{"gene":"ZHX3","run_date":"2026-06-11T09:02:06","timeline":{"discoveries":[{"year":2003,"finding":"ZHX3 (956 amino acids, two zinc-finger motifs and five homeodomains) was identified as a ZHX1-interacting protein via yeast two-hybrid. ZHX3 dimerizes with both ZHX1 and ZHX3 itself (homodimerization), interacts with the activation domain of NF-YA, and functions as a ubiquitous transcriptional repressor localized in the nucleus. Domain mapping showed the dimerization domain, NF-YA interaction domain, and repressor domain all map to a region including the HD1 region; two nuclear localization signals were mapped to the N-terminal through Znf1 and the HD2 region.","method":"Yeast two-hybrid screen, molecular cloning, in vitro and in vivo protein-protein interaction assays, nuclear localization experiments, domain-mapping/deletion analysis","journal":"The Biochemical journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (yeast two-hybrid, in vitro/in vivo interaction assays, domain mapping) in a single rigorous founding study","pmids":["12659632"],"is_preprint":false},{"year":2003,"finding":"ZHX2 forms a heterodimer with ZHX3 via a region containing each HD1 domain, demonstrated by in vitro and in vivo protein-protein interaction assays using mouse ZHX2 and ZHX3.","method":"In vitro and in vivo protein-protein interaction assays (heterodimerization analysis), molecular cloning of mouse ZHX2 and ZHX3 cDNAs","journal":"Gene","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal in vitro and in vivo interaction assays with domain mapping, replicates and extends findings from PMID:12659632","pmids":["14659886"],"is_preprint":false},{"year":2009,"finding":"ZHX3 represses transcription from the promoters of cancer marker genes PKM and HKII in hepatocytes, as demonstrated by reporter gene (luciferase) analysis using the promoter regions of these genes in hepatoma cells.","method":"Reporter gene (luciferase) assay with ZHX3 expression in hepatoma cell lines","journal":"Frontiers in bioscience (Landmark edition)","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — reporter assay is a direct functional readout, but single lab, single method","pmids":["19273305"],"is_preprint":false},{"year":2021,"finding":"ZHX3 is ubiquitinated and subsequently degraded by the E3 ubiquitin ligase TRIM21, identifying TRIM21 as the writer of ZHX3 ubiquitination. Additionally, ZHX3 transcriptionally represses expression of RGS2 (regulator of G protein signaling 2), promoting migration and invasion of urothelial carcinoma of the bladder cells.","method":"Co-immunoprecipitation (coIP), mass spectrometry (MS) for TRIM21 interaction, RNA-seq for RGS2 repression, in vitro and in vivo migration/invasion assays","journal":"Cancer science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — coIP plus MS for binding partner identification, RNA-seq for downstream target, functional assays; single lab but multiple orthogonal methods","pmids":["33440047"],"is_preprint":false}],"current_model":"ZHX3 is a nuclear transcriptional repressor that homodimerizes and heterodimerizes with ZHX family members (ZHX1, ZHX2, ZHX3) via their HD1 domains, interacts with the NF-YA activation domain, represses target gene promoters (including PKM, HKII, and RGS2), and is itself regulated by TRIM21-mediated ubiquitination and proteasomal degradation."},"narrative":{"mechanistic_narrative":"ZHX3 is a ubiquitously expressed nuclear transcriptional repressor that controls target gene expression through dimerization within the ZHX family and engagement of the transcription factor NF-Y [PMID:12659632]. It contains two zinc-finger motifs and five homeodomains; its HD1 region carries the overlapping dimerization, NF-YA-interaction, and repressor activities, while nuclear localization signals map to the N-terminal/Znf1 and HD2 regions [PMID:12659632]. ZHX3 homodimerizes and heterodimerizes with other ZHX family members, including ZHX1 [PMID:12659632] and ZHX2 [PMID:14659886], and binds the activation domain of NF-YA to repress transcription [PMID:12659632]. Functionally, ZHX3 represses the promoters of the metabolic/cancer marker genes PKM and HKII in hepatocytes [PMID:19273305] and represses RGS2 to promote migration and invasion of urothelial bladder carcinoma cells [PMID:33440047]. ZHX3 protein levels are controlled by the E3 ubiquitin ligase TRIM21, which ubiquitinates ZHX3 and targets it for degradation [PMID:33440047].","teleology":[{"year":2003,"claim":"Established ZHX3's core identity by showing it is a nuclear repressor that dimerizes and contacts the general transcription factor NF-Y, defining the molecular surfaces through which it acts.","evidence":"Yeast two-hybrid screen, in vitro/in vivo interaction assays, and domain-mapping/deletion analysis of cloned ZHX3","pmids":["12659632"],"confidence":"High","gaps":["No direct DNA-binding site or target promoter identified at this stage","Repression mechanism downstream of NF-YA binding not defined","No endogenous target genes characterized"]},{"year":2003,"claim":"Extended the ZHX dimerization network by demonstrating ZHX3 heterodimerizes with ZHX2 through the HD1 domain, establishing combinatorial partnering within the family.","evidence":"Reciprocal in vitro and in vivo interaction assays with mouse ZHX2 and ZHX3 cDNAs","pmids":["14659886"],"confidence":"High","gaps":["Functional consequence of ZHX2-ZHX3 heterodimers on transcription not measured","Relative affinities and preference among homo- vs heterodimers unknown"]},{"year":2009,"claim":"Connected ZHX3 to specific gene targets by showing it represses the PKM and HKII promoters, providing the first defined endogenous-relevant transcriptional outputs.","evidence":"Luciferase reporter assays with PKM and HKII promoter regions in hepatoma cell lines","pmids":["19273305"],"confidence":"Medium","gaps":["Reporter assay only; direct binding to these promoters not shown","Single lab, single method","Effect on endogenous PKM/HKII expression and metabolism not established"]},{"year":2021,"claim":"Defined how ZHX3 abundance is controlled and linked its repressor activity to cancer cell behavior by identifying TRIM21 as the ubiquitin ligase degrading ZHX3 and RGS2 as a repressed target driving invasion.","evidence":"CoIP plus mass spectrometry for TRIM21, RNA-seq for RGS2 repression, and in vitro/in vivo migration and invasion assays in bladder urothelial carcinoma cells","pmids":["33440047"],"confidence":"Medium","gaps":["Direct ZHX3 occupancy of the RGS2 promoter not demonstrated","TRIM21 ubiquitination site(s) on ZHX3 and signals regulating this degradation unknown","Single lab; reciprocal validation of binding partners limited"]},{"year":null,"claim":"How ZHX3 selects its DNA targets and whether its repression depends on chromatin-modifying cofactors beyond NF-Y interaction remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No defined ZHX3 DNA-binding consensus despite zinc fingers and homeodomains","Co-repressor machinery executing repression not identified","Genome-wide direct binding map (e.g. ChIP-seq) absent"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[0,2,3]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[0]}],"pathway":[{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[0,2,3]}],"complexes":[],"partners":["ZHX1","ZHX2","NFYA","TRIM21"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9H4I2","full_name":"Zinc fingers and homeoboxes protein 3","aliases":["Triple homeobox protein 1","Zinc finger and homeodomain protein 3"],"length_aa":956,"mass_kda":104.7,"function":"Acts as a transcriptional repressor. Involved in the early stages of mesenchymal stem cell (MSC) osteogenic differentiation. Is a regulator of podocyte gene expression during primary glomerula disease. Binds to promoter DNA","subcellular_location":"Nucleus","url":"https://www.uniprot.org/uniprotkb/Q9H4I2/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/ZHX3","classification":"Not Classified","n_dependent_lines":1,"n_total_lines":1208,"dependency_fraction":0.0008278145695364238},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"CSNK2B","stoichiometry":0.2},{"gene":"HDAC1","stoichiometry":0.2},{"gene":"HDAC2","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/ZHX3","total_profiled":1310},"omim":[{"mim_id":"609598","title":"ZINC FINGER AND HOMEODOMAIN PROTEIN 3; ZHX3","url":"https://www.omim.org/entry/609598"},{"mim_id":"609185","title":"ZINC FINGER AND HOMEODOMAIN PROTEIN 2; ZHX2","url":"https://www.omim.org/entry/609185"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nucleoplasm","reliability":"Supported"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/ZHX3"},"hgnc":{"alias_symbol":["KIAA0395"],"prev_symbol":["TIX1"]},"alphafold":{"accession":"Q9H4I2","domains":[{"cath_id":"3.30.160","chopping":"78-155","consensus_level":"high","plddt":78.3206,"start":78,"end":155},{"cath_id":"1.10.10,1.10.10","chopping":"318-376","consensus_level":"high","plddt":92.4875,"start":318,"end":376},{"cath_id":"1.10.10.60","chopping":"504-555","consensus_level":"high","plddt":89.9431,"start":504,"end":555},{"cath_id":"1.10.10.60","chopping":"622-682","consensus_level":"high","plddt":86.9959,"start":622,"end":682},{"cath_id":"1.10.10.60","chopping":"771-839","consensus_level":"medium","plddt":91.0119,"start":771,"end":839},{"cath_id":"1.10.10.60","chopping":"841-895","consensus_level":"medium","plddt":87.0867,"start":841,"end":895}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9H4I2","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9H4I2-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9H4I2-F1-predicted_aligned_error_v6.png","plddt_mean":59.22},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=ZHX3","jax_strain_url":"https://www.jax.org/strain/search?query=ZHX3"},"sequence":{"accession":"Q9H4I2","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9H4I2.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9H4I2/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9H4I2"}},"corpus_meta":[{"pmid":"12659632","id":"PMC_12659632","title":"Analysis of zinc-fingers and homeoboxes (ZHX)-1-interacting proteins: molecular cloning and characterization of a member of the ZHX family, ZHX3.","date":"2003","source":"The Biochemical journal","url":"https://pubmed.ncbi.nlm.nih.gov/12659632","citation_count":52,"is_preprint":false},{"pmid":"14659886","id":"PMC_14659886","title":"The mouse zinc-fingers and homeoboxes (ZHX) family; ZHX2 forms a heterodimer with ZHX3.","date":"2003","source":"Gene","url":"https://pubmed.ncbi.nlm.nih.gov/14659886","citation_count":39,"is_preprint":false},{"pmid":"33440047","id":"PMC_33440047","title":"ZHX3 promotes the progression of urothelial carcinoma of the bladder via repressing of RGS2 and is a novel substrate of TRIM21.","date":"2021","source":"Cancer science","url":"https://pubmed.ncbi.nlm.nih.gov/33440047","citation_count":28,"is_preprint":false},{"pmid":"19273305","id":"PMC_19273305","title":"ZHX2 and ZHX3 repress cancer markers in normal hepatocytes.","date":"2009","source":"Frontiers in bioscience (Landmark edition)","url":"https://pubmed.ncbi.nlm.nih.gov/19273305","citation_count":26,"is_preprint":false},{"pmid":"34883128","id":"PMC_34883128","title":"A novel polysaccharides-based bioflocculant produced by Bacillus subtilis ZHX3 and its application in the treatment of multiple pollutants.","date":"2021","source":"Chemosphere","url":"https://pubmed.ncbi.nlm.nih.gov/34883128","citation_count":11,"is_preprint":false},{"pmid":"35720472","id":"PMC_35720472","title":"Attenuated ZHX3 expression is predictive of poor outcome for liver cancer: Indication for personalized therapy.","date":"2022","source":"Oncology letters","url":"https://pubmed.ncbi.nlm.nih.gov/35720472","citation_count":4,"is_preprint":false},{"pmid":"27062826","id":"PMC_27062826","title":"[Effect of echinacoside-containing serum in promoting mesenchymal stem cell osteogenic differentiation and ZHX₃ expression in rats].","date":"2015","source":"Zhongguo Zhong yao za zhi = Zhongguo zhongyao zazhi = China journal of Chinese materia medica","url":"https://pubmed.ncbi.nlm.nih.gov/27062826","citation_count":1,"is_preprint":false},{"pmid":"42231399","id":"PMC_42231399","title":"lncRNA-operated stem cell fate control: RPS3 mediates the in situ Gm16751-Zhx3 interactome to program spermatogonial stem cell differentiation.","date":"2026","source":"Cell & bioscience","url":"https://pubmed.ncbi.nlm.nih.gov/42231399","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.10.17.683150","title":"Transcriptomic and protein analysis of human cortex reveals genes and pathways linked to NPTX2 disruption in Alzheimer’s disease","date":"2025-10-20","source":"bioRxiv","url":"https://doi.org/10.1101/2025.10.17.683150","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.02.28.25323104","title":"Intragenic deletions from whole genome sequencing of 1054 suicide deaths","date":"2025-03-06","source":"bioRxiv","url":"https://doi.org/10.1101/2025.02.28.25323104","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":7538,"output_tokens":1107,"usd":0.019609,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":7653,"output_tokens":1857,"usd":0.042345,"stage2_stop_reason":"end_turn"},"total_usd":0.061954,"stage1_batch_id":"msgbatch_01UT1AahFPCAyn2uVweS7A9z","stage2_batch_id":"msgbatch_01HuKWtKNT4Pqj1bc8eEP5RV","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2003,\n      \"finding\": \"ZHX3 (956 amino acids, two zinc-finger motifs and five homeodomains) was identified as a ZHX1-interacting protein via yeast two-hybrid. ZHX3 dimerizes with both ZHX1 and ZHX3 itself (homodimerization), interacts with the activation domain of NF-YA, and functions as a ubiquitous transcriptional repressor localized in the nucleus. Domain mapping showed the dimerization domain, NF-YA interaction domain, and repressor domain all map to a region including the HD1 region; two nuclear localization signals were mapped to the N-terminal through Znf1 and the HD2 region.\",\n      \"method\": \"Yeast two-hybrid screen, molecular cloning, in vitro and in vivo protein-protein interaction assays, nuclear localization experiments, domain-mapping/deletion analysis\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (yeast two-hybrid, in vitro/in vivo interaction assays, domain mapping) in a single rigorous founding study\",\n      \"pmids\": [\"12659632\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"ZHX2 forms a heterodimer with ZHX3 via a region containing each HD1 domain, demonstrated by in vitro and in vivo protein-protein interaction assays using mouse ZHX2 and ZHX3.\",\n      \"method\": \"In vitro and in vivo protein-protein interaction assays (heterodimerization analysis), molecular cloning of mouse ZHX2 and ZHX3 cDNAs\",\n      \"journal\": \"Gene\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal in vitro and in vivo interaction assays with domain mapping, replicates and extends findings from PMID:12659632\",\n      \"pmids\": [\"14659886\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"ZHX3 represses transcription from the promoters of cancer marker genes PKM and HKII in hepatocytes, as demonstrated by reporter gene (luciferase) analysis using the promoter regions of these genes in hepatoma cells.\",\n      \"method\": \"Reporter gene (luciferase) assay with ZHX3 expression in hepatoma cell lines\",\n      \"journal\": \"Frontiers in bioscience (Landmark edition)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — reporter assay is a direct functional readout, but single lab, single method\",\n      \"pmids\": [\"19273305\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"ZHX3 is ubiquitinated and subsequently degraded by the E3 ubiquitin ligase TRIM21, identifying TRIM21 as the writer of ZHX3 ubiquitination. Additionally, ZHX3 transcriptionally represses expression of RGS2 (regulator of G protein signaling 2), promoting migration and invasion of urothelial carcinoma of the bladder cells.\",\n      \"method\": \"Co-immunoprecipitation (coIP), mass spectrometry (MS) for TRIM21 interaction, RNA-seq for RGS2 repression, in vitro and in vivo migration/invasion assays\",\n      \"journal\": \"Cancer science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — coIP plus MS for binding partner identification, RNA-seq for downstream target, functional assays; single lab but multiple orthogonal methods\",\n      \"pmids\": [\"33440047\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ZHX3 is a nuclear transcriptional repressor that homodimerizes and heterodimerizes with ZHX family members (ZHX1, ZHX2, ZHX3) via their HD1 domains, interacts with the NF-YA activation domain, represses target gene promoters (including PKM, HKII, and RGS2), and is itself regulated by TRIM21-mediated ubiquitination and proteasomal degradation.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"ZHX3 is a ubiquitously expressed nuclear transcriptional repressor that controls target gene expression through dimerization within the ZHX family and engagement of the transcription factor NF-Y [#0]. It contains two zinc-finger motifs and five homeodomains; its HD1 region carries the overlapping dimerization, NF-YA-interaction, and repressor activities, while nuclear localization signals map to the N-terminal/Znf1 and HD2 regions [#0]. ZHX3 homodimerizes and heterodimerizes with other ZHX family members, including ZHX1 [#0] and ZHX2 [#1], and binds the activation domain of NF-YA to repress transcription [#0]. Functionally, ZHX3 represses the promoters of the metabolic/cancer marker genes PKM and HKII in hepatocytes [#2] and represses RGS2 to promote migration and invasion of urothelial bladder carcinoma cells [#3]. ZHX3 protein levels are controlled by the E3 ubiquitin ligase TRIM21, which ubiquitinates ZHX3 and targets it for degradation [#3].\",\n  \"teleology\": [\n    {\n      \"year\": 2003,\n      \"claim\": \"Established ZHX3's core identity by showing it is a nuclear repressor that dimerizes and contacts the general transcription factor NF-Y, defining the molecular surfaces through which it acts.\",\n      \"evidence\": \"Yeast two-hybrid screen, in vitro/in vivo interaction assays, and domain-mapping/deletion analysis of cloned ZHX3\",\n      \"pmids\": [\"12659632\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"No direct DNA-binding site or target promoter identified at this stage\",\n        \"Repression mechanism downstream of NF-YA binding not defined\",\n        \"No endogenous target genes characterized\"\n      ]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Extended the ZHX dimerization network by demonstrating ZHX3 heterodimerizes with ZHX2 through the HD1 domain, establishing combinatorial partnering within the family.\",\n      \"evidence\": \"Reciprocal in vitro and in vivo interaction assays with mouse ZHX2 and ZHX3 cDNAs\",\n      \"pmids\": [\"14659886\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Functional consequence of ZHX2-ZHX3 heterodimers on transcription not measured\",\n        \"Relative affinities and preference among homo- vs heterodimers unknown\"\n      ]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Connected ZHX3 to specific gene targets by showing it represses the PKM and HKII promoters, providing the first defined endogenous-relevant transcriptional outputs.\",\n      \"evidence\": \"Luciferase reporter assays with PKM and HKII promoter regions in hepatoma cell lines\",\n      \"pmids\": [\"19273305\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Reporter assay only; direct binding to these promoters not shown\",\n        \"Single lab, single method\",\n        \"Effect on endogenous PKM/HKII expression and metabolism not established\"\n      ]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Defined how ZHX3 abundance is controlled and linked its repressor activity to cancer cell behavior by identifying TRIM21 as the ubiquitin ligase degrading ZHX3 and RGS2 as a repressed target driving invasion.\",\n      \"evidence\": \"CoIP plus mass spectrometry for TRIM21, RNA-seq for RGS2 repression, and in vitro/in vivo migration and invasion assays in bladder urothelial carcinoma cells\",\n      \"pmids\": [\"33440047\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Direct ZHX3 occupancy of the RGS2 promoter not demonstrated\",\n        \"TRIM21 ubiquitination site(s) on ZHX3 and signals regulating this degradation unknown\",\n        \"Single lab; reciprocal validation of binding partners limited\"\n      ]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How ZHX3 selects its DNA targets and whether its repression depends on chromatin-modifying cofactors beyond NF-Y interaction remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"No defined ZHX3 DNA-binding consensus despite zinc fingers and homeodomains\",\n        \"Co-repressor machinery executing repression not identified\",\n        \"Genome-wide direct binding map (e.g. ChIP-seq) absent\"\n      ]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [0, 2, 3]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [0, 2, 3]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"ZHX1\", \"ZHX2\", \"NFYA\", \"TRIM21\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}