{"gene":"UBE3C","run_date":"2026-06-10T10:51:56","timeline":{"discoveries":[{"year":2013,"finding":"UBE3C enhances proteasome processivity by ubiquitinating partially proteolyzed substrates; knockdown causes slower and incomplete degradation of destabilizing domain-GFP reporters, and this processivity function requires UBE3C catalytic activity, its ability to bind the proteasome, and lysine residues on the substrate.","method":"Forward genetic screen, siRNA knockdown, active-site mutagenesis, proteasome-binding mutants, substrate lysine-less mutants, polyubiquitination assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — multiple orthogonal methods (genetic screen, mutagenesis, in vitro ubiquitination, cell-based degradation assays) in a single rigorous study","pmids":["24158444"],"is_preprint":false},{"year":2017,"finding":"UBE3C (and USP14) dynamically cycle on and off the 26S proteasome, and the presence of ubiquitinated substrate proteins promotes their stable association; ubiquitin conjugates on the proteasome also enhance USP14 binding, which in turn further stabilizes UBE3C binding.","method":"Biochemical fractionation, purified proteasome binding assays with recombinant USP14, inhibitor treatments (IU-1, ubiquitin aldehyde), cell-based ubiquitination blockade experiments","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — reciprocal biochemical reconstitution with purified components plus cell-based corroboration, multiple orthogonal approaches","pmids":["28396413"],"is_preprint":false},{"year":2019,"finding":"Acute protein misfolding (unfolding of AgDD) rapidly recruits UBE3C to the 26S proteasome and triggers ubiquitylation of the proteasomal ubiquitin receptors RPN10 and RPN13; this is an immediate cellular response to misfolded monomers/oligomers rather than insoluble aggregates.","method":"Global diglycine-capture (K-GG) ubiquitin proteomics, engineered FKBP-based destabilizing domain system, quantitative MS","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — global ubiquitin proteomics with temporal resolution and orthogonal biochemical validation in a well-controlled inducible misfolding system","pmids":["31375563"],"is_preprint":false},{"year":2020,"finding":"Crystal structure of the UBE3C HECT domain (aa 693–1083) at 2.7 Å reveals an open, L-shaped bilobed conformation; Lys903 is the major autoubiquitination site; deletion of the last three C-terminal residues abolishes activity; mutations of Gln961 and Ser1049 substantially reduce autoubiquitination; the N-terminal region (aa 693–743) and a loop (aa 758–762) in the N-lobe affect stability and activity; these regions are involved in E2-E3 transthiolation.","method":"X-ray crystallography (2.7 Å), in vitro ubiquitination assays, site-directed mutagenesis, deletion analysis","journal":"The Biochemical journal","confidence":"High","confidence_rationale":"Tier 1 / Moderate — crystal structure combined with multiple in vitro mutagenesis and activity assays in a single study","pmids":["32039437"],"is_preprint":false},{"year":2021,"finding":"UBE3C assembles K29/K48-branched ubiquitin chains on VPS34, promoting VPS34 binding to proteasomes and its degradation, thereby suppressing autophagosome formation and maturation; TRABID deubiquitinase reverses this modification. Under ER/proteotoxic stress, UBE3C is redistributed from phagophores to proteasomes, attenuating VPS34 ubiquitination and elevating autophagy.","method":"Co-immunoprecipitation, ubiquitin chain-linkage mass spectrometry, in vitro ubiquitination assays, siRNA knockdown, live-cell imaging, subcellular fractionation, in vivo liver metabolic studies","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — multiple orthogonal methods including in vitro reconstitution, MS-based chain-type identification, genetic perturbations, and in vivo validation; replicated across cell and animal models","pmids":["33637724"],"is_preprint":false},{"year":2021,"finding":"UBE3C interacts with ERα during mitosis in an estrogen-dependent manner; estrogen stimulates UBE3C E3 ligase activity in the presence of ERα in vitro; UBE3C ubiquitinates cyclin B1 (CCNB1) and promotes its degradation during mitosis; ERα, UBE3C, and CCNB1 co-localize in prophase nuclei and metaphase spindles; UBE3C depletion attenuates estrogen-dependent cell proliferation.","method":"Co-immunoprecipitation from mitotic MCF-7 cells, in vitro E3 ubiquitin ligase assay, siRNA knockdown, immunofluorescence co-localization","journal":"Molecular endocrinology (Baltimore, Md.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP, in vitro E3 assay, and cell imaging are orthogonal but from a single lab","pmids":["26389696"],"is_preprint":false},{"year":2021,"finding":"UBE3C promotes ubiquitination and proteasomal degradation of AXIN1, thereby increasing β-catenin nuclear accumulation and activating Wnt/β-catenin signaling in gastric cancer cells; knockdown increases AXIN1 and reduces nuclear β-catenin.","method":"Co-immunoprecipitation, ubiquitination assay, siRNA knockdown and overexpression, xenograft mouse model, western blot","journal":"Carcinogenesis","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — Co-IP plus ubiquitination assay plus in vivo validation, single lab","pmids":["32930707"],"is_preprint":false},{"year":2022,"finding":"Ube3c targets progesterone receptor (PR) for polyubiquitination and proteasomal degradation; P38α kinase phosphorylates Ube3c at serine 741, restraining its polyubiquitination activity toward PR; uterine-selective P38α deletion leads to excessive Ube3c-mediated PR degradation, defective uterine receptivity, and female infertility.","method":"Conditional knockout mouse model, Co-immunoprecipitation, ubiquitination assay, phosphorylation site mutagenesis, genetic rescue experiments","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — in vivo genetic evidence combined with biochemical ubiquitination assays and phosphorylation site mutagenesis in one rigorous study","pmids":["35914132"],"is_preprint":false},{"year":2024,"finding":"UBE3C assembles K33-branched ubiquitin chains on ATG4B at Lys119 without causing ATG4B degradation; this ubiquitination inhibits ATG4B activity and its interaction with LC3, suppressing autophagy flux; under starvation, the ATG4B–UBE3C interaction decreases with concomitant removal of K33-branched chains, allowing starvation-induced autophagy.","method":"Mass spectrometry identification, Co-immunoprecipitation, in vitro ubiquitination assay, site-directed mutagenesis (K119R ATG4B), autophagy flux assays, overexpression/knockout cell studies","journal":"Autophagy","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — in vitro reconstitution of ubiquitination, mutagenesis rescue (K119R), MS-based chain-type identification, and functional autophagy readouts in one study","pmids":["38146933"],"is_preprint":false},{"year":2024,"finding":"UBE3C sequentially follows BAP1 to control IRF3 stability during viral infection: in the early innate immune phase, BAP1 removes K48-linked ubiquitination from IRF3 in the nucleus; in the late phase, IFN-β-induced UBE3C mediates K48-linked ubiquitination of IRF3, promoting its proteasomal degradation to resolve the antiviral response.","method":"Co-immunoprecipitation, ubiquitination assay, siRNA knockdown, overexpression, temporal analysis of viral infection stages","journal":"Cell reports","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — reciprocal Co-IP and ubiquitination assays with temporal knockdown studies, single lab","pmids":["39120972"],"is_preprint":false},{"year":2024,"finding":"UBE3C facilitates ERAD of misfolded CFTR (ΔF508) and ΔY490-ABCB1 independently of RNF185/RNF5; UBE3C knockdown has limited impact on CFTR ubiquitination itself but stabilizes mature ΔF508-CFTR and increases its plasma membrane expression, as well as stabilizing a class VI CFTR mutant (T70-CFTR).","method":"siRNA knockdown, cell surface biotinylation, western blot, functional channel assays, combined RNF5/185 ablation with UBE3C knockdown","journal":"Cells","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — clean knockdown with defined protein stability and surface expression readouts, multiple CFTR mutant substrates tested, single lab","pmids":["38067172"],"is_preprint":false},{"year":2024,"finding":"UBE3C binds the 2C protein of EV-A71 (via its C-terminal domain) and promotes K33/K48-linked ubiquitination of 2C at Lys268, leading to its degradation and restriction of viral replication; the K268R mutant 2C resists UBE3C-mediated degradation; UBE3C also ubiquitinates 2C from CVB3 and CVA16.","method":"Co-immunoprecipitation, in vitro ubiquitination assay, site-directed mutagenesis (2C K268R), recombinant virus experiments, siRNA knockdown and overexpression, viral titer measurement","journal":"Journal of virology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, in vitro ubiquitination, and mutagenesis rescue are orthogonal but from a single lab","pmids":["39212385"],"is_preprint":false},{"year":2024,"finding":"Acute lower-level misfolded protein aggregates (oligomers) are degraded via a UBE3C-dependent proteasomal pathway that is independent of RPN13 ubiquitylation by UBE3C; higher aggregate burden activates NRF1-dependent proteasome upregulation instead; no evidence for autophagy involvement in aggregate turnover.","method":"Inducible agDD-GFP aggregate system, targeted gene knockdown, cryo-electron tomography, quantitative protein degradation assays","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — synchronous aggregate induction system, cryo-ET structural validation, orthogonal knockdown/KO experiments, distinguishes UBE3C-dependent vs NRF1-dependent pathways","pmids":["39636856"],"is_preprint":false},{"year":2025,"finding":"UBE3C interacts with BRAF V600E via the kinase domain and promotes its ubiquitination; BRAF V600E stability is modulated by UBE3C expression and also depends on HSP90 activity; UBE3C knockout increases BRAF V600E levels in multiple myeloma cells.","method":"Tandem affinity purification, Co-immunoprecipitation, ubiquitination assay, HSP90 inhibitor treatment, CRISPR/siRNA knockdown","journal":"Life sciences","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, Co-IP/pulldown with ubiquitination assay but limited mechanistic follow-up on linkage or site","pmids":["40602747"],"is_preprint":false},{"year":2025,"finding":"UBE3C catalyzes p53 ubiquitination, promoting its proteasomal degradation in pancreatic ductal adenocarcinoma cells; UBE3C knockdown increases p53 levels and promotes apoptosis, effects reversed by the p53 inhibitor pifithrin-α.","method":"Co-immunoprecipitation, ubiquitination assay, siRNA knockdown and overexpression, pharmacological p53 inhibitor rescue, cell proliferation and apoptosis assays","journal":"Molecular biology reports","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, Co-IP and ubiquitination assay without site mapping or mutagenesis","pmids":["40553397"],"is_preprint":false},{"year":2025,"finding":"The SLAP adaptor protein interacts with mLST8 and recruits UBE3C to mediate non-degradative ubiquitination of mLST8 at Lys86 and Lys215, reducing mTORC2 complex integrity and suppressing mTORC2-AKT signaling in colorectal cancer cells.","method":"Co-immunoprecipitation, ubiquitination assay with mutagenesis (K86R/K215R), Co-IP of SLAP-UBE3C interaction, mTORC2 integrity assays, xenograft mouse model","journal":"Cell death and differentiation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP identifying UBE3C as SLAP interactor plus site-specific mutagenesis and in vivo validation, single lab","pmids":["41398047"],"is_preprint":false},{"year":2025,"finding":"UBE3C regulates cellular composition of the murine cerebral cortex and human brain organoids; its loss favors neurogenesis and suppresses glial fate; disease-associated UBE3C mutations alter autoubiquitination activity and disrupt cortical lamination; proteomic profiling identifies Cbll1 (a m6A methyltransferase component) as a UBE3C substrate; the UBE3C–Cbll1 axis drives m6A mRNA methylation in neural progenitors; hyperactivation of m6A writers in UBE3C-deficient progenitors impairs cell cycle exit, reversible by the METTL3 inhibitor STM2457.","method":"Genetic complementation in mouse and human brain organoids, proteomic substrate profiling, autoubiquitination assays, disease-variant mutagenesis, METTL3 inhibitor rescue in vivo","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal methods (proteomics, genetics, in vivo rescue) in a preprint; not yet peer-reviewed","pmids":["bio_10.1101_2025.04.09.646620"],"is_preprint":true},{"year":2025,"finding":"PhIX-MS crosslinking coupled with cryo-EM places UBE3C/Hul5 along the 19S regulatory particle (RP) of the proteasome with its catalytic HECT domain positioned above the RPN11 deubiquitinase active site, enabling coupled ubiquitination and deubiquitination activities at the proteasome entry channel.","method":"Photo-induced in situ crosslinking mass spectrometry (PhIX-MS), cryo-electron microscopy, AlphaFold structural modeling","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — structural proteomics method is Tier 1 quality but preprint, single study, not independently replicated","pmids":["bio_10.1101_2025.07.31.667872"],"is_preprint":true},{"year":2025,"finding":"CRISPR loss-of-function screen in multiple myeloma cells shows UBE3C knockout markedly increases endogenous MYC protein levels, suggesting UBE3C negatively regulates MYC; paralogs UBE3A and UBE3B showed no measurable effect, indicating specificity.","method":"Genome-wide CRISPR-Cas9 loss-of-function screen, endogenous MYC-GFP reporter, FACS sorting, next-generation sequencing, functional validation of individual knockouts","journal":"Scientific reports","confidence":"Low","confidence_rationale":"Tier 3 / Weak — CRISPR screen with functional validation but mechanism of MYC regulation (direct vs indirect ubiquitination) not established","pmids":["41965876"],"is_preprint":false},{"year":2023,"finding":"UBE3C mediates ubiquitination-dependent proteasomal degradation of TP73, contributing to radioresistance in breast cancer cells; FOSB transcriptionally activates UBE3C, and LINC00963 recruits FOSB to the UBE3C promoter.","method":"Co-immunoprecipitation, immunofluorescence, siRNA knockdown and overexpression, in vitro/in vivo functional assays, rescue experiments","journal":"Journal of translational medicine","confidence":"Low","confidence_rationale":"Tier 3 / Weak — Co-IP and ubiquitination evidence for TP73 as substrate; limited biochemical mechanistic detail; single lab","pmids":["37173692"],"is_preprint":false},{"year":2025,"finding":"UBE3C promotes ubiquitination and degradation of AHNAK protein in osteosarcoma cells, suppressing ferroptosis; METTL5-mediated m6A modification of UBE3C mRNA enhances its stability via YTHDF1 binding.","method":"Co-immunoprecipitation, ubiquitination assay, siRNA knockdown, ferroptosis assays, m6A methylation analysis","journal":"Journal of molecular histology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, Co-IP and ubiquitination assay without site mapping","pmids":["40696164"],"is_preprint":false}],"current_model":"UBE3C is a HECT-domain E3 ubiquitin ligase (crystal structure resolved at 2.7 Å) that associates dynamically with the 26S proteasome in a substrate-dependent manner, where it enhances proteasomal processivity by ubiquitinating partially proteolyzed and misfolded substrates; it assembles atypical ubiquitin chain linkages (K29/K48-branched on VPS34; K33-branched on ATG4B; K33/K48-branched on viral 2C) to regulate autophagy, proteostasis, and antiviral immunity, and it targets diverse substrates including VPS34, cyclin B1, AXIN1, progesterone receptor, IRF3, CFTR, TP73, p53, and mLST8, with its activity modulated by ERα binding, P38α-mediated phosphorylation at Ser741, and substrate availability at the proteasome."},"narrative":{"mechanistic_narrative":"UBE3C is a HECT-domain E3 ubiquitin ligase that functions as a processivity factor for the 26S proteasome, ubiquitinating partially proteolyzed and misfolded substrates to ensure their complete degradation [PMID:24158444]. It cycles dynamically on and off the proteasome, with stable association driven by the presence of ubiquitinated substrate and stabilized cooperatively by USP14 binding [PMID:28396413]; structural placement of its catalytic HECT domain above the RPN11 deubiquitinase active site at the 19S regulatory particle positions UBE3C to couple ubiquitination with substrate processing at the proteasome entry channel [PMID:bio_10.1101_2025.07.31.667872]. Its catalytic HECT domain adopts an open, L-shaped bilobed fold whose C-terminal residues and N-lobe loops are required for E2-E3 transthiolation and activity, with Lys903 as the major autoubiquitination site [PMID:32039437]. UBE3C responds acutely to protein misfolding, being recruited to the proteasome and ubiquitinating the ubiquitin receptors RPN10 and RPN13 upon unfolding stress [PMID:31375563], and it clears low-level misfolded oligomers through a proteasomal route distinct from NRF1-dependent proteasome upregulation [PMID:39636856]; it also contributes to ER-associated degradation of misfolded CFTR (ΔF508) and ABCB1 [PMID:38067172]. Beyond canonical degradation, UBE3C assembles atypical branched ubiquitin chains to regulate autophagy: K29/K48-branched chains on VPS34 (reversed by TRABID) drive its proteasomal degradation and suppress autophagosome formation [PMID:33637724], and K33-branched chains on ATG4B Lys119 inhibit ATG4B activity and LC3 interaction non-degradatively [PMID:38146933]. UBE3C further targets specific substrates across diverse contexts — cyclin B1 during estrogen/ERα-stimulated mitosis [PMID:26389696], progesterone receptor under P38α-restrained control important for uterine receptivity [PMID:35914132], AXIN1 to activate Wnt/β-catenin signaling [PMID:32930707], IRF3 via K48 chains to resolve antiviral responses [PMID:39120972], picornaviral 2C protein via K33/K48 chains to restrict viral replication [PMID:39212385], and mLST8 (non-degradatively, recruited by the SLAP adaptor) to limit mTORC2-AKT signaling [PMID:41398047]. P38α-mediated phosphorylation at Ser741 restrains UBE3C ligase activity [PMID:35914132].","teleology":[{"year":2013,"claim":"Established UBE3C's core cellular role as a proteasome-associated processivity factor, answering why some substrates escape complete degradation.","evidence":"Forward genetic screen, siRNA knockdown, active-site and proteasome-binding mutagenesis, and substrate lysine-less mutants with polyubiquitination assays","pmids":["24158444"],"confidence":"High","gaps":["Did not define which proteasome subunit anchors UBE3C","Chain linkage type used at the proteasome not resolved"]},{"year":2017,"claim":"Showed UBE3C association with the proteasome is substrate-driven and cooperative with USP14, explaining how its recruitment is gated.","evidence":"Biochemical fractionation and purified proteasome binding assays with recombinant USP14, plus inhibitor and cell-based blockade experiments","pmids":["28396413"],"confidence":"High","gaps":["Molecular basis of the USP14–UBE3C cooperativity not structurally defined","Whether deubiquitination and ubiquitination are spatially coordinated unresolved at the time"]},{"year":2019,"claim":"Demonstrated UBE3C is an immediate responder to soluble misfolded protein, ubiquitinating proteasomal ubiquitin receptors RPN10/RPN13 as part of the response.","evidence":"Global K-GG ubiquitin proteomics with an inducible FKBP destabilizing-domain misfolding system and quantitative MS","pmids":["31375563"],"confidence":"High","gaps":["Functional consequence of RPN10/RPN13 ubiquitination for proteasome function not fully defined","Distinction between monomer/oligomer and aggregate substrates not yet established"]},{"year":2020,"claim":"Provided the structural framework for UBE3C catalysis, defining the HECT fold, autoubiquitination site, and residues required for transthiolation.","evidence":"2.7 Å X-ray crystallography of the HECT domain with in vitro ubiquitination, site-directed mutagenesis, and deletion analysis","pmids":["32039437"],"confidence":"High","gaps":["Structure of full-length protein and substrate-bound complexes not solved","Structural basis of branched-chain specificity not addressed"]},{"year":2021,"claim":"Revealed UBE3C builds atypical K29/K48-branched chains on VPS34 to suppress autophagy and that proteotoxic stress redistributes UBE3C between phagophores and proteasomes, linking proteostasis to autophagy regulation.","evidence":"Co-IP, ubiquitin chain-linkage MS, in vitro ubiquitination, knockdown, live-cell imaging, and in vivo liver metabolic studies","pmids":["33637724"],"confidence":"High","gaps":["Signals controlling UBE3C subcellular redistribution not fully defined","How TRABID is targeted to VPS34 chains unclear"]},{"year":2021,"claim":"Identified UBE3C as an ERα-stimulated mitotic ligase for cyclin B1 and as a Wnt-pathway regulator via AXIN1 degradation, extending its substrate repertoire to cell-cycle and signaling control.","evidence":"Co-IP from mitotic MCF-7 cells, in vitro E3 assays, immunofluorescence co-localization, knockdown, and xenograft models","pmids":["26389696","32930707"],"confidence":"Medium","gaps":["Ubiquitin linkage types on cyclin B1 and AXIN1 not mapped","Findings each from a single lab without reciprocal validation"]},{"year":2022,"claim":"Established physiological regulation of UBE3C by P38α phosphorylation at Ser741 and its in vivo importance, controlling progesterone receptor stability for uterine receptivity.","evidence":"Conditional knockout mice, Co-IP, ubiquitination assays, phosphorylation-site mutagenesis, and genetic rescue","pmids":["35914132"],"confidence":"High","gaps":["Whether Ser741 phosphorylation regulates UBE3C activity toward other substrates not tested","Mechanism by which phosphorylation restrains catalysis not structurally defined"]},{"year":2024,"claim":"Showed UBE3C can act non-degradatively, assembling K33-branched chains on ATG4B to inhibit its enzymatic activity, broadening the functional output of its ligase activity.","evidence":"MS site identification, Co-IP, in vitro ubiquitination, K119R mutagenesis rescue, and autophagy flux assays","pmids":["38146933"],"confidence":"High","gaps":["Deubiquitinase removing K33 chains during starvation not identified","Signal decreasing the ATG4B–UBE3C interaction unknown"]},{"year":2024,"claim":"Distinguished a UBE3C-dependent proteasomal route for clearing low-level misfolded oligomers from an NRF1-dependent pathway engaged at higher aggregate burden, and excluded autophagy in this turnover.","evidence":"Inducible agDD-GFP aggregate system, targeted knockdowns, cryo-electron tomography, and quantitative degradation assays","pmids":["39636856"],"confidence":"High","gaps":["How the cell switches between UBE3C- and NRF1-dependent responses not defined","Whether RPN13 ubiquitination plays any role in oligomer clearance contradicted prior model"]},{"year":2024,"claim":"Placed UBE3C in antiviral and ERAD contexts, mediating IRF3 K48 degradation to resolve interferon responses, restricting picornaviruses via 2C ubiquitination, and assisting degradation of misfolded CFTR.","evidence":"Co-IP, ubiquitination assays, site mutagenesis (2C K268R), recombinant virus and surface biotinylation experiments, knockdown","pmids":["39120972","39212385","38067172"],"confidence":"Medium","gaps":["IRF3 substrate handoff from BAP1 not reconstituted in vitro","CFTR effect appears largely independent of direct CFTR ubiquitination, mechanism unclear","Each result from a single lab"]},{"year":2025,"claim":"Identified adaptor-mediated and developmental roles — SLAP-recruited non-degradative mLST8 ubiquitination limiting mTORC2-AKT, and UBE3C control of cortical neural fate via the Cbll1–m6A axis.","evidence":"Co-IP, K86R/K215R mutagenesis, mTORC2 integrity assays, xenografts; brain organoid genetics, proteomic substrate profiling, and METTL3-inhibitor rescue (preprint)","pmids":["41398047","bio_10.1101_2025.04.09.646620"],"confidence":"Medium","gaps":["Disease-variant link to cortical phenotype remains in preprint and unreplicated","How SLAP redirects UBE3C specificity not structurally defined"]},{"year":2025,"claim":"Provided structural placement of UBE3C/Hul5 on the 19S regulatory particle above RPN11, proposing coupled ubiquitination and deubiquitination at the proteasome substrate entry channel.","evidence":"PhIX-MS crosslinking, cryo-EM, and AlphaFold modeling (preprint)","pmids":["bio_10.1101_2025.07.31.667872"],"confidence":"Medium","gaps":["Preprint, single study not independently replicated","Functional coupling between UBE3C and RPN11 not demonstrated biochemically"]},{"year":2025,"claim":"Extended the substrate list to oncogenic and tumor-suppressor proteins (p53, TP73, BRAF V600E, AHNAK) and MYC, implicating UBE3C in cancer phenotypes.","evidence":"Co-IP, ubiquitination assays, knockdown/CRISPR, MYC-GFP CRISPR screen with paralog specificity, and pharmacological rescues","pmids":["40553397","37173692","40602747","40696164","41965876"],"confidence":"Low","gaps":["Ubiquitination sites and chain linkages not mapped for these substrates","Direct vs indirect regulation of MYC not established","Each from a single lab without reciprocal validation"]},{"year":null,"claim":"How UBE3C achieves substrate and chain-linkage specificity — selecting among degradative K48 and non-degradative branched K29/K33 chains across its many substrates — and how adaptors, phosphorylation, and proteasome positioning collectively dictate outcome remain unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unifying structural model of substrate-bound UBE3C","Determinants of branched vs linear chain assembly unknown","Rules governing degradative vs non-degradative outcomes undefined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016874","term_label":"ligase activity","supporting_discovery_ids":[0,3,4,5,7,8,11,15]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,4,8,11]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[5,9]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[1,2]}],"pathway":[{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[0,1,2,12]},{"term_id":"R-HSA-9612973","term_label":"Autophagy","supporting_discovery_ids":[4,8]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[9,11]}],"complexes":[],"partners":["USP14","VPS34","ATG4B","ESR1","AXIN1","IRF3","MLST8","SLA"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q15386","full_name":"Ubiquitin-protein ligase E3C","aliases":["HECT-type ubiquitin transferase E3C","Homologous to E6AP carboxyl terminus homologous protein 2","HectH2","RTA-associated ubiquitin ligase","RAUL"],"length_aa":1083,"mass_kda":123.9,"function":"E3 ubiquitin-protein ligase that specifically catalyzes 'Lys-29'- and 'Lys-48'-linked polyubiquitin chains (PubMed:11278995, PubMed:12692129, PubMed:16341092, PubMed:16601690, PubMed:24158444, PubMed:24811749, PubMed:25752573, PubMed:25752577, PubMed:32039437, PubMed:33637724, PubMed:34239127). Accepts ubiquitin from the E2 ubiquitin-conjugating enzyme UBE2D1 in the form of a thioester and then directly transfers the ubiquitin to targeted substrates (PubMed:32039437, PubMed:9575161). Associates with the proteasome and promotes elongation of ubiquitin chains on substrates bound to the 26S proteasome (PubMed:24158444, PubMed:28396413, PubMed:31375563). Also catalyzes 'Lys-29'- and 'Lys-48'-linked ubiquitination of 26S proteasome subunit ADRM1/RPN13 in response to proteotoxic stress, impairing the ability of the proteasome to bind and degrade ubiquitin-conjugated proteins (PubMed:24811749, PubMed:31375563). Acts as a negative regulator of autophagy by mediating 'Lys-29'- and 'Lys-48'-linked ubiquitination of PIK3C3/VPS34, promoting its degradation (PubMed:33637724). Can assemble unanchored poly-ubiquitin chains in either 'Lys-29'- or 'Lys-48'-linked polyubiquitin chains; with some preference for 'Lys-48' linkages (PubMed:11278995, PubMed:16601690, PubMed:25752577). Acts as a negative regulator of type I interferon by mediating 'Lys-48'-linked ubiquitination of IRF3 and IRF7, leading to their degradation by the proteasome (PubMed:21167755). Catalyzes ubiquitination and degradation of CAND2 (PubMed:12692129)","subcellular_location":"","url":"https://www.uniprot.org/uniprotkb/Q15386/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/UBE3C","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":[{"gene":"PSMB7","stoichiometry":4.0},{"gene":"PSMD12","stoichiometry":4.0},{"gene":"CALM1","stoichiometry":0.2},{"gene":"CALM2","stoichiometry":0.2},{"gene":"CALM3","stoichiometry":0.2},{"gene":"PSMA1","stoichiometry":0.2},{"gene":"PSMA5","stoichiometry":0.2},{"gene":"PSMA6","stoichiometry":0.2},{"gene":"PSMB1","stoichiometry":0.2},{"gene":"PSMB2","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/UBE3C","total_profiled":1310},"omim":[{"mim_id":"620270","title":"NEURODEVELOPMENTAL DISORDER WITH ABSENT SPEECH AND MOVEMENT AND BEHAVIORAL ABNORMALITIES; NEDSMB","url":"https://www.omim.org/entry/620270"},{"mim_id":"620250","title":"NEURODEVELOPMENTAL DISORDER WITH SEIZURES, SPASTICITY, AND COMPLETE OR PARTIAL AGENESIS OF THE CORPUS CALLOSUM; NEDSSCC","url":"https://www.omim.org/entry/620250"},{"mim_id":"620209","title":"HECT DOMAIN E3 UBIQUITIN PROTEIN LIGASE 4; HECTD4","url":"https://www.omim.org/entry/620209"},{"mim_id":"614454","title":"UBIQUITIN PROTEIN LIGASE E3C; UBE3C","url":"https://www.omim.org/entry/614454"},{"mim_id":"182960","title":"NEURONOPATHY, DISTAL HEREDITARY MOTOR, AUTOSOMAL DOMINANT 1; HMND1","url":"https://www.omim.org/entry/182960"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Cytosol","reliability":"Approved"},{"location":"Vesicles","reliability":"Additional"},{"location":"Lipid droplets","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in 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\"method\": \"Forward genetic screen, siRNA knockdown, active-site mutagenesis, proteasome-binding mutants, substrate lysine-less mutants, polyubiquitination assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — multiple orthogonal methods (genetic screen, mutagenesis, in vitro ubiquitination, cell-based degradation assays) in a single rigorous study\",\n      \"pmids\": [\"24158444\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"UBE3C (and USP14) dynamically cycle on and off the 26S proteasome, and the presence of ubiquitinated substrate proteins promotes their stable association; ubiquitin conjugates on the proteasome also enhance USP14 binding, which in turn further stabilizes UBE3C binding.\",\n      \"method\": \"Biochemical fractionation, purified proteasome binding assays with recombinant USP14, inhibitor treatments (IU-1, ubiquitin aldehyde), cell-based ubiquitination blockade experiments\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — reciprocal biochemical reconstitution with purified components plus cell-based corroboration, multiple orthogonal approaches\",\n      \"pmids\": [\"28396413\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Acute protein misfolding (unfolding of AgDD) rapidly recruits UBE3C to the 26S proteasome and triggers ubiquitylation of the proteasomal ubiquitin receptors RPN10 and RPN13; this is an immediate cellular response to misfolded monomers/oligomers rather than insoluble aggregates.\",\n      \"method\": \"Global diglycine-capture (K-GG) ubiquitin proteomics, engineered FKBP-based destabilizing domain system, quantitative MS\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — global ubiquitin proteomics with temporal resolution and orthogonal biochemical validation in a well-controlled inducible misfolding system\",\n      \"pmids\": [\"31375563\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Crystal structure of the UBE3C HECT domain (aa 693–1083) at 2.7 Å reveals an open, L-shaped bilobed conformation; Lys903 is the major autoubiquitination site; deletion of the last three C-terminal residues abolishes activity; mutations of Gln961 and Ser1049 substantially reduce autoubiquitination; the N-terminal region (aa 693–743) and a loop (aa 758–762) in the N-lobe affect stability and activity; these regions are involved in E2-E3 transthiolation.\",\n      \"method\": \"X-ray crystallography (2.7 Å), in vitro ubiquitination assays, site-directed mutagenesis, deletion analysis\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — crystal structure combined with multiple in vitro mutagenesis and activity assays in a single study\",\n      \"pmids\": [\"32039437\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"UBE3C assembles K29/K48-branched ubiquitin chains on VPS34, promoting VPS34 binding to proteasomes and its degradation, thereby suppressing autophagosome formation and maturation; TRABID deubiquitinase reverses this modification. Under ER/proteotoxic stress, UBE3C is redistributed from phagophores to proteasomes, attenuating VPS34 ubiquitination and elevating autophagy.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitin chain-linkage mass spectrometry, in vitro ubiquitination assays, siRNA knockdown, live-cell imaging, subcellular fractionation, in vivo liver metabolic studies\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — multiple orthogonal methods including in vitro reconstitution, MS-based chain-type identification, genetic perturbations, and in vivo validation; replicated across cell and animal models\",\n      \"pmids\": [\"33637724\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"UBE3C interacts with ERα during mitosis in an estrogen-dependent manner; estrogen stimulates UBE3C E3 ligase activity in the presence of ERα in vitro; UBE3C ubiquitinates cyclin B1 (CCNB1) and promotes its degradation during mitosis; ERα, UBE3C, and CCNB1 co-localize in prophase nuclei and metaphase spindles; UBE3C depletion attenuates estrogen-dependent cell proliferation.\",\n      \"method\": \"Co-immunoprecipitation from mitotic MCF-7 cells, in vitro E3 ubiquitin ligase assay, siRNA knockdown, immunofluorescence co-localization\",\n      \"journal\": \"Molecular endocrinology (Baltimore, Md.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP, in vitro E3 assay, and cell imaging are orthogonal but from a single lab\",\n      \"pmids\": [\"26389696\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"UBE3C promotes ubiquitination and proteasomal degradation of AXIN1, thereby increasing β-catenin nuclear accumulation and activating Wnt/β-catenin signaling in gastric cancer cells; knockdown increases AXIN1 and reduces nuclear β-catenin.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, siRNA knockdown and overexpression, xenograft mouse model, western blot\",\n      \"journal\": \"Carcinogenesis\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — Co-IP plus ubiquitination assay plus in vivo validation, single lab\",\n      \"pmids\": [\"32930707\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Ube3c targets progesterone receptor (PR) for polyubiquitination and proteasomal degradation; P38α kinase phosphorylates Ube3c at serine 741, restraining its polyubiquitination activity toward PR; uterine-selective P38α deletion leads to excessive Ube3c-mediated PR degradation, defective uterine receptivity, and female infertility.\",\n      \"method\": \"Conditional knockout mouse model, Co-immunoprecipitation, ubiquitination assay, phosphorylation site mutagenesis, genetic rescue experiments\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — in vivo genetic evidence combined with biochemical ubiquitination assays and phosphorylation site mutagenesis in one rigorous study\",\n      \"pmids\": [\"35914132\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"UBE3C assembles K33-branched ubiquitin chains on ATG4B at Lys119 without causing ATG4B degradation; this ubiquitination inhibits ATG4B activity and its interaction with LC3, suppressing autophagy flux; under starvation, the ATG4B–UBE3C interaction decreases with concomitant removal of K33-branched chains, allowing starvation-induced autophagy.\",\n      \"method\": \"Mass spectrometry identification, Co-immunoprecipitation, in vitro ubiquitination assay, site-directed mutagenesis (K119R ATG4B), autophagy flux assays, overexpression/knockout cell studies\",\n      \"journal\": \"Autophagy\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — in vitro reconstitution of ubiquitination, mutagenesis rescue (K119R), MS-based chain-type identification, and functional autophagy readouts in one study\",\n      \"pmids\": [\"38146933\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"UBE3C sequentially follows BAP1 to control IRF3 stability during viral infection: in the early innate immune phase, BAP1 removes K48-linked ubiquitination from IRF3 in the nucleus; in the late phase, IFN-β-induced UBE3C mediates K48-linked ubiquitination of IRF3, promoting its proteasomal degradation to resolve the antiviral response.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, siRNA knockdown, overexpression, temporal analysis of viral infection stages\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — reciprocal Co-IP and ubiquitination assays with temporal knockdown studies, single lab\",\n      \"pmids\": [\"39120972\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"UBE3C facilitates ERAD of misfolded CFTR (ΔF508) and ΔY490-ABCB1 independently of RNF185/RNF5; UBE3C knockdown has limited impact on CFTR ubiquitination itself but stabilizes mature ΔF508-CFTR and increases its plasma membrane expression, as well as stabilizing a class VI CFTR mutant (T70-CFTR).\",\n      \"method\": \"siRNA knockdown, cell surface biotinylation, western blot, functional channel assays, combined RNF5/185 ablation with UBE3C knockdown\",\n      \"journal\": \"Cells\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — clean knockdown with defined protein stability and surface expression readouts, multiple CFTR mutant substrates tested, single lab\",\n      \"pmids\": [\"38067172\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"UBE3C binds the 2C protein of EV-A71 (via its C-terminal domain) and promotes K33/K48-linked ubiquitination of 2C at Lys268, leading to its degradation and restriction of viral replication; the K268R mutant 2C resists UBE3C-mediated degradation; UBE3C also ubiquitinates 2C from CVB3 and CVA16.\",\n      \"method\": \"Co-immunoprecipitation, in vitro ubiquitination assay, site-directed mutagenesis (2C K268R), recombinant virus experiments, siRNA knockdown and overexpression, viral titer measurement\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, in vitro ubiquitination, and mutagenesis rescue are orthogonal but from a single lab\",\n      \"pmids\": [\"39212385\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Acute lower-level misfolded protein aggregates (oligomers) are degraded via a UBE3C-dependent proteasomal pathway that is independent of RPN13 ubiquitylation by UBE3C; higher aggregate burden activates NRF1-dependent proteasome upregulation instead; no evidence for autophagy involvement in aggregate turnover.\",\n      \"method\": \"Inducible agDD-GFP aggregate system, targeted gene knockdown, cryo-electron tomography, quantitative protein degradation assays\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — synchronous aggregate induction system, cryo-ET structural validation, orthogonal knockdown/KO experiments, distinguishes UBE3C-dependent vs NRF1-dependent pathways\",\n      \"pmids\": [\"39636856\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"UBE3C interacts with BRAF V600E via the kinase domain and promotes its ubiquitination; BRAF V600E stability is modulated by UBE3C expression and also depends on HSP90 activity; UBE3C knockout increases BRAF V600E levels in multiple myeloma cells.\",\n      \"method\": \"Tandem affinity purification, Co-immunoprecipitation, ubiquitination assay, HSP90 inhibitor treatment, CRISPR/siRNA knockdown\",\n      \"journal\": \"Life sciences\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, Co-IP/pulldown with ubiquitination assay but limited mechanistic follow-up on linkage or site\",\n      \"pmids\": [\"40602747\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"UBE3C catalyzes p53 ubiquitination, promoting its proteasomal degradation in pancreatic ductal adenocarcinoma cells; UBE3C knockdown increases p53 levels and promotes apoptosis, effects reversed by the p53 inhibitor pifithrin-α.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, siRNA knockdown and overexpression, pharmacological p53 inhibitor rescue, cell proliferation and apoptosis assays\",\n      \"journal\": \"Molecular biology reports\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, Co-IP and ubiquitination assay without site mapping or mutagenesis\",\n      \"pmids\": [\"40553397\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"The SLAP adaptor protein interacts with mLST8 and recruits UBE3C to mediate non-degradative ubiquitination of mLST8 at Lys86 and Lys215, reducing mTORC2 complex integrity and suppressing mTORC2-AKT signaling in colorectal cancer cells.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay with mutagenesis (K86R/K215R), Co-IP of SLAP-UBE3C interaction, mTORC2 integrity assays, xenograft mouse model\",\n      \"journal\": \"Cell death and differentiation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP identifying UBE3C as SLAP interactor plus site-specific mutagenesis and in vivo validation, single lab\",\n      \"pmids\": [\"41398047\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"UBE3C regulates cellular composition of the murine cerebral cortex and human brain organoids; its loss favors neurogenesis and suppresses glial fate; disease-associated UBE3C mutations alter autoubiquitination activity and disrupt cortical lamination; proteomic profiling identifies Cbll1 (a m6A methyltransferase component) as a UBE3C substrate; the UBE3C–Cbll1 axis drives m6A mRNA methylation in neural progenitors; hyperactivation of m6A writers in UBE3C-deficient progenitors impairs cell cycle exit, reversible by the METTL3 inhibitor STM2457.\",\n      \"method\": \"Genetic complementation in mouse and human brain organoids, proteomic substrate profiling, autoubiquitination assays, disease-variant mutagenesis, METTL3 inhibitor rescue in vivo\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal methods (proteomics, genetics, in vivo rescue) in a preprint; not yet peer-reviewed\",\n      \"pmids\": [\"bio_10.1101_2025.04.09.646620\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"PhIX-MS crosslinking coupled with cryo-EM places UBE3C/Hul5 along the 19S regulatory particle (RP) of the proteasome with its catalytic HECT domain positioned above the RPN11 deubiquitinase active site, enabling coupled ubiquitination and deubiquitination activities at the proteasome entry channel.\",\n      \"method\": \"Photo-induced in situ crosslinking mass spectrometry (PhIX-MS), cryo-electron microscopy, AlphaFold structural modeling\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — structural proteomics method is Tier 1 quality but preprint, single study, not independently replicated\",\n      \"pmids\": [\"bio_10.1101_2025.07.31.667872\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"CRISPR loss-of-function screen in multiple myeloma cells shows UBE3C knockout markedly increases endogenous MYC protein levels, suggesting UBE3C negatively regulates MYC; paralogs UBE3A and UBE3B showed no measurable effect, indicating specificity.\",\n      \"method\": \"Genome-wide CRISPR-Cas9 loss-of-function screen, endogenous MYC-GFP reporter, FACS sorting, next-generation sequencing, functional validation of individual knockouts\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — CRISPR screen with functional validation but mechanism of MYC regulation (direct vs indirect ubiquitination) not established\",\n      \"pmids\": [\"41965876\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"UBE3C mediates ubiquitination-dependent proteasomal degradation of TP73, contributing to radioresistance in breast cancer cells; FOSB transcriptionally activates UBE3C, and LINC00963 recruits FOSB to the UBE3C promoter.\",\n      \"method\": \"Co-immunoprecipitation, immunofluorescence, siRNA knockdown and overexpression, in vitro/in vivo functional assays, rescue experiments\",\n      \"journal\": \"Journal of translational medicine\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — Co-IP and ubiquitination evidence for TP73 as substrate; limited biochemical mechanistic detail; single lab\",\n      \"pmids\": [\"37173692\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"UBE3C promotes ubiquitination and degradation of AHNAK protein in osteosarcoma cells, suppressing ferroptosis; METTL5-mediated m6A modification of UBE3C mRNA enhances its stability via YTHDF1 binding.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, siRNA knockdown, ferroptosis assays, m6A methylation analysis\",\n      \"journal\": \"Journal of molecular histology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, Co-IP and ubiquitination assay without site mapping\",\n      \"pmids\": [\"40696164\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"UBE3C is a HECT-domain E3 ubiquitin ligase (crystal structure resolved at 2.7 Å) that associates dynamically with the 26S proteasome in a substrate-dependent manner, where it enhances proteasomal processivity by ubiquitinating partially proteolyzed and misfolded substrates; it assembles atypical ubiquitin chain linkages (K29/K48-branched on VPS34; K33-branched on ATG4B; K33/K48-branched on viral 2C) to regulate autophagy, proteostasis, and antiviral immunity, and it targets diverse substrates including VPS34, cyclin B1, AXIN1, progesterone receptor, IRF3, CFTR, TP73, p53, and mLST8, with its activity modulated by ERα binding, P38α-mediated phosphorylation at Ser741, and substrate availability at the proteasome.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"UBE3C is a HECT-domain E3 ubiquitin ligase that functions as a processivity factor for the 26S proteasome, ubiquitinating partially proteolyzed and misfolded substrates to ensure their complete degradation [#0]. It cycles dynamically on and off the proteasome, with stable association driven by the presence of ubiquitinated substrate and stabilized cooperatively by USP14 binding [#1]; structural placement of its catalytic HECT domain above the RPN11 deubiquitinase active site at the 19S regulatory particle positions UBE3C to couple ubiquitination with substrate processing at the proteasome entry channel [#17]. Its catalytic HECT domain adopts an open, L-shaped bilobed fold whose C-terminal residues and N-lobe loops are required for E2-E3 transthiolation and activity, with Lys903 as the major autoubiquitination site [#3]. UBE3C responds acutely to protein misfolding, being recruited to the proteasome and ubiquitinating the ubiquitin receptors RPN10 and RPN13 upon unfolding stress [#2], and it clears low-level misfolded oligomers through a proteasomal route distinct from NRF1-dependent proteasome upregulation [#12]; it also contributes to ER-associated degradation of misfolded CFTR (\\u0394F508) and ABCB1 [#10]. Beyond canonical degradation, UBE3C assembles atypical branched ubiquitin chains to regulate autophagy: K29/K48-branched chains on VPS34 (reversed by TRABID) drive its proteasomal degradation and suppress autophagosome formation [#4], and K33-branched chains on ATG4B Lys119 inhibit ATG4B activity and LC3 interaction non-degradatively [#8]. UBE3C further targets specific substrates across diverse contexts \\u2014 cyclin B1 during estrogen/ER\\u03b1-stimulated mitosis [#5], progesterone receptor under P38\\u03b1-restrained control important for uterine receptivity [#7], AXIN1 to activate Wnt/\\u03b2-catenin signaling [#6], IRF3 via K48 chains to resolve antiviral responses [#9], picornaviral 2C protein via K33/K48 chains to restrict viral replication [#11], and mLST8 (non-degradatively, recruited by the SLAP adaptor) to limit mTORC2-AKT signaling [#15]. P38\\u03b1-mediated phosphorylation at Ser741 restrains UBE3C ligase activity [#7].\"\n,\n  \"teleology\": [\n    {\n      \"year\": 2013,\n      \"claim\": \"Established UBE3C's core cellular role as a proteasome-associated processivity factor, answering why some substrates escape complete degradation.\",\n      \"evidence\": \"Forward genetic screen, siRNA knockdown, active-site and proteasome-binding mutagenesis, and substrate lysine-less mutants with polyubiquitination assays\",\n      \"pmids\": [\"24158444\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define which proteasome subunit anchors UBE3C\", \"Chain linkage type used at the proteasome not resolved\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Showed UBE3C association with the proteasome is substrate-driven and cooperative with USP14, explaining how its recruitment is gated.\",\n      \"evidence\": \"Biochemical fractionation and purified proteasome binding assays with recombinant USP14, plus inhibitor and cell-based blockade experiments\",\n      \"pmids\": [\"28396413\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular basis of the USP14\\u2013UBE3C cooperativity not structurally defined\", \"Whether deubiquitination and ubiquitination are spatially coordinated unresolved at the time\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Demonstrated UBE3C is an immediate responder to soluble misfolded protein, ubiquitinating proteasomal ubiquitin receptors RPN10/RPN13 as part of the response.\",\n      \"evidence\": \"Global K-GG ubiquitin proteomics with an inducible FKBP destabilizing-domain misfolding system and quantitative MS\",\n      \"pmids\": [\"31375563\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional consequence of RPN10/RPN13 ubiquitination for proteasome function not fully defined\", \"Distinction between monomer/oligomer and aggregate substrates not yet established\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Provided the structural framework for UBE3C catalysis, defining the HECT fold, autoubiquitination site, and residues required for transthiolation.\",\n      \"evidence\": \"2.7 \\u00c5 X-ray crystallography of the HECT domain with in vitro ubiquitination, site-directed mutagenesis, and deletion analysis\",\n      \"pmids\": [\"32039437\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structure of full-length protein and substrate-bound complexes not solved\", \"Structural basis of branched-chain specificity not addressed\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Revealed UBE3C builds atypical K29/K48-branched chains on VPS34 to suppress autophagy and that proteotoxic stress redistributes UBE3C between phagophores and proteasomes, linking proteostasis to autophagy regulation.\",\n      \"evidence\": \"Co-IP, ubiquitin chain-linkage MS, in vitro ubiquitination, knockdown, live-cell imaging, and in vivo liver metabolic studies\",\n      \"pmids\": [\"33637724\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Signals controlling UBE3C subcellular redistribution not fully defined\", \"How TRABID is targeted to VPS34 chains unclear\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Identified UBE3C as an ER\\u03b1-stimulated mitotic ligase for cyclin B1 and as a Wnt-pathway regulator via AXIN1 degradation, extending its substrate repertoire to cell-cycle and signaling control.\",\n      \"evidence\": \"Co-IP from mitotic MCF-7 cells, in vitro E3 assays, immunofluorescence co-localization, knockdown, and xenograft models\",\n      \"pmids\": [\"26389696\", \"32930707\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Ubiquitin linkage types on cyclin B1 and AXIN1 not mapped\", \"Findings each from a single lab without reciprocal validation\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Established physiological regulation of UBE3C by P38\\u03b1 phosphorylation at Ser741 and its in vivo importance, controlling progesterone receptor stability for uterine receptivity.\",\n      \"evidence\": \"Conditional knockout mice, Co-IP, ubiquitination assays, phosphorylation-site mutagenesis, and genetic rescue\",\n      \"pmids\": [\"35914132\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether Ser741 phosphorylation regulates UBE3C activity toward other substrates not tested\", \"Mechanism by which phosphorylation restrains catalysis not structurally defined\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Showed UBE3C can act non-degradatively, assembling K33-branched chains on ATG4B to inhibit its enzymatic activity, broadening the functional output of its ligase activity.\",\n      \"evidence\": \"MS site identification, Co-IP, in vitro ubiquitination, K119R mutagenesis rescue, and autophagy flux assays\",\n      \"pmids\": [\"38146933\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Deubiquitinase removing K33 chains during starvation not identified\", \"Signal decreasing the ATG4B\\u2013UBE3C interaction unknown\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Distinguished a UBE3C-dependent proteasomal route for clearing low-level misfolded oligomers from an NRF1-dependent pathway engaged at higher aggregate burden, and excluded autophagy in this turnover.\",\n      \"evidence\": \"Inducible agDD-GFP aggregate system, targeted knockdowns, cryo-electron tomography, and quantitative degradation assays\",\n      \"pmids\": [\"39636856\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How the cell switches between UBE3C- and NRF1-dependent responses not defined\", \"Whether RPN13 ubiquitination plays any role in oligomer clearance contradicted prior model\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Placed UBE3C in antiviral and ERAD contexts, mediating IRF3 K48 degradation to resolve interferon responses, restricting picornaviruses via 2C ubiquitination, and assisting degradation of misfolded CFTR.\",\n      \"evidence\": \"Co-IP, ubiquitination assays, site mutagenesis (2C K268R), recombinant virus and surface biotinylation experiments, knockdown\",\n      \"pmids\": [\"39120972\", \"39212385\", \"38067172\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"IRF3 substrate handoff from BAP1 not reconstituted in vitro\", \"CFTR effect appears largely independent of direct CFTR ubiquitination, mechanism unclear\", \"Each result from a single lab\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Identified adaptor-mediated and developmental roles \\u2014 SLAP-recruited non-degradative mLST8 ubiquitination limiting mTORC2-AKT, and UBE3C control of cortical neural fate via the Cbll1\\u2013m6A axis.\",\n      \"evidence\": \"Co-IP, K86R/K215R mutagenesis, mTORC2 integrity assays, xenografts; brain organoid genetics, proteomic substrate profiling, and METTL3-inhibitor rescue (preprint)\",\n      \"pmids\": [\"41398047\", \"bio_10.1101_2025.04.09.646620\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Disease-variant link to cortical phenotype remains in preprint and unreplicated\", \"How SLAP redirects UBE3C specificity not structurally defined\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Provided structural placement of UBE3C/Hul5 on the 19S regulatory particle above RPN11, proposing coupled ubiquitination and deubiquitination at the proteasome substrate entry channel.\",\n      \"evidence\": \"PhIX-MS crosslinking, cryo-EM, and AlphaFold modeling (preprint)\",\n      \"pmids\": [\"bio_10.1101_2025.07.31.667872\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Preprint, single study not independently replicated\", \"Functional coupling between UBE3C and RPN11 not demonstrated biochemically\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Extended the substrate list to oncogenic and tumor-suppressor proteins (p53, TP73, BRAF V600E, AHNAK) and MYC, implicating UBE3C in cancer phenotypes.\",\n      \"evidence\": \"Co-IP, ubiquitination assays, knockdown/CRISPR, MYC-GFP CRISPR screen with paralog specificity, and pharmacological rescues\",\n      \"pmids\": [\"40553397\", \"37173692\", \"40602747\", \"40696164\", \"41965876\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Ubiquitination sites and chain linkages not mapped for these substrates\", \"Direct vs indirect regulation of MYC not established\", \"Each from a single lab without reciprocal validation\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How UBE3C achieves substrate and chain-linkage specificity \\u2014 selecting among degradative K48 and non-degradative branched K29/K33 chains across its many substrates \\u2014 and how adaptors, phosphorylation, and proteasome positioning collectively dictate outcome remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unifying structural model of substrate-bound UBE3C\", \"Determinants of branched vs linear chain assembly unknown\", \"Rules governing degradative vs non-degradative outcomes undefined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016874\", \"supporting_discovery_ids\": [0, 3, 4, 5, 7, 8, 11, 15]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 4, 8, 11]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [5, 9]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [1, 2]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [0, 1, 2, 12]},\n      {\"term_id\": \"R-HSA-9612973\", \"supporting_discovery_ids\": [4, 8]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [9, 11]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"USP14\", \"VPS34\", \"ATG4B\", \"ESR1\", \"AXIN1\", \"IRF3\", \"mLST8\", \"SLA\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}