecipitation","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP and ChIP with reporter assays; single lab","pmids":["19008232"],"is_preprint":false},{"year":2022,"finding":"Hypoxia-induced EYA3 couples with SIX5 and the histone acetyltransferase p300 to form a complex that transactivates EGFR, VEGFD, and multiple MMPs (MMP3, MMP7, MMP8, MMP21, MMP26) by binding their promoters in colorectal cancer. Disruption of the EYA3-SIX5-p300 complex decreases expression of these targets and inhibits CRC cell growth.","method":"Co-immunoprecipitation, mass spectrometry, chromatin immunoprecipitation (ChIP), tumor xenograft model, benzarone inhibitor treatment","journal":"Annals of translational medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP/MS for complex identification plus ChIP for promoter occupancy plus in vivo xenograft; single lab","pmids":["35957720"],"is_preprint":false},{"year":2023,"finding":"RBFOX2 regulates alternative splicing of EYA3 exon 7, generating tissue-specific isoforms. Different EYA3 isoforms differentially partner with either SIX4 or ZBTB1 transcription factors to dictate gene expression during myogenesis. EYA3 expression is required for myoblast proliferation and differentiation.","method":"Mass spectrometry-based proteomics, genome-wide transcriptomics, knockdown experiments, alternative splicing analysis","journal":"iScience","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — MS-based interactome combined with transcriptomic analysis and functional knockdown; single lab","pmids":["38026174"],"is_preprint":false},{"year":2021,"finding":"A missense variant in EYA3 (p.Asn358Ser) increases the half-life of the mutated protein without affecting its ability to dephosphorylate H2AFX following DNA damage repair pathway induction. Knockdown of eya3 in zebrafish embryos produces craniofacial abnormalities consistent with oculo-auriculo-vertebral spectrum.","method":"Cellular protein stability assay, H2AFX dephosphorylation assay, zebrafish knockdown morphant analysis, exome sequencing","journal":"Human genetics","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — in vitro phosphatase activity assay and in vivo zebrafish model; single lab","pmids":["33475861"],"is_preprint":false},{"year":2024,"finding":"EYA3 promotes gastric cancer cell proliferation by activating the mTORC1 signaling pathway and inhibiting autophagy. EYA3 silencing reduces cell proliferation in vitro and slows tumor growth in vivo in a xenograft model.","method":"Gene silencing, gene set enrichment analysis, in vitro proliferation assays, xenograft tumor model","journal":"Scientific reports","confidence":"Low","confidence_rationale":"Tier 3 / Weak — pathway association by GSEA with knockdown phenotype; no direct biochemical mechanism established; single lab","pmids":["39550476"],"is_preprint":false}],"current_model":"EYA3 is a multifunctional protein that acts as a transcriptional co-activator (partnering with SIX, SKI, and p300-containing complexes to regulate developmental and oncogenic target genes) and as a tyrosine phosphatase (dephosphorylating substrates including H2AX-pY142 to promote DNA damage repair and cell survival, and WDR1 to remodel the actin cytoskeleton); its apparent Ser/Thr phosphatase activity is not intrinsic but arises from direct recruitment of the PP2A-B55α holoenzyme—an interaction structurally defined by cryo-EM—through which EYA3 directs dephosphorylation of c-Myc pT58 to stabilize Myc and drive PD-L1 upregulation, immune evasion, and tumor progression, while its own tyrosine phosphorylation state is dynamically regulated by Src kinase and EYA3 autodephosphorylation to control subcellular localization and proliferative signaling."},"narrative":{"mechanistic_narrative":"EYA3 is a dual-function protein that operates both as a transcriptional co-activator in developmental and oncogenic programs and as a tyrosine phosphatase that controls DNA damage repair, cytoskeletal dynamics, and cell survival [PMID:31515519, PMID:29440662, PMID:19008232]. As a co-activator, EYA3 partners with SIX-family transcription factors and additional cofactors at target promoters: it synergizes with Six1 (enhanced by Tef and Hlf) to drive photoperiodic TSHβ induction [PMID:21129973, PMID:20434341], joins Ski and Six1 at the MEF3 element to activate myogenin during muscle differentiation [PMID:19008232], and under hypoxia assembles with SIX5 and the acetyltransferase p300 to transactivate EGFR, VEGFD, and multiple MMPs in colorectal cancer [PMID:35957720]; isoform choice driven by RBFOX2-dependent splicing of exon 7 redirects EYA3 between SIX4 and ZBTB1 partners during myogenesis [PMID:38026174]. Its intrinsic catalytic activity is a tyrosine phosphatase that dephosphorylates H2AX-pY142 to promote DNA repair and cell survival and WDR1 to remodel the actin cytoskeleton, with Src-mediated tyrosine phosphorylation and EYA3 autodephosphorylation tuning its localization and proliferative output [PMID:29440662, PMID:31847183, PMID:33649104]. EYA3's apparent Ser/Thr phosphatase activity is not intrinsic but arises from direct recruitment of the PP2A-B55α holoenzyme through an extended N-terminal peptide defined by cryo-EM, redirecting PP2A-B55α to dephosphorylate c-Myc pT58 and thereby stabilize Myc [PMID:29535359, PMID:40247147, PMID:40414499, PMID:39975004]. Through this Myc-stabilizing axis and NF-κB/CCL2 signaling, EYA3 upregulates PD-L1, suppresses CD8+ T cell and NK cell anti-tumor responses, and promotes breast cancer growth and metastasis [PMID:29757193, PMID:40333987, PMID:39211066]. A missense EYA3 variant and zebrafish knockdown link the gene to oculo-auriculo-vertebral spectrum craniofacial development [PMID:33475861].","teleology":[{"year":2008,"claim":"Established EYA3 as a component of a sequence-specific transcriptional activator complex, answering how it contributes to developmental gene programs.","evidence":"ChIP, reciprocal Co-IP, and reporter assays showing Ski-Six1-Eya3 binding to the Myog MEF3 site during myoblast differentiation","pmids":["19008232"],"confidence":"Medium","gaps":["Direct DNA contact by EYA3 itself not demonstrated","Role of phosphatase activity in this transcriptional function unaddressed"]},{"year":2010,"claim":"Placed Eya3 upstream of TSHβ in the photoperiodic response, defining a physiological context for its co-activator function with Six1.","evidence":"Genome-wide expression analysis and transcriptional reporter assays with acute light stimulation in mice, replicated in sheep","pmids":["21129973","20434341"],"confidence":"Medium","gaps":["Molecular mechanism of light-induced Eya3 induction not resolved","Whether catalytic activity is required for TSHβ activation unknown"]},{"year":2012,"claim":"Showed how EYA3 is post-transcriptionally controlled in cancer and linked it to DNA repair-mediated chemoresistance.","evidence":"miR-708 3'-UTR luciferase reporter, EYA3 knockdown, and DNA damage/chemosensitivity assays in Ewing sarcoma","pmids":["22723308"],"confidence":"Medium","gaps":["Direct EYA3 substrate in the repair pathway not identified here","Phosphatase requirement not tested"]},{"year":2018,"claim":"Resolved the long-standing puzzle of EYA Ser/Thr phosphatase activity by showing it is borrowed from PP2A-B55α and redirected to stabilize c-Myc via pT58 dephosphorylation.","evidence":"Co-IP, in vitro phosphatase assays, mutagenesis, and breast cancer xenograft metastasis model","pmids":["29535359"],"confidence":"High","gaps":["Structural basis of recruitment not yet defined at this stage","Generality across substrates beyond c-Myc unclear"]},{"year":2018,"claim":"Defined EYA3 substrate specificity and regulation of its tyrosine phosphatase, identifying WDR1 as a substrate and Src as the upstream kinase controlling localization.","evidence":"Phosphotyrosine peptide microarray, in vitro phosphatase assays, subcellular fractionation, and actin imaging","pmids":["29440662"],"confidence":"Medium","gaps":["WDR1 dephosphorylation site not mapped","Single-lab finding without reciprocal validation"]},{"year":2018,"claim":"Demonstrated that EYA3-directed Myc stabilization drives immune evasion via PD-L1, linking the phosphatase axis to anti-tumor immunity.","evidence":"Knockdown, phosphatase assays, CD8+ T cell depletion, and PD-L1 rescue in immune-competent mice","pmids":["29757193"],"confidence":"High","gaps":["Mechanism linking Myc to PD-L1 transcription not fully detailed","Tissue scope beyond model unclear"]},{"year":2018,"claim":"Validated EYA3 tyrosine phosphatase activity as a druggable driver of cell survival under DNA damage in a non-cancer disease setting.","evidence":"Phosphatase-dead transgenic mouse and pharmacological inhibition in rat pulmonary hypertension model","pmids":["31515519"],"confidence":"High","gaps":["Relevant substrate(s) in pulmonary arterial smooth muscle not pinpointed","Selectivity of inhibitor for EYA3 vs paralogs unaddressed"]},{"year":2019,"claim":"Mapped Src phosphorylation sites on EYA3 and tied specific tyrosines to its proliferative function, refining the autodephosphorylation regulatory loop.","evidence":"Native and bottom-up mass spectrometry, site-directed mutagenesis, and cell cycle analysis in HEK293T cells","pmids":["31847183"],"confidence":"Medium","gaps":["Downstream effectors of proliferative tyrosines not identified","In vivo relevance of autodephosphorylation kinetics untested"]},{"year":2021,"claim":"Confirmed H2AX-pY142 as an in vivo EYA3 substrate and linked tyrosine phosphatase activity to angiogenesis and repair-driven tumor survival.","evidence":"Genetic knockdown, benzarone inhibition, xenograft and patient-derived xenograft with H2AX-pY142 readout","pmids":["33649104"],"confidence":"Medium","gaps":["Mechanism connecting H2AX dephosphorylation to VEGFA regulation unclear","Single-lab finding"]},{"year":2021,"claim":"Connected EYA3 to a human developmental disorder, showing a stabilizing missense variant and a craniofacial phenotype on knockdown.","evidence":"Protein stability and H2AFX dephosphorylation assays, exome sequencing, and zebrafish morphant analysis","pmids":["33475861"],"confidence":"Medium","gaps":["Causality of the variant beyond a single family not established","Mechanism linking increased half-life to phenotype unknown"]},{"year":2022,"claim":"Identified a hypoxia-induced EYA3-SIX5-p300 transcriptional complex driving pro-tumorigenic target genes in colorectal cancer.","evidence":"Co-IP/MS, ChIP for promoter occupancy, xenograft, and benzarone treatment","pmids":["35957720"],"confidence":"Medium","gaps":["Whether phosphatase activity is required for complex function not separated","Single-lab finding"]},{"year":2023,"claim":"Showed that RBFOX2-dependent alternative splicing of EYA3 dictates which transcription-factor partner it engages during myogenesis.","evidence":"MS-based interactome proteomics, transcriptomics, and knockdown with alternative splicing analysis","pmids":["38026174"],"confidence":"Medium","gaps":["Functional consequences of SIX4 vs ZBTB1 partnering on specific genes not fully resolved","Single-lab finding"]},{"year":2025,"claim":"Provided the structural basis for EYA3 recruitment of PP2A-B55α and demonstrated that recruitment dictates phosphosite-selective dephosphorylation.","evidence":"Cryo-EM, NMR structures and NMR-based dephosphorylation assays of PP2A:B55 with Eya3, plus a competitive B55i peptide that elevates Myc pT58 in TNBC cells","pmids":["40247147","40414499","39975004"],"confidence":"High","gaps":["Whether other Eya-family members redirect PP2A to distinct substrates in cells untested","Therapeutic feasibility of disrupting the interaction in vivo unestablished"]},{"year":2025,"claim":"Extended EYA3's pro-metastatic role to innate immunity, showing NF-κB/CCL2-mediated suppression of NK cells in the pre-metastatic niche.","evidence":"Knockdown with NF-κB pathway rescue, in vitro NK activation assays, and in vivo pre-metastatic niche analysis in TNBC","pmids":["40333987","39211066"],"confidence":"High","gaps":["Direct mechanism by which EYA3 activates NF-κB not defined","Relationship between NF-κB and the Myc/PD-L1 axis unresolved"]},{"year":null,"claim":"How EYA3 integrates its transcriptional co-activator and dual phosphatase activities within a single cell, and whether these functions are coordinately or independently deployed across tissues, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified model linking catalytic and co-activator roles","Substrate repertoire of the tyrosine phosphatase incompletely mapped","Mechanism coupling EYA3 to mTORC1/autophagy not biochemically defined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[5,6,9]},{"term_id":"GO:0016787","term_label":"hydrolase activity","supporting_discovery_ids":[0,6,9]},{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[10,11,12]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[0,1,2]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[6]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[6]}],"pathway":[{"term_id":"R-HSA-73894","term_label":"DNA Repair","supporting_discovery_ids":[8,9,14]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[10,11,12]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[3,4]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[3,4,5]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[11,13,14]}],"complexes":["EYA3-SIX5-p300","Ski-Six1-Eya3","PP2A-B55α (recruited holoenzyme)"],"partners":["PP2A-B55Α","SIX1","SIX5","SIX4","EP300","SKI","SRC","ZBTB1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q99504","full_name":"Protein phosphatase EYA3","aliases":["Eyes absent homolog 3"],"length_aa":573,"mass_kda":62.7,"function":"Tyrosine phosphatase that specifically dephosphorylates 'Tyr-142' of histone H2AX (H2AXY142ph). 'Tyr-142' phosphorylation of histone H2AX plays a central role in DNA repair and acts as a mark that distinguishes between apoptotic and repair responses to genotoxic stress. Promotes efficient DNA repair by dephosphorylating H2AX, promoting the recruitment of DNA repair complexes containing MDC1 (PubMed:19234442, PubMed:19351884). Its function as histone phosphatase probably explains its role in transcription regulation during organogenesis. Coactivates SIX1, and seems to coactivate SIX2, SIX4 and SIX5. The repression of precursor cell proliferation in myoblasts by SIX1 is switched to activation through recruitment of EYA3 to the SIX1-DACH1 complex and seems to be dependent on EYA3 phosphatase activity (By similarity). May be involved in development of the eye","subcellular_location":"Cytoplasm; Nucleus","url":"https://www.uniprot.org/uniprotkb/Q99504/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/EYA3","classification":"Not Classified","n_dependent_lines":10,"n_total_lines":1208,"dependency_fraction":0.008278145695364239},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/EYA3","total_profiled":1310},"omim":[{"mim_id":"608389","title":"BRANCHIOOTIC SYNDROME 3; BOS3","url":"https://www.omim.org/entry/608389"},{"mim_id":"603550","title":"EYA TRANSCRIPTIONAL COACTIVATOR AND PHOSPHATASE 4; EYA4","url":"https://www.omim.org/entry/603550"},{"mim_id":"601655","title":"EYA TRANSCRIPTIONAL COACTIVATOR AND PHOSPHATASE 3; EYA3","url":"https://www.omim.org/entry/601655"},{"mim_id":"601654","title":"EYA TRANSCRIPTIONAL COACTIVATOR AND PHOSPHATASE 2; EYA2","url":"https://www.omim.org/entry/601654"},{"mim_id":"601653","title":"EYA TRANSCRIPTIONAL COACTIVATOR AND PHOSPHATASE 1; EYA1","url":"https://www.omim.org/entry/601653"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Enhanced","locations":[{"location":"Nucleoplasm","reliability":"Enhanced"},{"location":"Centrosome","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/EYA3"},"hgnc":{"alias_symbol":["DKFZp686C132"],"prev_symbol":[]},"alphafold":{"accession":"Q99504","domains":[{"cath_id":"3.40.50.12350","chopping":"303-386_405-569","consensus_level":"medium","plddt":97.1333,"start":303,"end":569}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q99504","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q99504-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q99504-F1-predicted_aligned_error_v6.png","plddt_mean":65.19},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=EYA3","jax_strain_url":"https://www.jax.org/strain/search?query=EYA3"},"sequence":{"accession":"Q99504","fasta_url":"https://rest.uniprot.org/uniprotkb/Q99504.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q99504/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q99504"}},"corpus_meta":[{"pmid":"21129973","id":"PMC_21129973","title":"Acute induction of Eya3 by late-night light stimulation triggers TSHβ expression in photoperiodism.","date":"2010","source":"Current biology : CB","url":"https://pubmed.ncbi.nlm.nih.gov/21129973","citation_count":97,"is_preprint":false},{"pmid":"22723308","id":"PMC_22723308","title":"EWS/FLI1 regulates EYA3 in Ewing sarcoma via modulation of miRNA-708, resulting in increased cell survival and chemoresistance.","date":"2012","source":"Molecular cancer research : MCR","url":"https://pubmed.ncbi.nlm.nih.gov/22723308","citation_count":78,"is_preprint":false},{"pmid":"29535359","id":"PMC_29535359","title":"Eya3 partners with PP2A to induce c-Myc stabilization and tumor progression.","date":"2018","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/29535359","citation_count":74,"is_preprint":false},{"pmid":"20434341","id":"PMC_20434341","title":"Identification of Eya3 and TAC1 as long-day signals in the sheep pituitary.","date":"2010","source":"Current biology : CB","url":"https://pubmed.ncbi.nlm.nih.gov/20434341","citation_count":69,"is_preprint":false},{"pmid":"29757193","id":"PMC_29757193","title":"Eya3 promotes breast tumor-associated immune suppression via threonine phosphatase-mediated PD-L1 upregulation.","date":"2018","source":"The Journal of clinical investigation","url":"https://pubmed.ncbi.nlm.nih.gov/29757193","citation_count":41,"is_preprint":false},{"pmid":"19008232","id":"PMC_19008232","title":"Ski regulates muscle terminal differentiation by transcriptional activation of Myog in a complex with Six1 and Eya3.","date":"2008","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/19008232","citation_count":36,"is_preprint":false},{"pmid":"19102749","id":"PMC_19102749","title":"Pleiotropic effects in Eya3 knockout mice.","date":"2008","source":"BMC developmental biology","url":"https://pubmed.ncbi.nlm.nih.gov/19102749","citation_count":33,"is_preprint":false},{"pmid":"31515519","id":"PMC_31515519","title":"The EYA3 tyrosine phosphatase activity promotes pulmonary vascular remodeling in pulmonary arterial hypertension.","date":"2019","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/31515519","citation_count":27,"is_preprint":false},{"pmid":"33475861","id":"PMC_33475861","title":"A recurrent missense variant in EYA3 gene is associated with oculo-auriculo-vertebral spectrum.","date":"2021","source":"Human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/33475861","citation_count":21,"is_preprint":false},{"pmid":"29440662","id":"PMC_29440662","title":"WDR1 is a novel EYA3 substrate and its dephosphorylation induces modifications of the cellular actin cytoskeleton.","date":"2018","source":"Scientific 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medicine","url":"https://pubmed.ncbi.nlm.nih.gov/35957720","citation_count":11,"is_preprint":false},{"pmid":"40247147","id":"PMC_40247147","title":"Cryo-EM structures of PP2A:B55 with p107 and Eya3 define substrate recruitment.","date":"2025","source":"Nature structural & molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/40247147","citation_count":7,"is_preprint":false},{"pmid":"38026174","id":"PMC_38026174","title":"RBFOX2 regulated EYA3 isoforms partner with SIX4 or ZBTB1 to control transcription during myogenesis.","date":"2023","source":"iScience","url":"https://pubmed.ncbi.nlm.nih.gov/38026174","citation_count":7,"is_preprint":false},{"pmid":"31847183","id":"PMC_31847183","title":"Analysis of EYA3 Phosphorylation by Src Kinase Identifies Residues Involved in Cell Proliferation.","date":"2019","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/31847183","citation_count":7,"is_preprint":false},{"pmid":"34089890","id":"PMC_34089890","title":"Neuroendocrine regulation of reproduction in Atlantic cod (Gadus morhua): Evidence of Eya3 as an integrator of photoperiodic cues and nutritional regulation to initiate sexual maturation.","date":"2021","source":"Comparative biochemistry and physiology. 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Eya3 alters PP2A regulation of c-Myc by enabling PP2A-B55α to dephosphorylate pT58 on c-Myc, increasing c-Myc stability—in direct contrast to PP2A-B56α-mediated dephosphorylation of pS62 which destabilizes c-Myc. Eya3 and PP2A-B55α promote metastasis in a xenograft model of breast cancer.\",\n      \"method\": \"Co-immunoprecipitation, in vitro phosphatase assays, mutagenesis, xenograft model\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — multiple orthogonal methods including in vitro assays, Co-IP, and in vivo xenograft; subsequently replicated and structurally validated by later studies\",\n      \"pmids\": [\"29535359\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Cryo-EM and NMR structures of PP2A:B55 with Eya3 reveal that Eya3 binds B55α through an extended peptide in the N-terminal domain. Eya3 and PP2A substrates share a common set of interaction pockets on B55α but with distinct binding mechanisms. The core B55 recruitment motif in Eya3 is conserved across the Eya family. NMR-based dephosphorylation assays demonstrated that B55 recruitment by Eya3 directs selective dephosphorylation of specific phosphosites.\",\n      \"method\": \"Cryo-electron microscopy, NMR spectroscopy, NMR-based dephosphorylation assays\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structure plus NMR functional validation in a single rigorous study, independently corroborated by a second structural study (PMID:40414499)\",\n      \"pmids\": [\"40247147\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Cryo-EM structures of PP2A-B55α bound with Eya3 show that Eya3 binds B55α through an extended peptide in the Eya3 N-terminal domain. A competitive inhibitory peptide (B55i) disrupts the B55α–Eya3 interaction in vitro; when expressed in TNBC cells, B55i increases Myc pT58 and decreases Myc protein levels, confirming functional relevance of the Eya3–PP2A-B55α interaction for Myc stability.\",\n      \"method\": \"Cryo-electron microscopy, in vitro binding/inhibition assays, cell-based Myc phosphorylation/protein level assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — cryo-EM structure plus functional cell-based validation; single lab but multiple orthogonal methods\",\n      \"pmids\": [\"40414499\", \"39975004\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Eya3 utilizes its Thr phosphatase activity (via PP2A-B55α) to dephosphorylate Myc at pT58, producing stabilized Myc, which is required for Eya3-mediated upregulation of PD-L1. Eya3 loss decreases tumor growth in immune-competent mice, increases CD8+ T cell infiltration, and CD8+ T cell depletion reverses the effects of Eya3 knockdown.\",\n      \"method\": \"Knockdown experiments, phosphatase assays, CD8+ T cell depletion, in vivo tumor growth assays, rescue experiments (PD-L1 re-expression)\",\n      \"journal\": \"The Journal of clinical investigation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods including in vitro phosphatase assays, in vivo immune depletion, and functional rescue experiments\",\n      \"pmids\": [\"29757193\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"EYA3 up-regulates NF-κB signaling to enhance CCL2 expression, which suppresses cytotoxic NK cell activation and infiltration into the pre-metastatic niche (PMN), thereby promoting TNBC metastasis. Restoration of NF-κB signaling downstream of Eya3 knockdown rescues metastasis without restoring primary tumor growth, isolating EYA3/NF-κB effects to the metastatic site.\",\n      \"method\": \"Knockdown experiments, NF-κB pathway rescue, in vitro NK cell activation assays, in vivo PMN analysis\",\n      \"journal\": \"Science advances\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — epistatic rescue experiments combined with in vitro and in vivo validation across multiple orthogonal methods; published peer-reviewed and preprint versions\",\n      \"pmids\": [\"40333987\", \"39211066\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"EYA3 tyrosine phosphatase activity promotes the survival of pulmonary arterial smooth muscle cells under DNA-damaging conditions. Transgenic mice with an inactivating mutation in the EYA3 tyrosine phosphatase domain are significantly protected from vascular remodeling. Pharmacological inhibition of EYA3 tyrosine phosphatase substantially reverses vascular remodeling in a rat model of pulmonary hypertension.\",\n      \"method\": \"Transgenic mouse with phosphatase-dead EYA3 mutation, pharmacological inhibition, rat disease model\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — active-site mutagenesis in vivo, pharmacological inhibition, and disease model with clear phenotypic readout\",\n      \"pmids\": [\"31515519\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"WDR1 is a substrate of EYA3 tyrosine phosphatase but not EYA1. Src kinase phosphorylates tyrosine residues in EYA3, and EYA3 can autodephosphorylate these residues. Src phosphorylation of EYA3 controls its subcellular localization (nuclear and cytoskeletal). EYA3-mediated dephosphorylation of WDR1 induces major changes in cellular actin cytoskeleton organization.\",\n      \"method\": \"Phosphotyrosine peptide microarray, in vitro phosphatase assays, subcellular fractionation/localization, actin cytoskeleton imaging\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro phosphatase assay with substrate identified by peptide microarray, functional cytoskeletal readout; single lab\",\n      \"pmids\": [\"29440662\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Src kinase phosphorylates EYA3 at 13 tyrosine residues with different phosphorylation and autodephosphorylation kinetics. Residues Y77, Y96, and Y237 are involved in cell proliferation; mutation of these three residues abolishes the pro-proliferative effect of EYA3 overexpression. EYA3 controls its own phosphorylation state through autodephosphorylation.\",\n      \"method\": \"Native and bottom-up mass spectrometry, site-directed mutagenesis, cell cycle analysis in HEK293T cells\",\n      \"journal\": \"International journal of molecular sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1-2 / Moderate — mutagenesis plus mass spectrometry mapping and functional cell cycle assay; single lab\",\n      \"pmids\": [\"31847183\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"EYA3 is regulated by the EWS/FLI1 fusion protein via repression of miR-708, which targets the EYA3 3'-UTR, rather than through direct promoter binding. EYA3 knockdown in Ewing sarcoma cells sensitizes them to DNA-damaging chemotherapeutics and impairs DNA damage repair, implicating EYA3 as a mediator of chemoresistance.\",\n      \"method\": \"miRNA luciferase reporter assay, EYA3 knockdown, DNA damage/repair assays, chemotherapy sensitivity assays\",\n      \"journal\": \"Molecular cancer research : MCR\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reporter assay establishing miR-708 targeting of EYA3 3'-UTR, knockdown with functional DNA repair readout; single lab\",\n      \"pmids\": [\"22723308\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"EYA3 tyrosine phosphatase activity regulates VEGFA levels in Ewing sarcoma tumors and promotes DNA damage repair and cell survival. Pharmacological inhibition of EYA3 tyrosine phosphatase (benzarone) inhibits tumor growth and angiogenesis. Elevated EYA3 substrate H2AX-pY142 upon EYA3 loss confirms H2AX-pY142 as an EYA3 substrate in vivo.\",\n      \"method\": \"Genetic knockdown and pharmacological inhibition, xenograft tumor model, patient-derived xenograft, H2AX-pY142 substrate measurement\",\n      \"journal\": \"Molecular cancer therapeutics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo pharmacological and genetic approaches with substrate engagement readout; single lab\",\n      \"pmids\": [\"33649104\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Eya3 and its partner Six1 synergistically activate TSHβ expression in the pars tuberalis upon long-day photoperiod stimulation, and this activation is further enhanced by Tef and Hlf. Eya3 expression is acutely induced by late-night light stimulation and precedes TSHβ induction, placing Eya3 upstream of TSHβ in the photoperiodic pathway.\",\n      \"method\": \"Genome-wide expression analysis, transcriptional reporter assays, acute light stimulation experiments in CBA/N mice\",\n      \"journal\": \"Current biology : CB\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — transcriptional reporter assay establishing synergistic activation, genome-wide expression analysis for pathway placement; replicated in sheep (PMID:20434341)\",\n      \"pmids\": [\"21129973\", \"20434341\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Ski directly interacts with Six1 and Eya3 via the Dachshund homology domain of Ski to form a complex that binds the MEF3 site on the Myog regulatory region and activates myogenin (Myog) transcription during muscle terminal differentiation. This interaction is required for Ski-mediated promotion of myoblast differentiation.\",\n      \"method\": \"Chromatin immunoprecipitation (ChIP), transcriptional reporter assays, retroviral overexpression/knockdown, co-immunoprecipitation\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP and ChIP with reporter assays; single lab\",\n      \"pmids\": [\"19008232\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Hypoxia-induced EYA3 couples with SIX5 and the histone acetyltransferase p300 to form a complex that transactivates EGFR, VEGFD, and multiple MMPs (MMP3, MMP7, MMP8, MMP21, MMP26) by binding their promoters in colorectal cancer. Disruption of the EYA3-SIX5-p300 complex decreases expression of these targets and inhibits CRC cell growth.\",\n      \"method\": \"Co-immunoprecipitation, mass spectrometry, chromatin immunoprecipitation (ChIP), tumor xenograft model, benzarone inhibitor treatment\",\n      \"journal\": \"Annals of translational medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP/MS for complex identification plus ChIP for promoter occupancy plus in vivo xenograft; single lab\",\n      \"pmids\": [\"35957720\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"RBFOX2 regulates alternative splicing of EYA3 exon 7, generating tissue-specific isoforms. Different EYA3 isoforms differentially partner with either SIX4 or ZBTB1 transcription factors to dictate gene expression during myogenesis. EYA3 expression is required for myoblast proliferation and differentiation.\",\n      \"method\": \"Mass spectrometry-based proteomics, genome-wide transcriptomics, knockdown experiments, alternative splicing analysis\",\n      \"journal\": \"iScience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — MS-based interactome combined with transcriptomic analysis and functional knockdown; single lab\",\n      \"pmids\": [\"38026174\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"A missense variant in EYA3 (p.Asn358Ser) increases the half-life of the mutated protein without affecting its ability to dephosphorylate H2AFX following DNA damage repair pathway induction. Knockdown of eya3 in zebrafish embryos produces craniofacial abnormalities consistent with oculo-auriculo-vertebral spectrum.\",\n      \"method\": \"Cellular protein stability assay, H2AFX dephosphorylation assay, zebrafish knockdown morphant analysis, exome sequencing\",\n      \"journal\": \"Human genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — in vitro phosphatase activity assay and in vivo zebrafish model; single lab\",\n      \"pmids\": [\"33475861\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"EYA3 promotes gastric cancer cell proliferation by activating the mTORC1 signaling pathway and inhibiting autophagy. EYA3 silencing reduces cell proliferation in vitro and slows tumor growth in vivo in a xenograft model.\",\n      \"method\": \"Gene silencing, gene set enrichment analysis, in vitro proliferation assays, xenograft tumor model\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — pathway association by GSEA with knockdown phenotype; no direct biochemical mechanism established; single lab\",\n      \"pmids\": [\"39550476\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"EYA3 is a multifunctional protein that acts as a transcriptional co-activator (partnering with SIX, SKI, and p300-containing complexes to regulate developmental and oncogenic target genes) and as a tyrosine phosphatase (dephosphorylating substrates including H2AX-pY142 to promote DNA damage repair and cell survival, and WDR1 to remodel the actin cytoskeleton); its apparent Ser/Thr phosphatase activity is not intrinsic but arises from direct recruitment of the PP2A-B55α holoenzyme—an interaction structurally defined by cryo-EM—through which EYA3 directs dephosphorylation of c-Myc pT58 to stabilize Myc and drive PD-L1 upregulation, immune evasion, and tumor progression, while its own tyrosine phosphorylation state is dynamically regulated by Src kinase and EYA3 autodephosphorylation to control subcellular localization and proliferative signaling.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"EYA3 is a dual-function protein that operates both as a transcriptional co-activator in developmental and oncogenic programs and as a tyrosine phosphatase that controls DNA damage repair, cytoskeletal dynamics, and cell survival [#5, #6, #11]. As a co-activator, EYA3 partners with SIX-family transcription factors and additional cofactors at target promoters: it synergizes with Six1 (enhanced by Tef and Hlf) to drive photoperiodic TSH\\u03b2 induction [#10], joins Ski and Six1 at the MEF3 element to activate myogenin during muscle differentiation [#11], and under hypoxia assembles with SIX5 and the acetyltransferase p300 to transactivate EGFR, VEGFD, and multiple MMPs in colorectal cancer [#12]; isoform choice driven by RBFOX2-dependent splicing of exon 7 redirects EYA3 between SIX4 and ZBTB1 partners during myogenesis [#13]. Its intrinsic catalytic activity is a tyrosine phosphatase that dephosphorylates H2AX-pY142 to promote DNA repair and cell survival and WDR1 to remodel the actin cytoskeleton, with Src-mediated tyrosine phosphorylation and EYA3 autodephosphorylation tuning its localization and proliferative output [#6, #7, #9]. EYA3's apparent Ser/Thr phosphatase activity is not intrinsic but arises from direct recruitment of the PP2A-B55\\u03b1 holoenzyme through an extended N-terminal peptide defined by cryo-EM, redirecting PP2A-B55\\u03b1 to dephosphorylate c-Myc pT58 and thereby stabilize Myc [#0, #1, #2]. Through this Myc-stabilizing axis and NF-\\u03baB/CCL2 signaling, EYA3 upregulates PD-L1, suppresses CD8+ T cell and NK cell anti-tumor responses, and promotes breast cancer growth and metastasis [#3, #4]. A missense EYA3 variant and zebrafish knockdown link the gene to oculo-auriculo-vertebral spectrum craniofacial development [#14].\",\n  \"teleology\": [\n    {\n      \"year\": 2008,\n      \"claim\": \"Established EYA3 as a component of a sequence-specific transcriptional activator complex, answering how it contributes to developmental gene programs.\",\n      \"evidence\": \"ChIP, reciprocal Co-IP, and reporter assays showing Ski-Six1-Eya3 binding to the Myog MEF3 site during myoblast differentiation\",\n      \"pmids\": [\"19008232\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct DNA contact by EYA3 itself not demonstrated\", \"Role of phosphatase activity in this transcriptional function unaddressed\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Placed Eya3 upstream of TSH\\u03b2 in the photoperiodic response, defining a physiological context for its co-activator function with Six1.\",\n      \"evidence\": \"Genome-wide expression analysis and transcriptional reporter assays with acute light stimulation in mice, replicated in sheep\",\n      \"pmids\": [\"21129973\", \"20434341\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular mechanism of light-induced Eya3 induction not resolved\", \"Whether catalytic activity is required for TSH\\u03b2 activation unknown\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Showed how EYA3 is post-transcriptionally controlled in cancer and linked it to DNA repair-mediated chemoresistance.\",\n      \"evidence\": \"miR-708 3'-UTR luciferase reporter, EYA3 knockdown, and DNA damage/chemosensitivity assays in Ewing sarcoma\",\n      \"pmids\": [\"22723308\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct EYA3 substrate in the repair pathway not identified here\", \"Phosphatase requirement not tested\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Resolved the long-standing puzzle of EYA Ser/Thr phosphatase activity by showing it is borrowed from PP2A-B55\\u03b1 and redirected to stabilize c-Myc via pT58 dephosphorylation.\",\n      \"evidence\": \"Co-IP, in vitro phosphatase assays, mutagenesis, and breast cancer xenograft metastasis model\",\n      \"pmids\": [\"29535359\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of recruitment not yet defined at this stage\", \"Generality across substrates beyond c-Myc unclear\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Defined EYA3 substrate specificity and regulation of its tyrosine phosphatase, identifying WDR1 as a substrate and Src as the upstream kinase controlling localization.\",\n      \"evidence\": \"Phosphotyrosine peptide microarray, in vitro phosphatase assays, subcellular fractionation, and actin imaging\",\n      \"pmids\": [\"29440662\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"WDR1 dephosphorylation site not mapped\", \"Single-lab finding without reciprocal validation\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Demonstrated that EYA3-directed Myc stabilization drives immune evasion via PD-L1, linking the phosphatase axis to anti-tumor immunity.\",\n      \"evidence\": \"Knockdown, phosphatase assays, CD8+ T cell depletion, and PD-L1 rescue in immune-competent mice\",\n      \"pmids\": [\"29757193\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism linking Myc to PD-L1 transcription not fully detailed\", \"Tissue scope beyond model unclear\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Validated EYA3 tyrosine phosphatase activity as a druggable driver of cell survival under DNA damage in a non-cancer disease setting.\",\n      \"evidence\": \"Phosphatase-dead transgenic mouse and pharmacological inhibition in rat pulmonary hypertension model\",\n      \"pmids\": [\"31515519\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Relevant substrate(s) in pulmonary arterial smooth muscle not pinpointed\", \"Selectivity of inhibitor for EYA3 vs paralogs unaddressed\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Mapped Src phosphorylation sites on EYA3 and tied specific tyrosines to its proliferative function, refining the autodephosphorylation regulatory loop.\",\n      \"evidence\": \"Native and bottom-up mass spectrometry, site-directed mutagenesis, and cell cycle analysis in HEK293T cells\",\n      \"pmids\": [\"31847183\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Downstream effectors of proliferative tyrosines not identified\", \"In vivo relevance of autodephosphorylation kinetics untested\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Confirmed H2AX-pY142 as an in vivo EYA3 substrate and linked tyrosine phosphatase activity to angiogenesis and repair-driven tumor survival.\",\n      \"evidence\": \"Genetic knockdown, benzarone inhibition, xenograft and patient-derived xenograft with H2AX-pY142 readout\",\n      \"pmids\": [\"33649104\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism connecting H2AX dephosphorylation to VEGFA regulation unclear\", \"Single-lab finding\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Connected EYA3 to a human developmental disorder, showing a stabilizing missense variant and a craniofacial phenotype on knockdown.\",\n      \"evidence\": \"Protein stability and H2AFX dephosphorylation assays, exome sequencing, and zebrafish morphant analysis\",\n      \"pmids\": [\"33475861\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Causality of the variant beyond a single family not established\", \"Mechanism linking increased half-life to phenotype unknown\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Identified a hypoxia-induced EYA3-SIX5-p300 transcriptional complex driving pro-tumorigenic target genes in colorectal cancer.\",\n      \"evidence\": \"Co-IP/MS, ChIP for promoter occupancy, xenograft, and benzarone treatment\",\n      \"pmids\": [\"35957720\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether phosphatase activity is required for complex function not separated\", \"Single-lab finding\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Showed that RBFOX2-dependent alternative splicing of EYA3 dictates which transcription-factor partner it engages during myogenesis.\",\n      \"evidence\": \"MS-based interactome proteomics, transcriptomics, and knockdown with alternative splicing analysis\",\n      \"pmids\": [\"38026174\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional consequences of SIX4 vs ZBTB1 partnering on specific genes not fully resolved\", \"Single-lab finding\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Provided the structural basis for EYA3 recruitment of PP2A-B55\\u03b1 and demonstrated that recruitment dictates phosphosite-selective dephosphorylation.\",\n      \"evidence\": \"Cryo-EM, NMR structures and NMR-based dephosphorylation assays of PP2A:B55 with Eya3, plus a competitive B55i peptide that elevates Myc pT58 in TNBC cells\",\n      \"pmids\": [\"40247147\", \"40414499\", \"39975004\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether other Eya-family members redirect PP2A to distinct substrates in cells untested\", \"Therapeutic feasibility of disrupting the interaction in vivo unestablished\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Extended EYA3's pro-metastatic role to innate immunity, showing NF-\\u03baB/CCL2-mediated suppression of NK cells in the pre-metastatic niche.\",\n      \"evidence\": \"Knockdown with NF-\\u03baB pathway rescue, in vitro NK activation assays, and in vivo pre-metastatic niche analysis in TNBC\",\n      \"pmids\": [\"40333987\", \"39211066\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct mechanism by which EYA3 activates NF-\\u03baB not defined\", \"Relationship between NF-\\u03baB and the Myc/PD-L1 axis unresolved\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How EYA3 integrates its transcriptional co-activator and dual phosphatase activities within a single cell, and whether these functions are coordinately or independently deployed across tissues, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified model linking catalytic and co-activator roles\", \"Substrate repertoire of the tyrosine phosphatase incompletely mapped\", \"Mechanism coupling EYA3 to mTORC1/autophagy not biochemically defined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [5, 6, 9]},\n      {\"term_id\": \"GO:0016787\", \"supporting_discovery_ids\": [0, 6, 9]},\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [10, 11, 12]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [0, 1, 2]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [6]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [6]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [8, 9, 14]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [10, 11, 12]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [3, 4]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [3, 4, 5]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [11, 13, 14]}\n    ],\n    \"complexes\": [\n      \"EYA3-SIX5-p300\",\n      \"Ski-Six1-Eya3\",\n      \"PP2A-B55\\u03b1 (recruited holoenzyme)\"\n    ],\n    \"partners\": [\n      \"PP2A-B55\\u03b1\",\n      \"SIX1\",\n      \"SIX5\",\n      \"SIX4\",\n      \"EP300\",\n      \"SKI\",\n      \"SRC\",\n      \"ZBTB1\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"tie","faith_supported":6,"faith_total":6,"faith_pct":100.0}}