{"gene":"TRIM35","run_date":"2026-06-10T10:51:56","timeline":{"discoveries":[{"year":2014,"finding":"TRIM35 physically interacts with pyruvate kinase isoform M2 (PKM2) via its coiled-coil domain, suppresses PKM2 tyrosine-105 (Y105) phosphorylation, and thereby reduces the Warburg effect and tumorigenicity in hepatocellular carcinoma cells.","method":"Mass spectrometry, Co-IP, domain deletion mutants, PKM2 Y105 phosphorylation assay, HCC cell proliferation and xenograft assays","journal":"Oncogene","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP with domain mapping, phosphorylation assay, and in vivo xenograft validation; replicated in principle across multiple subsequent studies","pmids":["25263439"],"is_preprint":false},{"year":2015,"finding":"TRIM35 negatively regulates TLR7/9-mediated type I interferon production by interacting with IRF7, promoting its K48-linked polyubiquitination, and inducing its proteasome-dependent degradation, acting as a negative feedback regulator.","method":"Co-IP, ubiquitination assay (K48-linkage), proteasome inhibitor rescue, TLR7/9 stimulation assays, overexpression/knockdown","journal":"FEBS letters","confidence":"High","confidence_rationale":"Tier 2 / Moderate — Co-IP, specific ubiquitin-linkage typing, proteasome-dependent degradation confirmed; single lab but multiple orthogonal methods","pmids":["25907537"],"is_preprint":false},{"year":2022,"finding":"TRIM35 ubiquitinates PKM2 and regulates the transition between its tetrameric (active, high-affinity) and dimeric (less active) forms, thereby suppressing the Warburg effect in breast cancer cells.","method":"Ubiquitination assays, co-IP, PKM2 tetramer/dimer gel assays, cell proliferation/migration/invasion assays, xenograft","journal":"International journal of oncology","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — single lab, ubiquitination and tetramer/dimer transition assays performed; mechanism partially characterized","pmids":["36196894"],"is_preprint":false},{"year":2022,"finding":"TRIM35, acting as an E3 ubiquitin ligase, promotes K48-linked ubiquitination and proteasomal degradation of nuclear PKM2 (S37P-PKM2) in cardiomyocytes; cardiomyocyte-specific TRIM35 overexpression in mice reduces nuclear PKM2 and GATA4/6 levels while increasing P53, producing dilated cardiomyopathy.","method":"Cardiomyocyte-specific TRIM35 transgenic mice, PKM2 cardiac-specific knockout mice, ubiquitination assays, Co-IP, cardiac echocardiography, IHC, patient LV samples","journal":"Science translational medicine","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo cardiac-specific overexpression and PKM2 KO mouse models, mechanistic Co-IP and ubiquitination assays, translational patient data; multiple orthogonal methods","pmids":["36322626"],"is_preprint":false},{"year":2021,"finding":"TRIM35 functions as an E3 ubiquitin ligase to mediate the ubiquitination and degradation of CLOCK, a key regulator of circadian rhythmicity, in diffuse large B-cell lymphoma (DLBCL) cells, suppressing DLBCL proliferation and modulating NK cell infiltration.","method":"Overexpression in DLBCL cell lines, ubiquitination assay, co-IP, NK cell co-culture/infiltration assays, tumor xenograft","journal":"Journal of immunology research","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — single lab, ubiquitination and degradation of CLOCK demonstrated by Co-IP and overexpression; limited mechanistic depth on NK cell link","pmids":["34124276"],"is_preprint":false},{"year":2022,"finding":"TRIM35 interacts with TIGAR and promotes its polyubiquitination and proteasomal degradation; TRIM35 knockdown alleviates renal ischemia-reperfusion injury by reducing oxidative stress and enhancing mitochondrial fusion via stabilization of TIGAR.","method":"Co-IP, ubiquitination assay, TRIM35 knockdown (siRNA), mitochondrial morphology analysis, oxidative stress assays, renal IRI mouse model","journal":"International journal of biological macromolecules","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — single lab, Co-IP and ubiquitination assays performed, in vivo mouse IRI model; mechanistic link to mitochondrial fusion via TIGAR established","pmids":["35421414"],"is_preprint":false},{"year":2022,"finding":"TRIM35 promotes breast cancer cell apoptosis by ubiquitinating and degrading PDK1, leading to inactivation of AKT signaling.","method":"Ubiquitination assay, co-IP, overexpression/knockdown, AKT pathway western blot, xenograft mouse model","journal":"Neoplasma","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — single lab, Co-IP and ubiquitination of PDK1 demonstrated with downstream AKT readout; xenograft validation","pmids":["35081724"],"is_preprint":false},{"year":2023,"finding":"TRIM35 (E3 ligase) mediates K63-linked ubiquitination of LSD1 at lysine-422, repressing LSD1 demethylase activity; suppressed LSD1 activity facilitates ERGIC1 transcription, autophagy inhibition, and IFNGR1 stabilization, activating IFN-γ signaling and increasing MHC class I expression in NSCLC cells.","method":"Co-IP, K63-specific ubiquitination assay, LSD1 demethylase activity assay, site-directed mutagenesis (K422), gene expression analysis, NSCLC cell and mouse models","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — site-directed mutagenesis of ubiquitination site, enzymatic (demethylase) activity assay, downstream pathway validation; single lab with multiple orthogonal methods","pmids":["37979167"],"is_preprint":false},{"year":2024,"finding":"TRIM35 directly monoubiquitinates lysine-120 (K120) on histone H2B in postnatal cardiomyocytes, promoting chromatin remodeling and accessibility of P53 to its transcriptional promoter targets, thereby increasing P53 transcriptional output and contributing to dilated cardiomyopathy.","method":"ChIP-seq, RNA-seq, overexpression transgenic mice (cardiomyocyte-specific), siRNA knockdown in primary cardiomyocytes, adenovirus-mediated gene delivery, patient DCM LV samples","journal":"Circulation research","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — ChIP-seq, direct H2B K120 monoubiquitination, in vivo cardiomyocyte-specific transgenic model, translational human DCM data; multiple orthogonal methods in single study","pmids":["38860363"],"is_preprint":false},{"year":2024,"finding":"TRIM35 interacts with and ubiquitinates SLC7A5 (an amino acid transporter), increasing its stability and enhancing amino acid transport, which activates mTORC1 signaling in cardiac fibroblasts to promote fibroblast proliferation, migration, and differentiation, driving cardiac remodeling.","method":"LC-MS/MS substrate identification, Co-IP, ubiquitination assay, fibroblast-specific Trim35 conditional knockout mice (TAC surgery), adenoviral/AAV overexpression, mTORC1 pathway assays, RNA-seq","journal":"Cell communication and signaling","confidence":"High","confidence_rationale":"Tier 2 / Strong — fibroblast-specific conditional KO mice, LC-MS/MS substrate discovery, Co-IP and ubiquitination of SLC7A5, mTORC1 rescue experiments; multiple orthogonal methods","pmids":["39304904"],"is_preprint":false},{"year":2023,"finding":"TRIM35 interacts with adenovirus E1A protein and promotes its K48-linked ubiquitination at lysine residues K253 and K285, leading to E1A degradation and inhibition of HAdV replication.","method":"Co-IP, K48-linked ubiquitination assay, site-directed mutagenesis of E1A (K253A, K285A), viral replication assays, shTRIM35 oncolytic adenovirus construct","journal":"Journal of virology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, K48-specific ubiquitination, mutagenesis of substrate sites; single lab with two orthogonal methods","pmids":["37578239"],"is_preprint":false},{"year":2024,"finding":"TRIM35 negatively regulates the cGAS-STING antiviral signaling pathway by directly interacting with STING and removing K63-linked ubiquitin from STING via its RING domain (C36 and C44 sites acting as deubiquitinase-like activity), impairing STING interaction with TBK1/IKKε and attenuating IFN-β production.","method":"Co-IP, co-localization (cytoplasm), K63-ubiquitin assay, RING domain mutagenesis (C36A, C44A), TBK1/IKKε interaction assays, IFN-β reporter assay","journal":"Inflammation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, site-directed RING domain mutagenesis, specific K63-deubiquitination of STING, downstream TBK1 assay; single lab, multiple methods","pmids":["39088122"],"is_preprint":false},{"year":2021,"finding":"Porcine TRIM35 directly interacts with TRAF3 and catalyzes K63-linked polyubiquitination of TRAF3, leading to upregulation of IFN-β production; RING and PRY/SPRY domains are essential for E3 ligase activity.","method":"Co-IP, K63-linked ubiquitination assay, domain deletion mutants (RING, PRY/SPRY), IFN-β luciferase reporter, overexpression/knockdown in ST cells, JEV infection model","journal":"Developmental and comparative immunology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, K63-specific ubiquitination, domain requirement mapping; ortholog (porcine) data with domain architecture consistent with human gene; single lab","pmids":["34626690"],"is_preprint":false},{"year":2023,"finding":"TRIM35 interacts with IRF5 and promotes its ubiquitination and degradation; loss of TRIM35 leads to increased IRF5 levels and enhanced LDHA expression and glycolysis in hepatocellular carcinoma cells.","method":"Co-immunoprecipitation, ubiquitination assay, mass spectrometry metabolite analysis, western blot, qPCR, HCC cell functional assays","journal":"Journal of digestive diseases","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — Co-IP and ubiquitination assay support TRIM35-IRF5 interaction and degradation; single lab, limited mechanistic depth","pmids":["37594849"],"is_preprint":false},{"year":2007,"finding":"Hls5 (mouse ortholog of TRIM35/HLS5) relocalizes from cytoplasmic granules to the nucleus under GATA-1 influence, associates with both FOG-1 and GATA-1, and suppresses GATA-1-mediated transactivation and GATA-1 DNA binding, thereby impeding erythroid maturation.","method":"Co-IP (Hls5/FOG-1/GATA-1), subcellular localization (fluorescence microscopy), GATA-1 transactivation reporter assay, GATA-1 DNA binding assay (ChIP/EMSA), erythroid differentiation assays","journal":"Blood","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, transcriptional reporter, DNA binding assay; mouse ortholog, multiple orthogonal methods in single study","pmids":["18063753"],"is_preprint":false},{"year":2003,"finding":"HLS5 (human TRIM35), an RBCC family protein, localizes to cytoplasmic granules and punctate nuclear bodies; enforced expression in HeLa cells inhibits cell growth, clonogenicity, and tumorigenicity, establishing it as a candidate tumor suppressor at chromosome 8p21.","method":"cDNA cloning, subcellular localization (immunofluorescence), HeLa cell overexpression, growth inhibition and colony-formation assays, xenograft tumorigenicity assays","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — direct localization imaging, functional tumor suppression in cell and xenograft assays; foundational study, multiple readouts","pmids":["14662771"],"is_preprint":false},{"year":2025,"finding":"TRIM35 directly binds genomic promoters (DNA-binding activity), interacts with histone H3, and catalyzes non-proteolytic (monoubiquitination) ubiquitination of H3, serving as a recruitment signal for p300 acetyltransferase, leading to H3K27 acetylation and transcriptional activation of HSPA6, which suppresses breast cancer progression.","method":"ChIP-PCR, Co-IP, ubiquitination assay (non-proteolytic), p300 recruitment assay, H3K27ac western blot, HSPA6 expression analysis, breast cancer cell functional assays","journal":"Cell death discovery","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — ChIP, Co-IP, ubiquitination and downstream H3K27ac assays; single lab, novel mechanism not yet replicated","pmids":["41136372"],"is_preprint":false},{"year":2025,"finding":"Endothelial TRIM35 inhibits MMP10 expression and secretion by promoting K63-linked ubiquitination of RelB and maintaining its nuclear localization, thereby suppressing noncanonical NF-κB-driven MMP10 transcription; TRIM35 endothelial-specific conditional knockout leads to increased MMP10 release and promotes vascular graft calcification via SMC paracrine signaling.","method":"Endothelial-specific TRIM35 cKO mice (arterial isograft model), single-cell RNA-seq, Co-IP, K63-ubiquitination assay of RelB, RelB nuclear localization assay, MMP10 ELISA, in vitro SMC calcification co-culture","journal":"Advanced science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — conditional endothelial KO in vivo, K63-ubiquitination of RelB, RelB nuclear localization, MMP10 functional rescue; single lab, multiple orthogonal methods","pmids":["39865905"],"is_preprint":false},{"year":2025,"finding":"TRIM35 ubiquitinates EIF3E and promotes its degradation, modulating the CDK4/Cyclin D1 signaling pathway to suppress endometrial cancer cell proliferation.","method":"Co-IP, ubiquitination assay, EIF3E overexpression rescue, CDK4/Cyclin D1 western blot, cell proliferation and invasion assays","journal":"Cellular signalling","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, Co-IP and ubiquitination assay; limited mechanistic follow-up; no independent replication","pmids":["41429342"],"is_preprint":false}],"current_model":"TRIM35 is a RING-domain E3 ubiquitin ligase (and in some contexts a deubiquitinase/ubiquitin editor) that suppresses tumor growth and regulates innate immunity and cardiac homeostasis through substrate-specific ubiquitination: it interacts with and ubiquitinates PKM2 (blocking Y105 phosphorylation and the Warburg effect), IRF7 (K48-linked degradation to dampen TLR7/9-driven IFN production), LSD1 (K63-linked ubiquitination inhibiting demethylase activity to enhance tumor immunogenicity), nuclear histone H2B (K120 monoubiquitination promoting P53 transcriptional access in cardiomyocytes), STING (removal of K63-linked ubiquitin attenuating cGAS-STING signaling), SLC7A5 (stabilizing the amino acid transporter to activate mTORC1 in cardiac fibroblasts), PDK1, TIGAR, CLOCK, EIF3E, viral E1A, and RelB, while also localizing to the nucleus under GATA-1 direction to co-repress GATA-1 transcriptional activity during erythroid differentiation."},"narrative":{"mechanistic_narrative":"TRIM35 (HLS5) is a RING-domain TRIM-family E3 ubiquitin ligase that functions as a tumor suppressor and a substrate-specific regulator of cellular metabolism, innate immune signaling, and cardiovascular homeostasis, originally identified as a chromosome 8p21 candidate tumor suppressor whose enforced expression inhibits growth, clonogenicity, and tumorigenicity [PMID:14662771]. A core oncometabolic activity is its control of pyruvate kinase M2: TRIM35 binds PKM2 through its coiled-coil domain, suppresses PKM2 Y105 phosphorylation, and shifts PKM2 toward its tetrameric state, thereby blunting the Warburg effect and tumorigenicity [PMID:25263439, PMID:36196894]. Through substrate ubiquitination TRIM35 restrains multiple pro-tumor and pro-glycolytic factors, including K48-linked degradation of PDK1 to inactivate AKT signaling [PMID:35081724], degradation of EIF3E to modulate CDK4/Cyclin D1 [PMID:41429342], and K63-linked ubiquitination of LSD1 at K422 that represses its demethylase activity to enhance IFN-γ signaling and MHC class I expression and tumor immunogenicity [PMID:37979167]. TRIM35 also acts directly on chromatin: it binds promoters, monoubiquitinates histone H2B at K120 and histone H3, and recruits p300, thereby remodeling chromatin to control P53 and HSPA6 transcriptional programs [PMID:38860363, PMID:41136372]. In innate immunity TRIM35 is a bidirectional regulator, promoting K48-linked degradation of IRF7 to dampen TLR7/9-driven type I interferon as a negative feedback loop [PMID:25907537] and removing K63-linked ubiquitin from STING to attenuate cGAS-STING signaling [PMID:39088122]. In the cardiovascular system, cardiomyocyte TRIM35 drives dilated cardiomyopathy by degrading nuclear PKM2 and enhancing P53 output [PMID:36322626, PMID:38860363], while it stabilizes SLC7A5 to activate mTORC1 in cardiac fibroblasts [PMID:39304904] and ubiquitinates RelB in endothelium to restrain MMP10-driven vascular calcification [PMID:39865905].","teleology":[{"year":2003,"claim":"Established TRIM35/HLS5 as a candidate tumor suppressor, answering whether this 8p21 RBCC protein has growth-regulatory function before any catalytic mechanism was known.","evidence":"cDNA cloning, immunofluorescence localization, and growth/clonogenicity/xenograft assays in HeLa cells","pmids":["14662771"],"confidence":"Medium","gaps":["No molecular substrate or enzymatic activity defined","Mechanism of growth suppression unknown"]},{"year":2007,"claim":"Linked TRIM35 to transcriptional control by showing its ortholog co-represses GATA-1, addressing how it might influence differentiation programs.","evidence":"Co-IP, subcellular localization, GATA-1 reporter and DNA-binding assays in erythroid differentiation models (mouse ortholog Hls5)","pmids":["18063753"],"confidence":"Medium","gaps":["Repression mechanism not tied to ubiquitin ligase activity","Mouse ortholog data only"]},{"year":2014,"claim":"Identified PKM2 as a TRIM35 partner and defined its anti-Warburg activity, revealing how the tumor suppressor reprograms cancer metabolism.","evidence":"Mass spectrometry, reciprocal Co-IP with domain mapping, PKM2 Y105 phosphorylation assay, HCC proliferation and xenograft assays","pmids":["25263439"],"confidence":"High","gaps":["Whether PKM2 regulation requires ubiquitination not resolved here","Coiled-coil binding mapped but ligase mechanism on PKM2 unclear"]},{"year":2015,"claim":"Defined TRIM35 as a negative feedback regulator of innate immunity by targeting IRF7 for degradation, establishing its E3 ligase role in interferon control.","evidence":"Co-IP, K48-linkage ubiquitination assay, proteasome inhibitor rescue, TLR7/9 stimulation with overexpression/knockdown","pmids":["25907537"],"confidence":"High","gaps":["Ubiquitination site on IRF7 not mapped","Single lab"]},{"year":2021,"claim":"Extended TRIM35 substrate scope to CLOCK and TRAF3, showing context-dependent degradative versus activating ubiquitination in lymphoma and antiviral immunity.","evidence":"Co-IP, ubiquitination assays, IFN-β reporter, NK co-culture, JEV infection model (porcine TRIM35 for TRAF3)","pmids":["34124276","34626690"],"confidence":"Medium","gaps":["TRAF3 data is from porcine ortholog","Mechanistic depth on NK link and ubiquitin linkage outcomes limited"]},{"year":2022,"claim":"Demonstrated TRIM35's role in cardiac and renal pathophysiology and broadened its oncometabolic substrate set, showing the same ligase activity drives disparate disease phenotypes.","evidence":"Cardiomyocyte-specific transgenic and PKM2-KO mice, renal IRI model, Co-IP and ubiquitination assays for PKM2, TIGAR, PDK1; breast cancer PKM2 tetramer assays","pmids":["36322626","35421414","35081724","36196894"],"confidence":"High","gaps":["TIGAR, PDK1 findings are single-lab Medium evidence","Tissue-specific determinants of substrate selection unclear"]},{"year":2023,"claim":"Revealed non-degradative K63 ubiquitination as a TRIM35 mechanism via LSD1, linking the ligase to chromatin/immunogenicity, and expanded substrates to E1A and IRF5.","evidence":"Co-IP, K63-specific and site-directed ubiquitination assays (LSD1 K422; E1A K253/K285), demethylase activity assay, viral replication and HCC metabolite assays","pmids":["37979167","37578239","37594849"],"confidence":"High","gaps":["IRF5 link is single-lab Weak evidence","How TRIM35 selects K48 versus K63 linkages not established"]},{"year":2024,"claim":"Defined direct histone modification and transporter stabilization activities, showing TRIM35 acts on chromatin (H2B K120 monoubiquitination) and on SLC7A5 to drive cardiac remodeling, and removes K63-Ub from STING.","evidence":"ChIP-seq, cardiomyocyte- and fibroblast-specific transgenic/cKO mice, LC-MS/MS substrate ID, Co-IP, ubiquitination assays, STING RING-domain mutagenesis (C36/C44)","pmids":["38860363","39304904","39088122"],"confidence":"High","gaps":["STING deubiquitinase-like activity is Medium single-lab evidence","How a RING ligase performs ubiquitin removal mechanistically unresolved"]},{"year":2025,"claim":"Established TRIM35 as a chromatin-binding transcriptional activator (H3 monoubiquitination, p300 recruitment) and a vascular regulator via RelB, broadening its mechanistic repertoire beyond degradative ubiquitination.","evidence":"ChIP-PCR, endothelial-specific cKO mice, scRNA-seq, Co-IP, K63-ubiquitination of RelB, H3K27ac and MMP10 functional assays","pmids":["41136372","39865905","41429342"],"confidence":"Medium","gaps":["Direct DNA-binding mode and sequence specificity uncharacterized","EIF3E finding is Low-confidence single lab","Non-proteolytic H3 ubiquitination mechanism novel and unreplicated"]},{"year":null,"claim":"It remains unresolved how a single RING-domain ligase achieves its broad substrate range and switches between K48-degradative, K63-non-degradative, monoubiquitinating, and ubiquitin-editing activities in a tissue- and context-specific manner.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structural model of substrate recognition","Determinants of linkage-type and proteolytic versus non-proteolytic outcomes unknown","Direct DNA-binding specificity undefined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016740","term_label":"transferase activity","supporting_discovery_ids":[1,3,6,7,8,9,10,12,16,17]},{"term_id":"GO:0016874","term_label":"ligase activity","supporting_discovery_ids":[1,3,7,8,9]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[1,3,6,9,10,18]},{"term_id":"GO:0003677","term_label":"DNA binding","supporting_discovery_ids":[16]},{"term_id":"GO:0042393","term_label":"histone binding","supporting_discovery_ids":[8,16]},{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[8,14,16]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[3,8,14,15,16]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[11,14,15]}],"pathway":[{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[0,2,3,13]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[1,7,11,12]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[8,14,16,17]},{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[1,3,6,9,18]},{"term_id":"R-HSA-9612973","term_label":"Autophagy","supporting_discovery_ids":[7]}],"complexes":[],"partners":["PKM2","IRF7","LSD1","SLC7A5","STING1","RELB","TIGAR","PDK1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9UPQ4","full_name":"E3 ubiquitin-protein ligase TRIM35","aliases":["Hemopoietic lineage switch protein 5"],"length_aa":493,"mass_kda":56.5,"function":"E3 ubiquitin-protein ligase that participates in multiple biological processes including cell death, glucose metabolism, and in particular, the innate immune response. Mediates 'Lys-63'-linked polyubiquitination of TRAF3 thereby promoting type I interferon production via RIG-I signaling pathway (PubMed:32562145). Can also catalyze 'Lys-48'-linked polyubiquitination and proteasomal degradation of viral proteins such as influenza virus PB2 (PubMed:32562145). Acts as a negative feedback regulator of TLR7- and TLR9-triggered signaling. Mechanistically, promotes the 'Lys-48'-linked ubiquitination of IRF7 and induces its degradation via a proteasome-dependent pathway (PubMed:25907537). Reduces FGFR1-dependent tyrosine phosphorylation of PKM, inhibiting PKM-dependent lactate production, glucose metabolism, and cell growth (PubMed:25263439)","subcellular_location":"Cytoplasm; Nucleus","url":"https://www.uniprot.org/uniprotkb/Q9UPQ4/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/TRIM35","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/TRIM35","total_profiled":1310},"omim":[{"mim_id":"617007","title":"TRIPARTITE MOTIF-CONTAINING PROTEIN 35; TRIM35","url":"https://www.omim.org/entry/617007"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Nucleoplasm","reliability":"Approved"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/TRIM35"},"hgnc":{"alias_symbol":["KIAA1098","MAIR","HLS5"],"prev_symbol":[]},"alphafold":{"accession":"Q9UPQ4","domains":[{"cath_id":"3.30.40.10","chopping":"15-90","consensus_level":"medium","plddt":88.5426,"start":15,"end":90},{"cath_id":"-","chopping":"183-297","consensus_level":"high","plddt":95.3306,"start":183,"end":297},{"cath_id":"2.60.120.920","chopping":"305-375_390-479","consensus_level":"high","plddt":89.0218,"start":305,"end":479}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9UPQ4","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9UPQ4-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9UPQ4-F1-predicted_aligned_error_v6.png","plddt_mean":88.25},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=TRIM35","jax_strain_url":"https://www.jax.org/strain/search?query=TRIM35"},"sequence":{"accession":"Q9UPQ4","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9UPQ4.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9UPQ4/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9UPQ4"}},"corpus_meta":[{"pmid":"12874256","id":"PMC_12874256","title":"Paired activating and inhibitory immunoglobulin-like receptors, MAIR-I and MAIR-II, regulate mast cell and macrophage activation.","date":"2003","source":"The Journal of experimental medicine","url":"https://pubmed.ncbi.nlm.nih.gov/12874256","citation_count":85,"is_preprint":false},{"pmid":"25263439","id":"PMC_25263439","title":"TRIM35 Interacts with pyruvate kinase isoform M2 to suppress the Warburg effect and tumorigenicity in hepatocellular carcinoma.","date":"2014","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/25263439","citation_count":78,"is_preprint":false},{"pmid":"25576919","id":"PMC_25576919","title":"Co-expression of PKM2 and TRIM35 predicts survival and recurrence in hepatocellular carcinoma.","date":"2015","source":"Oncotarget","url":"https://pubmed.ncbi.nlm.nih.gov/25576919","citation_count":66,"is_preprint":false},{"pmid":"28394882","id":"PMC_28394882","title":"miR-4417 Targets Tripartite Motif-Containing 35 (TRIM35) and Regulates Pyruvate Kinase Muscle 2 (PKM2) Phosphorylation to Promote Proliferation and Suppress Apoptosis in Hepatocellular Carcinoma Cells.","date":"2017","source":"Medical science monitor : international medical journal of experimental and clinical research","url":"https://pubmed.ncbi.nlm.nih.gov/28394882","citation_count":35,"is_preprint":false},{"pmid":"25907537","id":"PMC_25907537","title":"TRIM35 negatively regulates TLR7- and TLR9-mediated type I interferon production by targeting IRF7.","date":"2015","source":"FEBS letters","url":"https://pubmed.ncbi.nlm.nih.gov/25907537","citation_count":32,"is_preprint":false},{"pmid":"36196894","id":"PMC_36196894","title":"TRIM35 ubiquitination regulates the expression of PKM2 tetramer and dimer and affects the malignant behaviour of breast cancer by regulating the Warburg effect.","date":"2022","source":"International journal of oncology","url":"https://pubmed.ncbi.nlm.nih.gov/36196894","citation_count":28,"is_preprint":false},{"pmid":"14662771","id":"PMC_14662771","title":"HLS5, a novel RBCC (ring finger, B box, coiled-coil) family member isolated from a hemopoietic lineage switch, is a candidate tumor suppressor.","date":"2003","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/14662771","citation_count":28,"is_preprint":false},{"pmid":"18097021","id":"PMC_18097021","title":"Caspase-independent cell death by CD300LF (MAIR-V), an inhibitory immunoglobulin-like receptor on myeloid cells.","date":"2008","source":"Journal of immunology (Baltimore, Md. : 1950)","url":"https://pubmed.ncbi.nlm.nih.gov/18097021","citation_count":28,"is_preprint":false},{"pmid":"36322626","id":"PMC_36322626","title":"TRIM35-mediated degradation of nuclear PKM2 destabilizes GATA4/6 and induces P53 in cardiomyocytes to promote heart failure.","date":"2022","source":"Science translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/36322626","citation_count":26,"is_preprint":false},{"pmid":"17202337","id":"PMC_17202337","title":"Dual assemblies of an 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biological macromolecules","url":"https://pubmed.ncbi.nlm.nih.gov/35421414","citation_count":13,"is_preprint":false},{"pmid":"39304904","id":"PMC_39304904","title":"TRIM35 triggers cardiac remodeling by regulating SLC7A5-mediated amino acid transport and mTORC1 activation in fibroblasts.","date":"2024","source":"Cell communication and signaling : CCS","url":"https://pubmed.ncbi.nlm.nih.gov/39304904","citation_count":12,"is_preprint":false},{"pmid":"38860363","id":"PMC_38860363","title":"TRIM35 Monoubiquitinates H2B in Cardiac Cells, Implications for Heart Failure.","date":"2024","source":"Circulation research","url":"https://pubmed.ncbi.nlm.nih.gov/38860363","citation_count":12,"is_preprint":false},{"pmid":"37594849","id":"PMC_37594849","title":"IRF5 promotes glycolysis in the progression of hepatocellular carcinoma and is regulated by TRIM35.","date":"2023","source":"Journal of digestive diseases","url":"https://pubmed.ncbi.nlm.nih.gov/37594849","citation_count":12,"is_preprint":false},{"pmid":"34626690","id":"PMC_34626690","title":"Porcine TRIM35 positively regulate TRAF3-mediated IFN-β production and inhibit Japanese encephalitis virus replication.","date":"2021","source":"Developmental and comparative immunology","url":"https://pubmed.ncbi.nlm.nih.gov/34626690","citation_count":9,"is_preprint":false},{"pmid":"33140535","id":"PMC_33140535","title":"MAIR-II deficiency ameliorates cardiac remodelling post-myocardial infarction by suppressing TLR9-mediated macrophage activation.","date":"2020","source":"Journal of cellular and molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/33140535","citation_count":8,"is_preprint":false},{"pmid":"18294079","id":"PMC_18294079","title":"Expression of a splicing isoform of MAIR-V (CD300LF), an inhibitory immunoglobulin-like receptor on myeloid cells.","date":"2008","source":"Hybridoma (2005)","url":"https://pubmed.ncbi.nlm.nih.gov/18294079","citation_count":6,"is_preprint":false},{"pmid":"37578239","id":"PMC_37578239","title":"Inhibition of human adenovirus replication by TRIM35-mediated degradation of E1A.","date":"2023","source":"Journal of virology","url":"https://pubmed.ncbi.nlm.nih.gov/37578239","citation_count":6,"is_preprint":false},{"pmid":"39954170","id":"PMC_39954170","title":"Downregulated TRIM35 Alleviates Doxorubicin-Induced Cardiotoxicity by Suppressing Oxidative Stress and Inflammation via Inhibiting TLR4/NF-κB Pathway.","date":"2025","source":"Cardiovascular drugs and therapy","url":"https://pubmed.ncbi.nlm.nih.gov/39954170","citation_count":4,"is_preprint":false},{"pmid":"18063753","id":"PMC_18063753","title":"Hls5 regulated erythroid differentiation by modulating GATA-1 activity.","date":"2007","source":"Blood","url":"https://pubmed.ncbi.nlm.nih.gov/18063753","citation_count":4,"is_preprint":false},{"pmid":"32955098","id":"PMC_32955098","title":"Tripartite motif-containing 35 (TRIM35) is up-regulated in UUO-induced renal fibrosis animal model.","date":"2020","source":"Histology and histopathology","url":"https://pubmed.ncbi.nlm.nih.gov/32955098","citation_count":3,"is_preprint":false},{"pmid":"36445078","id":"PMC_36445078","title":"Duck TRIM35 Promotes Tembusu Virus Replication by Interfering with RIG-I-Mediated Antiviral Signaling in Duck Embryo Fibroblasts.","date":"2022","source":"Microbiology spectrum","url":"https://pubmed.ncbi.nlm.nih.gov/36445078","citation_count":2,"is_preprint":false},{"pmid":"39865905","id":"PMC_39865905","title":"Endothelial TRIM35-Regulated MMP10 Release Exacerbates Calcification of Vascular Grafts.","date":"2025","source":"Advanced science (Weinheim, Baden-Wurttemberg, Germany)","url":"https://pubmed.ncbi.nlm.nih.gov/39865905","citation_count":1,"is_preprint":false},{"pmid":"41136372","id":"PMC_41136372","title":"TRIM35, a novel DNA-binding protein, epigenetically modifies H3 to promote HSPA6 transcription and suppress breast cancer progression.","date":"2025","source":"Cell death discovery","url":"https://pubmed.ncbi.nlm.nih.gov/41136372","citation_count":1,"is_preprint":false},{"pmid":"38642184","id":"PMC_38642184","title":"HBO regulates the Warburg effect of hypoxic HCC cells through miR-103a-3p/TRIM35.","date":"2024","source":"Discover oncology","url":"https://pubmed.ncbi.nlm.nih.gov/38642184","citation_count":1,"is_preprint":false},{"pmid":"28397257","id":"PMC_28397257","title":"Retracted: Mice lacking the Raf-1 kinase inhibitor protein exhibit exaggerated hypoxia-induced pulmonary hypertension, by I Morecroft, B Doyle, M Nilsen, W Kolch, K Mair and MR MacLean. British Journal of Pharmacology, volume 163(5): 948-963, published in June 2011; DOI 10.1111/j.1476-5381.2011.01305.x.","date":"2017","source":"British journal of pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/28397257","citation_count":1,"is_preprint":false},{"pmid":"39088122","id":"PMC_39088122","title":"TRIM35 Negatively Regulates the cGAS-STING-Mediated Signaling Pathway by Attenuating K63-Linked Ubiquitination of STING.","date":"2024","source":"Inflammation","url":"https://pubmed.ncbi.nlm.nih.gov/39088122","citation_count":0,"is_preprint":false},{"pmid":"41429342","id":"PMC_41429342","title":"TRIM35 inhibits endometrial cancer progression via ubiquitination and degradation of EIF3E.","date":"2025","source":"Cellular signalling","url":"https://pubmed.ncbi.nlm.nih.gov/41429342","citation_count":0,"is_preprint":false},{"pmid":"41351040","id":"PMC_41351040","title":"tsRNA-3040b accumulates R-loop to regulate Trim35 transcription, which leads to disordered glycolysis and promotes PAECs proliferation.","date":"2025","source":"Cellular & molecular biology letters","url":"https://pubmed.ncbi.nlm.nih.gov/41351040","citation_count":0,"is_preprint":false},{"pmid":"42180910","id":"PMC_42180910","title":"Knockdown of TRIM35 suppresses cell proliferation and metastasis by modulating the PPAR signalling pathway in papillary thyroid cancer cells.","date":"2026","source":"Translational cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/42180910","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":20866,"output_tokens":4946,"usd":0.068394,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":13145,"output_tokens":4048,"usd":0.083462,"stage2_stop_reason":"end_turn"},"total_usd":0.151856,"stage1_batch_id":"msgbatch_017qqVz33Ga63SCDwzb35XPb","stage2_batch_id":"msgbatch_01Scr2Ei9x4HJMiRGxwrmvLK","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2014,\n      \"finding\": \"TRIM35 physically interacts with pyruvate kinase isoform M2 (PKM2) via its coiled-coil domain, suppresses PKM2 tyrosine-105 (Y105) phosphorylation, and thereby reduces the Warburg effect and tumorigenicity in hepatocellular carcinoma cells.\",\n      \"method\": \"Mass spectrometry, Co-IP, domain deletion mutants, PKM2 Y105 phosphorylation assay, HCC cell proliferation and xenograft assays\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP with domain mapping, phosphorylation assay, and in vivo xenograft validation; replicated in principle across multiple subsequent studies\",\n      \"pmids\": [\"25263439\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"TRIM35 negatively regulates TLR7/9-mediated type I interferon production by interacting with IRF7, promoting its K48-linked polyubiquitination, and inducing its proteasome-dependent degradation, acting as a negative feedback regulator.\",\n      \"method\": \"Co-IP, ubiquitination assay (K48-linkage), proteasome inhibitor rescue, TLR7/9 stimulation assays, overexpression/knockdown\",\n      \"journal\": \"FEBS letters\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, specific ubiquitin-linkage typing, proteasome-dependent degradation confirmed; single lab but multiple orthogonal methods\",\n      \"pmids\": [\"25907537\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"TRIM35 ubiquitinates PKM2 and regulates the transition between its tetrameric (active, high-affinity) and dimeric (less active) forms, thereby suppressing the Warburg effect in breast cancer cells.\",\n      \"method\": \"Ubiquitination assays, co-IP, PKM2 tetramer/dimer gel assays, cell proliferation/migration/invasion assays, xenograft\",\n      \"journal\": \"International journal of oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — single lab, ubiquitination and tetramer/dimer transition assays performed; mechanism partially characterized\",\n      \"pmids\": [\"36196894\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"TRIM35, acting as an E3 ubiquitin ligase, promotes K48-linked ubiquitination and proteasomal degradation of nuclear PKM2 (S37P-PKM2) in cardiomyocytes; cardiomyocyte-specific TRIM35 overexpression in mice reduces nuclear PKM2 and GATA4/6 levels while increasing P53, producing dilated cardiomyopathy.\",\n      \"method\": \"Cardiomyocyte-specific TRIM35 transgenic mice, PKM2 cardiac-specific knockout mice, ubiquitination assays, Co-IP, cardiac echocardiography, IHC, patient LV samples\",\n      \"journal\": \"Science translational medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo cardiac-specific overexpression and PKM2 KO mouse models, mechanistic Co-IP and ubiquitination assays, translational patient data; multiple orthogonal methods\",\n      \"pmids\": [\"36322626\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"TRIM35 functions as an E3 ubiquitin ligase to mediate the ubiquitination and degradation of CLOCK, a key regulator of circadian rhythmicity, in diffuse large B-cell lymphoma (DLBCL) cells, suppressing DLBCL proliferation and modulating NK cell infiltration.\",\n      \"method\": \"Overexpression in DLBCL cell lines, ubiquitination assay, co-IP, NK cell co-culture/infiltration assays, tumor xenograft\",\n      \"journal\": \"Journal of immunology research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — single lab, ubiquitination and degradation of CLOCK demonstrated by Co-IP and overexpression; limited mechanistic depth on NK cell link\",\n      \"pmids\": [\"34124276\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"TRIM35 interacts with TIGAR and promotes its polyubiquitination and proteasomal degradation; TRIM35 knockdown alleviates renal ischemia-reperfusion injury by reducing oxidative stress and enhancing mitochondrial fusion via stabilization of TIGAR.\",\n      \"method\": \"Co-IP, ubiquitination assay, TRIM35 knockdown (siRNA), mitochondrial morphology analysis, oxidative stress assays, renal IRI mouse model\",\n      \"journal\": \"International journal of biological macromolecules\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — single lab, Co-IP and ubiquitination assays performed, in vivo mouse IRI model; mechanistic link to mitochondrial fusion via TIGAR established\",\n      \"pmids\": [\"35421414\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"TRIM35 promotes breast cancer cell apoptosis by ubiquitinating and degrading PDK1, leading to inactivation of AKT signaling.\",\n      \"method\": \"Ubiquitination assay, co-IP, overexpression/knockdown, AKT pathway western blot, xenograft mouse model\",\n      \"journal\": \"Neoplasma\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — single lab, Co-IP and ubiquitination of PDK1 demonstrated with downstream AKT readout; xenograft validation\",\n      \"pmids\": [\"35081724\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"TRIM35 (E3 ligase) mediates K63-linked ubiquitination of LSD1 at lysine-422, repressing LSD1 demethylase activity; suppressed LSD1 activity facilitates ERGIC1 transcription, autophagy inhibition, and IFNGR1 stabilization, activating IFN-γ signaling and increasing MHC class I expression in NSCLC cells.\",\n      \"method\": \"Co-IP, K63-specific ubiquitination assay, LSD1 demethylase activity assay, site-directed mutagenesis (K422), gene expression analysis, NSCLC cell and mouse models\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — site-directed mutagenesis of ubiquitination site, enzymatic (demethylase) activity assay, downstream pathway validation; single lab with multiple orthogonal methods\",\n      \"pmids\": [\"37979167\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"TRIM35 directly monoubiquitinates lysine-120 (K120) on histone H2B in postnatal cardiomyocytes, promoting chromatin remodeling and accessibility of P53 to its transcriptional promoter targets, thereby increasing P53 transcriptional output and contributing to dilated cardiomyopathy.\",\n      \"method\": \"ChIP-seq, RNA-seq, overexpression transgenic mice (cardiomyocyte-specific), siRNA knockdown in primary cardiomyocytes, adenovirus-mediated gene delivery, patient DCM LV samples\",\n      \"journal\": \"Circulation research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — ChIP-seq, direct H2B K120 monoubiquitination, in vivo cardiomyocyte-specific transgenic model, translational human DCM data; multiple orthogonal methods in single study\",\n      \"pmids\": [\"38860363\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"TRIM35 interacts with and ubiquitinates SLC7A5 (an amino acid transporter), increasing its stability and enhancing amino acid transport, which activates mTORC1 signaling in cardiac fibroblasts to promote fibroblast proliferation, migration, and differentiation, driving cardiac remodeling.\",\n      \"method\": \"LC-MS/MS substrate identification, Co-IP, ubiquitination assay, fibroblast-specific Trim35 conditional knockout mice (TAC surgery), adenoviral/AAV overexpression, mTORC1 pathway assays, RNA-seq\",\n      \"journal\": \"Cell communication and signaling\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — fibroblast-specific conditional KO mice, LC-MS/MS substrate discovery, Co-IP and ubiquitination of SLC7A5, mTORC1 rescue experiments; multiple orthogonal methods\",\n      \"pmids\": [\"39304904\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"TRIM35 interacts with adenovirus E1A protein and promotes its K48-linked ubiquitination at lysine residues K253 and K285, leading to E1A degradation and inhibition of HAdV replication.\",\n      \"method\": \"Co-IP, K48-linked ubiquitination assay, site-directed mutagenesis of E1A (K253A, K285A), viral replication assays, shTRIM35 oncolytic adenovirus construct\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, K48-specific ubiquitination, mutagenesis of substrate sites; single lab with two orthogonal methods\",\n      \"pmids\": [\"37578239\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"TRIM35 negatively regulates the cGAS-STING antiviral signaling pathway by directly interacting with STING and removing K63-linked ubiquitin from STING via its RING domain (C36 and C44 sites acting as deubiquitinase-like activity), impairing STING interaction with TBK1/IKKε and attenuating IFN-β production.\",\n      \"method\": \"Co-IP, co-localization (cytoplasm), K63-ubiquitin assay, RING domain mutagenesis (C36A, C44A), TBK1/IKKε interaction assays, IFN-β reporter assay\",\n      \"journal\": \"Inflammation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, site-directed RING domain mutagenesis, specific K63-deubiquitination of STING, downstream TBK1 assay; single lab, multiple methods\",\n      \"pmids\": [\"39088122\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Porcine TRIM35 directly interacts with TRAF3 and catalyzes K63-linked polyubiquitination of TRAF3, leading to upregulation of IFN-β production; RING and PRY/SPRY domains are essential for E3 ligase activity.\",\n      \"method\": \"Co-IP, K63-linked ubiquitination assay, domain deletion mutants (RING, PRY/SPRY), IFN-β luciferase reporter, overexpression/knockdown in ST cells, JEV infection model\",\n      \"journal\": \"Developmental and comparative immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, K63-specific ubiquitination, domain requirement mapping; ortholog (porcine) data with domain architecture consistent with human gene; single lab\",\n      \"pmids\": [\"34626690\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"TRIM35 interacts with IRF5 and promotes its ubiquitination and degradation; loss of TRIM35 leads to increased IRF5 levels and enhanced LDHA expression and glycolysis in hepatocellular carcinoma cells.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, mass spectrometry metabolite analysis, western blot, qPCR, HCC cell functional assays\",\n      \"journal\": \"Journal of digestive diseases\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — Co-IP and ubiquitination assay support TRIM35-IRF5 interaction and degradation; single lab, limited mechanistic depth\",\n      \"pmids\": [\"37594849\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Hls5 (mouse ortholog of TRIM35/HLS5) relocalizes from cytoplasmic granules to the nucleus under GATA-1 influence, associates with both FOG-1 and GATA-1, and suppresses GATA-1-mediated transactivation and GATA-1 DNA binding, thereby impeding erythroid maturation.\",\n      \"method\": \"Co-IP (Hls5/FOG-1/GATA-1), subcellular localization (fluorescence microscopy), GATA-1 transactivation reporter assay, GATA-1 DNA binding assay (ChIP/EMSA), erythroid differentiation assays\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, transcriptional reporter, DNA binding assay; mouse ortholog, multiple orthogonal methods in single study\",\n      \"pmids\": [\"18063753\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"HLS5 (human TRIM35), an RBCC family protein, localizes to cytoplasmic granules and punctate nuclear bodies; enforced expression in HeLa cells inhibits cell growth, clonogenicity, and tumorigenicity, establishing it as a candidate tumor suppressor at chromosome 8p21.\",\n      \"method\": \"cDNA cloning, subcellular localization (immunofluorescence), HeLa cell overexpression, growth inhibition and colony-formation assays, xenograft tumorigenicity assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — direct localization imaging, functional tumor suppression in cell and xenograft assays; foundational study, multiple readouts\",\n      \"pmids\": [\"14662771\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"TRIM35 directly binds genomic promoters (DNA-binding activity), interacts with histone H3, and catalyzes non-proteolytic (monoubiquitination) ubiquitination of H3, serving as a recruitment signal for p300 acetyltransferase, leading to H3K27 acetylation and transcriptional activation of HSPA6, which suppresses breast cancer progression.\",\n      \"method\": \"ChIP-PCR, Co-IP, ubiquitination assay (non-proteolytic), p300 recruitment assay, H3K27ac western blot, HSPA6 expression analysis, breast cancer cell functional assays\",\n      \"journal\": \"Cell death discovery\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — ChIP, Co-IP, ubiquitination and downstream H3K27ac assays; single lab, novel mechanism not yet replicated\",\n      \"pmids\": [\"41136372\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Endothelial TRIM35 inhibits MMP10 expression and secretion by promoting K63-linked ubiquitination of RelB and maintaining its nuclear localization, thereby suppressing noncanonical NF-κB-driven MMP10 transcription; TRIM35 endothelial-specific conditional knockout leads to increased MMP10 release and promotes vascular graft calcification via SMC paracrine signaling.\",\n      \"method\": \"Endothelial-specific TRIM35 cKO mice (arterial isograft model), single-cell RNA-seq, Co-IP, K63-ubiquitination assay of RelB, RelB nuclear localization assay, MMP10 ELISA, in vitro SMC calcification co-culture\",\n      \"journal\": \"Advanced science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — conditional endothelial KO in vivo, K63-ubiquitination of RelB, RelB nuclear localization, MMP10 functional rescue; single lab, multiple orthogonal methods\",\n      \"pmids\": [\"39865905\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"TRIM35 ubiquitinates EIF3E and promotes its degradation, modulating the CDK4/Cyclin D1 signaling pathway to suppress endometrial cancer cell proliferation.\",\n      \"method\": \"Co-IP, ubiquitination assay, EIF3E overexpression rescue, CDK4/Cyclin D1 western blot, cell proliferation and invasion assays\",\n      \"journal\": \"Cellular signalling\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, Co-IP and ubiquitination assay; limited mechanistic follow-up; no independent replication\",\n      \"pmids\": [\"41429342\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"TRIM35 is a RING-domain E3 ubiquitin ligase (and in some contexts a deubiquitinase/ubiquitin editor) that suppresses tumor growth and regulates innate immunity and cardiac homeostasis through substrate-specific ubiquitination: it interacts with and ubiquitinates PKM2 (blocking Y105 phosphorylation and the Warburg effect), IRF7 (K48-linked degradation to dampen TLR7/9-driven IFN production), LSD1 (K63-linked ubiquitination inhibiting demethylase activity to enhance tumor immunogenicity), nuclear histone H2B (K120 monoubiquitination promoting P53 transcriptional access in cardiomyocytes), STING (removal of K63-linked ubiquitin attenuating cGAS-STING signaling), SLC7A5 (stabilizing the amino acid transporter to activate mTORC1 in cardiac fibroblasts), PDK1, TIGAR, CLOCK, EIF3E, viral E1A, and RelB, while also localizing to the nucleus under GATA-1 direction to co-repress GATA-1 transcriptional activity during erythroid differentiation.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"TRIM35 (HLS5) is a RING-domain TRIM-family E3 ubiquitin ligase that functions as a tumor suppressor and a substrate-specific regulator of cellular metabolism, innate immune signaling, and cardiovascular homeostasis, originally identified as a chromosome 8p21 candidate tumor suppressor whose enforced expression inhibits growth, clonogenicity, and tumorigenicity [#15]. A core oncometabolic activity is its control of pyruvate kinase M2: TRIM35 binds PKM2 through its coiled-coil domain, suppresses PKM2 Y105 phosphorylation, and shifts PKM2 toward its tetrameric state, thereby blunting the Warburg effect and tumorigenicity [#0, #2]. Through substrate ubiquitination TRIM35 restrains multiple pro-tumor and pro-glycolytic factors, including K48-linked degradation of PDK1 to inactivate AKT signaling [#6], degradation of EIF3E to modulate CDK4/Cyclin D1 [#18], and K63-linked ubiquitination of LSD1 at K422 that represses its demethylase activity to enhance IFN-\\u03b3 signaling and MHC class I expression and tumor immunogenicity [#7]. TRIM35 also acts directly on chromatin: it binds promoters, monoubiquitinates histone H2B at K120 and histone H3, and recruits p300, thereby remodeling chromatin to control P53 and HSPA6 transcriptional programs [#8, #16]. In innate immunity TRIM35 is a bidirectional regulator, promoting K48-linked degradation of IRF7 to dampen TLR7/9-driven type I interferon as a negative feedback loop [#1] and removing K63-linked ubiquitin from STING to attenuate cGAS-STING signaling [#11]. In the cardiovascular system, cardiomyocyte TRIM35 drives dilated cardiomyopathy by degrading nuclear PKM2 and enhancing P53 output [#3, #8], while it stabilizes SLC7A5 to activate mTORC1 in cardiac fibroblasts [#9] and ubiquitinates RelB in endothelium to restrain MMP10-driven vascular calcification [#17].\",\n  \"teleology\": [\n    {\n      \"year\": 2003,\n      \"claim\": \"Established TRIM35/HLS5 as a candidate tumor suppressor, answering whether this 8p21 RBCC protein has growth-regulatory function before any catalytic mechanism was known.\",\n      \"evidence\": \"cDNA cloning, immunofluorescence localization, and growth/clonogenicity/xenograft assays in HeLa cells\",\n      \"pmids\": [\"14662771\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"No molecular substrate or enzymatic activity defined\", \"Mechanism of growth suppression unknown\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Linked TRIM35 to transcriptional control by showing its ortholog co-represses GATA-1, addressing how it might influence differentiation programs.\",\n      \"evidence\": \"Co-IP, subcellular localization, GATA-1 reporter and DNA-binding assays in erythroid differentiation models (mouse ortholog Hls5)\",\n      \"pmids\": [\"18063753\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Repression mechanism not tied to ubiquitin ligase activity\", \"Mouse ortholog data only\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Identified PKM2 as a TRIM35 partner and defined its anti-Warburg activity, revealing how the tumor suppressor reprograms cancer metabolism.\",\n      \"evidence\": \"Mass spectrometry, reciprocal Co-IP with domain mapping, PKM2 Y105 phosphorylation assay, HCC proliferation and xenograft assays\",\n      \"pmids\": [\"25263439\"],\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Whether PKM2 regulation requires ubiquitination not resolved here\", \"Coiled-coil binding mapped but ligase mechanism on PKM2 unclear\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Defined TRIM35 as a negative feedback regulator of innate immunity by targeting IRF7 for degradation, establishing its E3 ligase role in interferon control.\",\n      \"evidence\": \"Co-IP, K48-linkage ubiquitination assay, proteasome inhibitor rescue, TLR7/9 stimulation with overexpression/knockdown\",\n      \"pmids\": [\"25907537\"],\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Ubiquitination site on IRF7 not mapped\", \"Single lab\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Extended TRIM35 substrate scope to CLOCK and TRAF3, showing context-dependent degradative versus activating ubiquitination in lymphoma and antiviral immunity.\",\n      \"evidence\": \"Co-IP, ubiquitination assays, IFN-\\u03b2 reporter, NK co-culture, JEV infection model (porcine TRIM35 for TRAF3)\",\n      \"pmids\": [\"34124276\", \"34626690\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"TRAF3 data is from porcine ortholog\", \"Mechanistic depth on NK link and ubiquitin linkage outcomes limited\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Demonstrated TRIM35's role in cardiac and renal pathophysiology and broadened its oncometabolic substrate set, showing the same ligase activity drives disparate disease phenotypes.\",\n      \"evidence\": \"Cardiomyocyte-specific transgenic and PKM2-KO mice, renal IRI model, Co-IP and ubiquitination assays for PKM2, TIGAR, PDK1; breast cancer PKM2 tetramer assays\",\n      \"pmids\": [\"36322626\", \"35421414\", \"35081724\", \"36196894\"],\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"TIGAR, PDK1 findings are single-lab Medium evidence\", \"Tissue-specific determinants of substrate selection unclear\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Revealed non-degradative K63 ubiquitination as a TRIM35 mechanism via LSD1, linking the ligase to chromatin/immunogenicity, and expanded substrates to E1A and IRF5.\",\n      \"evidence\": \"Co-IP, K63-specific and site-directed ubiquitination assays (LSD1 K422; E1A K253/K285), demethylase activity assay, viral replication and HCC metabolite assays\",\n      \"pmids\": [\"37979167\", \"37578239\", \"37594849\"],\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"IRF5 link is single-lab Weak evidence\", \"How TRIM35 selects K48 versus K63 linkages not established\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Defined direct histone modification and transporter stabilization activities, showing TRIM35 acts on chromatin (H2B K120 monoubiquitination) and on SLC7A5 to drive cardiac remodeling, and removes K63-Ub from STING.\",\n      \"evidence\": \"ChIP-seq, cardiomyocyte- and fibroblast-specific transgenic/cKO mice, LC-MS/MS substrate ID, Co-IP, ubiquitination assays, STING RING-domain mutagenesis (C36/C44)\",\n      \"pmids\": [\"38860363\", \"39304904\", \"39088122\"],\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"STING deubiquitinase-like activity is Medium single-lab evidence\", \"How a RING ligase performs ubiquitin removal mechanistically unresolved\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Established TRIM35 as a chromatin-binding transcriptional activator (H3 monoubiquitination, p300 recruitment) and a vascular regulator via RelB, broadening its mechanistic repertoire beyond degradative ubiquitination.\",\n      \"evidence\": \"ChIP-PCR, endothelial-specific cKO mice, scRNA-seq, Co-IP, K63-ubiquitination of RelB, H3K27ac and MMP10 functional assays\",\n      \"pmids\": [\"41136372\", \"39865905\", \"41429342\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Direct DNA-binding mode and sequence specificity uncharacterized\", \"EIF3E finding is Low-confidence single lab\", \"Non-proteolytic H3 ubiquitination mechanism novel and unreplicated\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unresolved how a single RING-domain ligase achieves its broad substrate range and switches between K48-degradative, K63-non-degradative, monoubiquitinating, and ubiquitin-editing activities in a tissue- and context-specific manner.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"No structural model of substrate recognition\", \"Determinants of linkage-type and proteolytic versus non-proteolytic outcomes unknown\", \"Direct DNA-binding specificity undefined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016740\", \"supporting_discovery_ids\": [1, 3, 6, 7, 8, 9, 10, 12, 16, 17]},\n      {\"term_id\": \"GO:0016874\", \"supporting_discovery_ids\": [1, 3, 7, 8, 9]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [1, 3, 6, 9, 10, 18]},\n      {\"term_id\": \"GO:0003677\", \"supporting_discovery_ids\": [16]},\n      {\"term_id\": \"GO:0042393\", \"supporting_discovery_ids\": [8, 16]},\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [8, 14, 16]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [3, 8, 14, 15, 16]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [11, 14, 15]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [0, 2, 3, 13]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [1, 7, 11, 12]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [8, 14, 16, 17]},\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [1, 3, 6, 9, 18]},\n      {\"term_id\": \"R-HSA-9612973\", \"supporting_discovery_ids\": [7]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"PKM2\", \"IRF7\", \"LSD1\", \"SLC7A5\", \"STING1\", \"RelB\", \"TIGAR\", \"PDK1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":6,"faith_pct":83.33333333333333}}