{"gene":"OTUD6B","run_date":"2026-06-10T05:19:53","timeline":{"discoveries":[{"year":2011,"finding":"OTUD6B encodes a catalytically active deubiquitinating enzyme; mutation of the conserved catalytic Cys residue abolished its deubiquitinating activity in vitro. Enforced OTUD6B expression in Ba/F3 cells blocked proliferation by arresting cells in G1 phase and downregulated cyclin D2.","method":"In vitro deubiquitinating assay with Cys mutant; cell cycle analysis by flow cytometry; immunoblot for cyclin D2","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 1–2 / Moderate — in vitro activity assay with mutagenesis plus cell-cycle phenotype, single lab","pmids":["21267069"],"is_preprint":false},{"year":2016,"finding":"OTUD6B associates with the translation initiation complex (48S preinitiation complex) and regulates protein synthesis downstream of mTORC1 in NSCLC cells. The two main splice isoforms have opposing roles: OTUD6B-1 (long) is inhibitory to protein synthesis and represses DNA synthesis, while OTUD6B-2 (short) stimulates protein synthesis and promotes DNA synthesis. OTUD6B-2 promotes cyclin D1 translation and regulates c-Myc protein stability.","method":"Co-immunoprecipitation with initiation complex components; [35S]-methionine incorporation; BrdU incorporation; immunoblot; isoform overexpression and knockdown","journal":"Molecular cancer research : MCR","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — multiple orthogonal methods (Co-IP, metabolic labeling, BrdU) in a single lab; isoform-specific mutagenesis not fully described in abstract","pmids":["27864334"],"is_preprint":false},{"year":2017,"finding":"Loss-of-function biallelic variants in OTUD6B in human patients cause reduced incorporation of 19S subunits into 26S proteasomes, decreased chymotrypsin-like proteasome activity, and accumulation of ubiquitin-protein conjugates in peripheral blood mononuclear cells, implicating OTUD6B in proteasome function. Homozygous Otud6b knockout mice were subviable, smaller, and had congenital heart defects.","method":"Proteasome activity assays (chymotrypsin-like activity) and native PAGE of 26S proteasome assembly in patient PBMCs; ubiquitin-conjugate accumulation by immunoblot; knockout mouse model","journal":"American journal of human genetics","confidence":"High","confidence_rationale":"Tier 2 / Strong — biochemical proteasome assays in patient cells plus orthologous mouse KO, replicated across multiple families","pmids":["28343629"],"is_preprint":false},{"year":2020,"finding":"OTUD6B directly interacts with pVHL (von Hippel-Lindau protein), decreases its ubiquitylation and proteasomal degradation, and thereby reduces HIF-1α accumulation in HCC cells under hypoxia. Importantly, this stabilization of pVHL is independent of OTUD6B's deubiquitylase catalytic activity. Mechanistically, OTUD6B couples pVHL with elongin B/C to form a more stable CBC-VHL E3 ligase complex; loss of OTUD6B causes dissociation of this complex and degradation of pVHL by WSB1. HIF-1α transcriptionally induces OTUD6B, forming a negative feedback loop.","method":"Co-immunoprecipitation; ubiquitylation assays; catalytic mutant overexpression; siRNA knockdown; HIF-1α reporter assays; immunoblot; ChIP","journal":"Advanced science (Weinheim, Baden-Wurttemberg, Germany)","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP, ubiquitylation assays, catalytic mutant analysis, and mechanistic pathway placement in single rigorous study","pmids":["32328410"],"is_preprint":false},{"year":2021,"finding":"Zebrafish otud6b (ortholog) interacts with IRF3 and IRF7, and removes TRAF6-mediated K63-linked polyubiquitin chains from IRF3 and IRF7, thereby suppressing IFN-1 antiviral signaling. The OTU catalytic domain is required for this activity. Additionally, otud6b attenuates TBK1 binding to IRF3 and IRF7, impairing their phosphorylation.","method":"Co-immunoprecipitation; ubiquitination assays (K63-specific); OTU domain mutant overexpression; zebrafish knockout survival assay; IFN reporter assays","journal":"Journal of immunology (Baltimore, Md. : 1950)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, ubiquitination assays, and in vivo knockout in zebrafish, single lab; zebrafish ortholog may not fully model human function","pmids":["34183367"],"is_preprint":false},{"year":2022,"finding":"OTUD6B is a bona fide deubiquitylase of LIN28B, a suppressor of microRNA biogenesis. OTUD6B stabilizes LIN28B in a cell-cycle-specific (G1/S) manner, and LIN28B stabilization drives MYC expression at G1/S, allowing rapid S-phase entry. Silencing OTUD6B or LIN28B inhibits multiple myeloma outgrowth in vivo.","method":"DUB siRNA screen; Co-IP; ubiquitination assays; cell cycle synchronization; in vivo xenograft; LIN28B protein stability assays","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, ubiquitination assay, cell-cycle-specific substrate stabilization, and in vivo validation; multiple orthogonal methods","pmids":["36059274"],"is_preprint":false},{"year":2022,"finding":"OTUD6B interacts with wild-type pVHL and most tumor-derived pVHL missense mutants (except pVHL I151T) in ccRCC cells, decreasing their ubiquitylation and proteasomal degradation. OTUD6B depletion enhanced cell migration and HIF-2α levels in a pVHL-dependent manner. The I151 residue of pVHL appears critical for the OTUD6B–pVHL interaction.","method":"Co-immunoprecipitation; ubiquitination assays; siRNA knockdown; migration assays; immunoblot for HIF-2α","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — Co-IP and ubiquitination assays across multiple pVHL mutants, single lab, builds on prior study","pmids":["35110537"],"is_preprint":false},{"year":2023,"finding":"Human OTUD6B stabilizes IRF3 protein by hydrolyzing K33-linked polyubiquitin chains at Lys315 of IRF3, thereby positively regulating IRF3-mediated type I IFN antiviral immune response. This opposes the negative regulatory role reported for zebrafish otud6b, establishing a species-distinct function for the human enzyme at this specific ubiquitin linkage.","method":"Ubiquitination assays (K33-specific); site-directed mutagenesis of IRF3 Lys315; immunoblot for IRF3 stability; OTUD6B overexpression mice challenged with RNA virus","journal":"mBio","confidence":"Medium","confidence_rationale":"Tier 1–2 / Moderate — mutagenesis identifying ubiquitin linkage and acceptor site, plus in vivo mouse model; single lab","pmids":["37650650"],"is_preprint":false},{"year":2023,"finding":"OTUD6B is a potent deubiquitinase of β-TrCP (an E3 ubiquitin ligase), stabilizing β-TrCP protein and thereby suppressing SNAIL levels and ESCC progression via an OTUD6B–β-TrCP–SNAIL axis. All-trans retinoic acid (ATRA) promotes OTUD6B translation to achieve this suppression.","method":"Co-immunoprecipitation; ubiquitination assays; immunoblot; ATRA treatment; ESCC xenograft models; SNAIL and β-TrCP protein stability assays","journal":"Advanced science (Weinheim, Baden-Wurttemberg, Germany)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, ubiquitination assay, and in vivo model, single lab","pmids":["37038094"],"is_preprint":false},{"year":2023,"finding":"MCTS1 interacts specifically with OTUD6B isoform 1 (OTUD6B-1) and enhances OTUD6B-mediated removal of K48-linked ubiquitin chains from LIN28B at G1/S, thereby reducing LIN28B degradation and promoting cyclin D1, cyclin E1, and c-Myc expression and LSCC cell proliferation.","method":"Co-immunoprecipitation; ubiquitination assays (K48-specific); shRNA knockdown; immunoblot; in vivo tumor models","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — Co-IP and K48-specific ubiquitination assay; single lab; confirms OTUD6B-LIN28B axis from prior study","pmids":["37634410"],"is_preprint":false},{"year":2024,"finding":"OTUD6B binds RIPK1, reduces its K48-linked ubiquitination, and increases RIPK1 protein stability in lung adenocarcinoma cells, thereby promoting LUAD cell proliferation and metastasis.","method":"IP mass spectrometry; co-immunoprecipitation; ubiquitination assays; siRNA knockdown; xenograft models","journal":"Biology direct","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — IP-MS substrate identification confirmed by Co-IP and ubiquitination assays, single lab","pmids":["38880876"],"is_preprint":false},{"year":2024,"finding":"OTUD6B promotes cholangiocarcinoma growth through deubiquitination of PTK2 (focal adhesion kinase), stabilizing it and leading to increased STAT3 phosphorylation, thereby driving CCA cell proliferation and cell cycle progression.","method":"Co-immunoprecipitation; ubiquitination assays; immunoblot for p-STAT3; siRNA knockdown; xenograft models; flow cytometry","journal":"Cell biology international","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — Co-IP and ubiquitination assays; single lab; in vivo xenograft confirmation","pmids":["39192576"],"is_preprint":false},{"year":2024,"finding":"OTUD6B mediates a Calpain-1/HIF-1α positive feedback loop in pulmonary arterial hypertension. Increased OTUD6B expression activates HIF-1α, increasing ET-1 and VEGF and causing endothelial injury; inhibiting Calpain-1 reduces the effect of OTUD6B on HIF-1α, and inhibiting HIF-1α reduces Calpain-1 and OTUD6B expression.","method":"Proteomics; siRNA knockdown in vitro; tracheal siOTUD6B infusion in mouse PAH model; Calpain-1 KO mice; immunoblot for HIF-1α, ET-1, VEGF","journal":"Cellular and molecular life sciences : CMLS","confidence":"Low","confidence_rationale":"Tier 3 / Weak — proteomic association and siRNA phenotype but direct deubiquitination of Calpain-1 not demonstrated; single lab","pmids":["38878112"],"is_preprint":false},{"year":2021,"finding":"OTUD6B directly interacts with OTUB1 (another DUB); this protein-protein interaction was detected both in cells by GFP-Trap immunoprecipitation and as a direct in vitro interaction by AlphaScreen assay.","method":"GFP-Trap co-immunoprecipitation; AlphaScreen proximity assay (in vitro direct interaction)","journal":"Methods in molecular biology (Clifton, N.J.)","confidence":"Medium","confidence_rationale":"Tier 1–2 / Moderate — direct in vitro AlphaScreen assay plus cell-based Co-IP; functional consequence not established","pmids":["33421002"],"is_preprint":false},{"year":2025,"finding":"OTUD6B localizes to centrosomes and the mitotic spindle, interacts with the kinesin KIFC1/HSET, and acts as a positive regulator of KIFC1 protein expression by preventing its polyubiquitination and premature proteasomal degradation during mitosis. Loss of OTUD6B increases KIFC1 polyubiquitination, causes multipolar spindles in centrosome-amplified cancer cells, and is lethal in TNBC cells with centrosome amplification. Phenotypic rescue requires OTUD6B catalytic activity.","method":"siRNA screen; co-immunoprecipitation; ubiquitination assays; CRISPR-Cas9 knockout; immunofluorescence/live imaging for centrosome/spindle localization; KIFC1 overexpression rescue; catalytic mutant rescue","journal":"EMBO reports","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — multiple orthogonal methods (Co-IP, ubiquitination assay, catalytic mutant rescue, KIFC1 OE rescue, CRISPR KO, direct localization), single rigorous study","pmids":["39789388"],"is_preprint":false},{"year":2025,"finding":"OTUD6B deubiquitinates and stabilizes DDX5, and this stabilization promotes STAT3 activation via DDX5 resolving the RNA G-quadruplex structure of STAT3, leading to increased CXCL11 transcription and CD8+ T cell recruitment in colorectal liver metastasis. All-trans retinoic acid upregulates OTUD6B to achieve this effect.","method":"Co-immunoprecipitation; ubiquitination assays; ATRA treatment; RNA G-quadruplex assay; immunofluorescence/flow cytometry for CD8+ T cell infiltration; in vivo mouse models (nude and immunocompetent)","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and ubiquitination assays with in vivo mechanistic validation; single lab; DDX5 deubiquitination confirmed","pmids":["40651961"],"is_preprint":false},{"year":2026,"finding":"OTUD6B binds the KH domain of FXR1 via its N-terminal region and removes K48-linked polyubiquitin chains from FXR1 in a catalysis-dependent manner, thereby stabilizing FXR1. Stabilized FXR1 binds and stabilizes MEK2 mRNA, increasing MEK2 expression and activating ERK signaling. FXR1 also upregulates OTUD6B expression, forming a feed-forward oncogenic loop driving CRC liver metastasis.","method":"Co-immunoprecipitation; K48-specific ubiquitination assays; domain mapping; mRNA stability assay for MEK2; immunoblot; in vivo liver metastasis models; catalytic mutant analysis","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — domain-specific Co-IP, K48-ubiquitination assays, catalytic mutant, and in vivo model; single lab","pmids":["42056075"],"is_preprint":false},{"year":2026,"finding":"OTUD6B localizes to stress granules (SGs) and regulates both their early assembly and clearance, partially dependent on its enzymatic activity. Using proximity proteomics and interactomics, OTUD6B was found to associate with the ATPase VCP/p97 through disordered regions involved in biomolecular condensation. OTUD6B promotes coalescence of VCP into SGs, accelerating SG assembly and post-stress clearance; VCP knockdown or inhibition phenocopied OTUD6B silencing.","method":"Proximity proteomics (BioID); co-immunoprecipitation; immunofluorescence; VCP inhibitor/siRNA; catalytic mutant; SG dynamics assays","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — proximity proteomics confirmed by Co-IP and immunofluorescence, catalytic mutant analysis; single lab","pmids":["41651815"],"is_preprint":false}],"current_model":"OTUD6B is a catalytically active OTU-family deubiquitinase with multiple cellular substrates: it stabilizes pVHL (enzyme-independently, by scaffolding the VHL–elongin B/C complex), LIN28B (via K48-deubiquitination to drive MYC/G1-S progression), KIFC1 (protecting it from mitotic degradation to enable centrosome clustering), IRF3 (via K33-deubiquitination to enhance antiviral IFN signaling in humans), β-TrCP, RIPK1, PTK2, DDX5, and FXR1, while its two splice isoforms oppositely regulate 48S preinitiation complex activity and protein synthesis; loss-of-function causes defective 26S proteasome assembly, and OTUD6B also localizes to centrosomes, the mitotic spindle, and stress granules where it modulates VCP-dependent granule dynamics."},"narrative":{"mechanistic_narrative":"OTUD6B is a catalytically active OTU-family deubiquitinase that controls the stability of diverse substrates to regulate cell proliferation, proteasome function, and stress responses [PMID:21267069, PMID:28343629]. Its catalytic cysteine is required for ubiquitin chain hydrolysis, and enforced expression arrests cells in G1 by downregulating cyclin D2 [PMID:21267069]. Biallelic loss-of-function variants in humans impair incorporation of 19S subunits into the 26S proteasome and cause accumulation of ubiquitin-protein conjugates, defining a Mendelian disorder with congenital heart defects recapitulated in Otud6b knockout mice [PMID:28343629]. A recurring mechanistic theme is catalysis-dependent removal of K48-linked polyubiquitin to stabilize pro-proliferative substrates: OTUD6B deubiquitinates LIN28B in a G1/S-restricted manner to drive MYC expression and S-phase entry [PMID:36059274, PMID:37634410], and stabilizes RIPK1, PTK2/FAK, and FXR1 to promote tumor proliferation and metastasis through downstream STAT3 and ERK signaling [PMID:38880876, PMID:39192576, PMID:42056075]. Beyond catalysis, OTUD6B can act as a scaffold: it stabilizes pVHL independently of its enzymatic activity by coupling pVHL with elongin B/C into a more stable CBC-VHL E3 ligase complex, thereby restraining HIF accumulation [PMID:32328410, PMID:35110537]. OTUD6B also localizes to centrosomes and the mitotic spindle, where it protects the kinesin KIFC1/HSET from premature mitotic degradation to enable bipolar spindle formation in centrosome-amplified cancer cells [PMID:39789388], and to stress granules, where it associates with the ATPase VCP/p97 to modulate granule assembly and clearance [PMID:41651815]. Two splice isoforms exert opposing control over the 48S preinitiation complex and protein synthesis downstream of mTORC1 [PMID:27864334].","teleology":[{"year":2011,"claim":"Established that OTUD6B is an enzymatically active deubiquitinase and linked its activity to cell-cycle control, defining the core biochemical identity of the protein.","evidence":"In vitro deubiquitinating assay with catalytic Cys mutant plus flow-cytometry cell-cycle analysis in Ba/F3 cells","pmids":["21267069"],"confidence":"Medium","gaps":["No physiological substrate identified at this stage","Mechanism linking DUB activity to cyclin D2 downregulation unresolved"]},{"year":2016,"claim":"Connected OTUD6B to translational control, showing its two splice isoforms oppositely regulate the 48S preinitiation complex and protein synthesis downstream of mTORC1.","evidence":"Co-IP with initiation complex components, [35S]-methionine and BrdU incorporation, isoform overexpression/knockdown in NSCLC cells","pmids":["27864334"],"confidence":"Medium","gaps":["Direct deubiquitination target within the initiation machinery not defined","Structural basis of isoform-specific opposing activity unknown"]},{"year":2017,"claim":"Defined OTUD6B as a human disease gene and tied it to proteasome biogenesis, showing loss-of-function impairs 26S assembly.","evidence":"Proteasome activity assays and native PAGE in patient PBMCs, ubiquitin-conjugate immunoblot, and Otud6b knockout mouse model","pmids":["28343629"],"confidence":"High","gaps":["Molecular mechanism by which OTUD6B promotes 19S incorporation not established","Substrate responsible for proteasome assembly phenotype unidentified"]},{"year":2020,"claim":"Revealed a catalysis-independent scaffolding function, showing OTUD6B stabilizes pVHL by assembling the CBC-VHL E3 complex, expanding its mechanistic repertoire beyond deubiquitination.","evidence":"Reciprocal Co-IP, ubiquitylation assays, catalytic mutant analysis, HIF-1α reporter and ChIP in HCC cells","pmids":["32328410"],"confidence":"High","gaps":["Structural detail of how OTUD6B couples pVHL to elongin B/C not resolved","Relationship between scaffolding and catalytic substrate pools unclear"]},{"year":2021,"claim":"Identified a direct OTUD6B–OTUB1 DUB-DUB interaction and, in zebrafish, an antiviral signaling role, broadening the partner and pathway landscape.","evidence":"GFP-Trap Co-IP and AlphaScreen for OTUB1; Co-IP, K63-specific ubiquitination assays and zebrafish knockout for IRF3/IRF7 regulation","pmids":["33421002","34183367"],"confidence":"Medium","gaps":["Functional consequence of OTUB1 interaction not established","Zebrafish ortholog function (negative IFN regulation) conflicts with later human data"]},{"year":2022,"claim":"Defined a cell-cycle-restricted substrate, LIN28B, establishing how OTUD6B drives MYC-dependent S-phase entry, and confirmed pVHL stabilization across tumor-derived mutants.","evidence":"DUB siRNA screen, reciprocal Co-IP, ubiquitination assays, cell-cycle synchronization and xenografts for LIN28B; Co-IP and migration assays across pVHL mutants in ccRCC","pmids":["36059274","35110537"],"confidence":"High","gaps":["Mechanism restricting LIN28B deubiquitination to G1/S not defined","Determinants of the pVHL I151 dependence for binding unresolved"]},{"year":2023,"claim":"Resolved a species-distinct antiviral role and expanded the catalytic substrate set, showing human OTUD6B cleaves K33-linked chains on IRF3 and K48 chains on β-TrCP and LIN28B (with MCTS1 as a coactivator).","evidence":"Linkage- and site-specific ubiquitination assays with IRF3 Lys315 mutagenesis and in vivo mouse virus challenge; Co-IP and ATRA studies for β-TrCP; MCTS1 Co-IP and K48 assays for LIN28B","pmids":["37650650","37038094","37634410"],"confidence":"Medium","gaps":["Reason for opposing human vs zebrafish IFN regulation mechanistically unexplained","How OTUD6B selects among K33 and K48 linkages on different substrates unknown"]},{"year":2024,"claim":"Extended the substrate catalog to RIPK1 and PTK2/FAK in cancer and implicated OTUD6B in a Calpain-1/HIF-1α loop, linking it to proliferation, metastasis, and vascular pathology.","evidence":"IP-MS, Co-IP and ubiquitination assays with xenografts for RIPK1 and PTK2; proteomics and siRNA in a mouse PAH model with Calpain-1 KO","pmids":["38880876","39192576","38878112"],"confidence":"Medium","gaps":["Direct deubiquitination of Calpain-1 not demonstrated","Whether RIPK1 and PTK2 stabilization share a common recognition mechanism unknown"]},{"year":2025,"claim":"Localized OTUD6B to centrosomes and the mitotic spindle and to stress granules, establishing roles in spindle integrity via KIFC1 and in condensate dynamics via VCP.","evidence":"siRNA screen, Co-IP, ubiquitination assays, CRISPR KO, catalytic-mutant and KIFC1 overexpression rescue with imaging; proximity proteomics and Co-IP with VCP plus SG dynamics assays; Co-IP and G-quadruplex assays for DDX5","pmids":["39789388","41651815","40651961"],"confidence":"High","gaps":["How OTUD6B is recruited to centrosomes vs stress granules not defined","Whether the proteasome-assembly defect connects to centrosome/spindle phenotypes unexplored"]},{"year":2026,"claim":"Defined FXR1 as a domain-specific K48 substrate, linking OTUD6B to an FXR1–MEK2–ERK feed-forward oncogenic loop in colorectal liver metastasis.","evidence":"Domain-mapping Co-IP, K48-specific ubiquitination assays, MEK2 mRNA stability assays, catalytic mutant and in vivo liver metastasis models","pmids":["42056075"],"confidence":"Medium","gaps":["Generalizability of the N-terminal/KH-domain recognition mode to other substrates untested","Direct structural basis of OTUD6B–FXR1 binding unresolved"]},{"year":null,"claim":"How OTUD6B achieves substrate and linkage selectivity (K48 vs K33), and how its catalytic, scaffolding, translational, and condensate functions are coordinated within a single protein, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structural model of substrate or linkage recognition","Mechanism integrating proteasome-assembly, spindle, and stress-granule roles unknown","Determinants switching between catalytic and scaffold-only modes undefined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,3,5,8,10,11,14,15,16]},{"term_id":"GO:0016787","term_label":"hydrolase activity","supporting_discovery_ids":[0,4,7]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[3]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[2]}],"localization":[{"term_id":"GO:0005815","term_label":"microtubule organizing center","supporting_discovery_ids":[14]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[14]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[17]}],"pathway":[{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[0,5,14]},{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[2,3]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[4,7]},{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[1,16]},{"term_id":"R-HSA-8953897","term_label":"Cellular responses to stimuli","supporting_discovery_ids":[17]}],"complexes":["CBC-VHL E3 ligase complex","48S preinitiation complex"],"partners":["VHL","LIN28B","KIFC1","IRF3","BTRC","RIPK1","DDX5","FXR1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q8N6M0","full_name":"Deubiquitinase OTUD6B","aliases":["DUBA-5","OTU domain-containing protein 6B"],"length_aa":293,"mass_kda":33.8,"function":"Deubiquitinating enzyme that may play a role in the ubiquitin-dependent regulation of protein synthesis, downstream of mTORC1 (PubMed:21267069, PubMed:27864334). May associate with the protein synthesis initiation complex and modify its ubiquitination to repress translation (PubMed:27864334). May also repress DNA synthesis and modify different cellular targets thereby regulating cell growth and proliferation (PubMed:27864334). May also play a role in proteasome assembly and function (PubMed:28343629) Stimulates protein synthesis. Influences the expression of CCND1/cyclin D1 by promoting its translation and regulates MYC/c-Myc protein stability","subcellular_location":"","url":"https://www.uniprot.org/uniprotkb/Q8N6M0/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/OTUD6B","classification":"Not Classified","n_dependent_lines":9,"n_total_lines":1208,"dependency_fraction":0.0074503311258278145},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/OTUD6B","total_profiled":1310},"omim":[{"mim_id":"617452","title":"INTELLECTUAL DEVELOPMENTAL DISORDER WITH DYSMORPHIC FACIES, SEIZURES, AND DISTAL LIMB ANOMALIES; IDDFSDA","url":"https://www.omim.org/entry/617452"},{"mim_id":"612021","title":"OTU DOMAIN-CONTAINING PROTEIN 6B; OTUD6B","url":"https://www.omim.org/entry/612021"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Nucleoplasm","reliability":"Approved"},{"location":"Golgi apparatus","reliability":"Approved"},{"location":"Cytosol","reliability":"Approved"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/OTUD6B"},"hgnc":{"alias_symbol":["CGI-77","DUBA5"],"prev_symbol":[]},"alphafold":{"accession":"Q8N6M0","domains":[{"cath_id":"3.90.70.80","chopping":"132-284","consensus_level":"high","plddt":94.8455,"start":132,"end":284},{"cath_id":"1.10.287","chopping":"12-73","consensus_level":"medium","plddt":88.04,"start":12,"end":73}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8N6M0","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q8N6M0-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q8N6M0-F1-predicted_aligned_error_v6.png","plddt_mean":84.88},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=OTUD6B","jax_strain_url":"https://www.jax.org/strain/search?query=OTUD6B"},"sequence":{"accession":"Q8N6M0","fasta_url":"https://rest.uniprot.org/uniprotkb/Q8N6M0.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q8N6M0/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8N6M0"}},"corpus_meta":[{"pmid":"34740994","id":"PMC_34740994","title":"LncRNA OTUD6B-AS1 promotes paclitaxel resistance in triple negative breast cancer by regulation of miR-26a-5p/MTDH pathway-mediated autophagy and genomic instability.","date":"2021","source":"Aging","url":"https://pubmed.ncbi.nlm.nih.gov/34740994","citation_count":62,"is_preprint":false},{"pmid":"28343629","id":"PMC_28343629","title":"Biallelic Variants in OTUD6B Cause an Intellectual Disability Syndrome Associated with Seizures and Dysmorphic Features.","date":"2017","source":"American journal of human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/28343629","citation_count":54,"is_preprint":false},{"pmid":"32328410","id":"PMC_32328410","title":"Deubiquitylase OTUD6B Governs pVHL Stability in an Enzyme-Independent Manner and Suppresses Hepatocellular Carcinoma Metastasis.","date":"2020","source":"Advanced science (Weinheim, Baden-Wurttemberg, 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Bio","url":"https://pubmed.ncbi.nlm.nih.gov/41909231","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":21087,"output_tokens":5042,"usd":0.069445,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":13327,"output_tokens":4123,"usd":0.084855,"stage2_stop_reason":"end_turn"},"total_usd":0.1543,"stage1_batch_id":"msgbatch_013a8chtv3AWyTqv4z4sEF4C","stage2_batch_id":"msgbatch_01AHsuW63vinsvuTgzmQ3n62","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2011,\n      \"finding\": \"OTUD6B encodes a catalytically active deubiquitinating enzyme; mutation of the conserved catalytic Cys residue abolished its deubiquitinating activity in vitro. Enforced OTUD6B expression in Ba/F3 cells blocked proliferation by arresting cells in G1 phase and downregulated cyclin D2.\",\n      \"method\": \"In vitro deubiquitinating assay with Cys mutant; cell cycle analysis by flow cytometry; immunoblot for cyclin D2\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — in vitro activity assay with mutagenesis plus cell-cycle phenotype, single lab\",\n      \"pmids\": [\"21267069\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"OTUD6B associates with the translation initiation complex (48S preinitiation complex) and regulates protein synthesis downstream of mTORC1 in NSCLC cells. The two main splice isoforms have opposing roles: OTUD6B-1 (long) is inhibitory to protein synthesis and represses DNA synthesis, while OTUD6B-2 (short) stimulates protein synthesis and promotes DNA synthesis. OTUD6B-2 promotes cyclin D1 translation and regulates c-Myc protein stability.\",\n      \"method\": \"Co-immunoprecipitation with initiation complex components; [35S]-methionine incorporation; BrdU incorporation; immunoblot; isoform overexpression and knockdown\",\n      \"journal\": \"Molecular cancer research : MCR\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — multiple orthogonal methods (Co-IP, metabolic labeling, BrdU) in a single lab; isoform-specific mutagenesis not fully described in abstract\",\n      \"pmids\": [\"27864334\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Loss-of-function biallelic variants in OTUD6B in human patients cause reduced incorporation of 19S subunits into 26S proteasomes, decreased chymotrypsin-like proteasome activity, and accumulation of ubiquitin-protein conjugates in peripheral blood mononuclear cells, implicating OTUD6B in proteasome function. Homozygous Otud6b knockout mice were subviable, smaller, and had congenital heart defects.\",\n      \"method\": \"Proteasome activity assays (chymotrypsin-like activity) and native PAGE of 26S proteasome assembly in patient PBMCs; ubiquitin-conjugate accumulation by immunoblot; knockout mouse model\",\n      \"journal\": \"American journal of human genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — biochemical proteasome assays in patient cells plus orthologous mouse KO, replicated across multiple families\",\n      \"pmids\": [\"28343629\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"OTUD6B directly interacts with pVHL (von Hippel-Lindau protein), decreases its ubiquitylation and proteasomal degradation, and thereby reduces HIF-1α accumulation in HCC cells under hypoxia. Importantly, this stabilization of pVHL is independent of OTUD6B's deubiquitylase catalytic activity. Mechanistically, OTUD6B couples pVHL with elongin B/C to form a more stable CBC-VHL E3 ligase complex; loss of OTUD6B causes dissociation of this complex and degradation of pVHL by WSB1. HIF-1α transcriptionally induces OTUD6B, forming a negative feedback loop.\",\n      \"method\": \"Co-immunoprecipitation; ubiquitylation assays; catalytic mutant overexpression; siRNA knockdown; HIF-1α reporter assays; immunoblot; ChIP\",\n      \"journal\": \"Advanced science (Weinheim, Baden-Wurttemberg, Germany)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP, ubiquitylation assays, catalytic mutant analysis, and mechanistic pathway placement in single rigorous study\",\n      \"pmids\": [\"32328410\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Zebrafish otud6b (ortholog) interacts with IRF3 and IRF7, and removes TRAF6-mediated K63-linked polyubiquitin chains from IRF3 and IRF7, thereby suppressing IFN-1 antiviral signaling. The OTU catalytic domain is required for this activity. Additionally, otud6b attenuates TBK1 binding to IRF3 and IRF7, impairing their phosphorylation.\",\n      \"method\": \"Co-immunoprecipitation; ubiquitination assays (K63-specific); OTU domain mutant overexpression; zebrafish knockout survival assay; IFN reporter assays\",\n      \"journal\": \"Journal of immunology (Baltimore, Md. : 1950)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, ubiquitination assays, and in vivo knockout in zebrafish, single lab; zebrafish ortholog may not fully model human function\",\n      \"pmids\": [\"34183367\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"OTUD6B is a bona fide deubiquitylase of LIN28B, a suppressor of microRNA biogenesis. OTUD6B stabilizes LIN28B in a cell-cycle-specific (G1/S) manner, and LIN28B stabilization drives MYC expression at G1/S, allowing rapid S-phase entry. Silencing OTUD6B or LIN28B inhibits multiple myeloma outgrowth in vivo.\",\n      \"method\": \"DUB siRNA screen; Co-IP; ubiquitination assays; cell cycle synchronization; in vivo xenograft; LIN28B protein stability assays\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, ubiquitination assay, cell-cycle-specific substrate stabilization, and in vivo validation; multiple orthogonal methods\",\n      \"pmids\": [\"36059274\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"OTUD6B interacts with wild-type pVHL and most tumor-derived pVHL missense mutants (except pVHL I151T) in ccRCC cells, decreasing their ubiquitylation and proteasomal degradation. OTUD6B depletion enhanced cell migration and HIF-2α levels in a pVHL-dependent manner. The I151 residue of pVHL appears critical for the OTUD6B–pVHL interaction.\",\n      \"method\": \"Co-immunoprecipitation; ubiquitination assays; siRNA knockdown; migration assays; immunoblot for HIF-2α\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — Co-IP and ubiquitination assays across multiple pVHL mutants, single lab, builds on prior study\",\n      \"pmids\": [\"35110537\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Human OTUD6B stabilizes IRF3 protein by hydrolyzing K33-linked polyubiquitin chains at Lys315 of IRF3, thereby positively regulating IRF3-mediated type I IFN antiviral immune response. This opposes the negative regulatory role reported for zebrafish otud6b, establishing a species-distinct function for the human enzyme at this specific ubiquitin linkage.\",\n      \"method\": \"Ubiquitination assays (K33-specific); site-directed mutagenesis of IRF3 Lys315; immunoblot for IRF3 stability; OTUD6B overexpression mice challenged with RNA virus\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — mutagenesis identifying ubiquitin linkage and acceptor site, plus in vivo mouse model; single lab\",\n      \"pmids\": [\"37650650\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"OTUD6B is a potent deubiquitinase of β-TrCP (an E3 ubiquitin ligase), stabilizing β-TrCP protein and thereby suppressing SNAIL levels and ESCC progression via an OTUD6B–β-TrCP–SNAIL axis. All-trans retinoic acid (ATRA) promotes OTUD6B translation to achieve this suppression.\",\n      \"method\": \"Co-immunoprecipitation; ubiquitination assays; immunoblot; ATRA treatment; ESCC xenograft models; SNAIL and β-TrCP protein stability assays\",\n      \"journal\": \"Advanced science (Weinheim, Baden-Wurttemberg, Germany)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, ubiquitination assay, and in vivo model, single lab\",\n      \"pmids\": [\"37038094\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"MCTS1 interacts specifically with OTUD6B isoform 1 (OTUD6B-1) and enhances OTUD6B-mediated removal of K48-linked ubiquitin chains from LIN28B at G1/S, thereby reducing LIN28B degradation and promoting cyclin D1, cyclin E1, and c-Myc expression and LSCC cell proliferation.\",\n      \"method\": \"Co-immunoprecipitation; ubiquitination assays (K48-specific); shRNA knockdown; immunoblot; in vivo tumor models\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — Co-IP and K48-specific ubiquitination assay; single lab; confirms OTUD6B-LIN28B axis from prior study\",\n      \"pmids\": [\"37634410\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"OTUD6B binds RIPK1, reduces its K48-linked ubiquitination, and increases RIPK1 protein stability in lung adenocarcinoma cells, thereby promoting LUAD cell proliferation and metastasis.\",\n      \"method\": \"IP mass spectrometry; co-immunoprecipitation; ubiquitination assays; siRNA knockdown; xenograft models\",\n      \"journal\": \"Biology direct\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — IP-MS substrate identification confirmed by Co-IP and ubiquitination assays, single lab\",\n      \"pmids\": [\"38880876\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"OTUD6B promotes cholangiocarcinoma growth through deubiquitination of PTK2 (focal adhesion kinase), stabilizing it and leading to increased STAT3 phosphorylation, thereby driving CCA cell proliferation and cell cycle progression.\",\n      \"method\": \"Co-immunoprecipitation; ubiquitination assays; immunoblot for p-STAT3; siRNA knockdown; xenograft models; flow cytometry\",\n      \"journal\": \"Cell biology international\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — Co-IP and ubiquitination assays; single lab; in vivo xenograft confirmation\",\n      \"pmids\": [\"39192576\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"OTUD6B mediates a Calpain-1/HIF-1α positive feedback loop in pulmonary arterial hypertension. Increased OTUD6B expression activates HIF-1α, increasing ET-1 and VEGF and causing endothelial injury; inhibiting Calpain-1 reduces the effect of OTUD6B on HIF-1α, and inhibiting HIF-1α reduces Calpain-1 and OTUD6B expression.\",\n      \"method\": \"Proteomics; siRNA knockdown in vitro; tracheal siOTUD6B infusion in mouse PAH model; Calpain-1 KO mice; immunoblot for HIF-1α, ET-1, VEGF\",\n      \"journal\": \"Cellular and molecular life sciences : CMLS\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — proteomic association and siRNA phenotype but direct deubiquitination of Calpain-1 not demonstrated; single lab\",\n      \"pmids\": [\"38878112\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"OTUD6B directly interacts with OTUB1 (another DUB); this protein-protein interaction was detected both in cells by GFP-Trap immunoprecipitation and as a direct in vitro interaction by AlphaScreen assay.\",\n      \"method\": \"GFP-Trap co-immunoprecipitation; AlphaScreen proximity assay (in vitro direct interaction)\",\n      \"journal\": \"Methods in molecular biology (Clifton, N.J.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — direct in vitro AlphaScreen assay plus cell-based Co-IP; functional consequence not established\",\n      \"pmids\": [\"33421002\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"OTUD6B localizes to centrosomes and the mitotic spindle, interacts with the kinesin KIFC1/HSET, and acts as a positive regulator of KIFC1 protein expression by preventing its polyubiquitination and premature proteasomal degradation during mitosis. Loss of OTUD6B increases KIFC1 polyubiquitination, causes multipolar spindles in centrosome-amplified cancer cells, and is lethal in TNBC cells with centrosome amplification. Phenotypic rescue requires OTUD6B catalytic activity.\",\n      \"method\": \"siRNA screen; co-immunoprecipitation; ubiquitination assays; CRISPR-Cas9 knockout; immunofluorescence/live imaging for centrosome/spindle localization; KIFC1 overexpression rescue; catalytic mutant rescue\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — multiple orthogonal methods (Co-IP, ubiquitination assay, catalytic mutant rescue, KIFC1 OE rescue, CRISPR KO, direct localization), single rigorous study\",\n      \"pmids\": [\"39789388\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"OTUD6B deubiquitinates and stabilizes DDX5, and this stabilization promotes STAT3 activation via DDX5 resolving the RNA G-quadruplex structure of STAT3, leading to increased CXCL11 transcription and CD8+ T cell recruitment in colorectal liver metastasis. All-trans retinoic acid upregulates OTUD6B to achieve this effect.\",\n      \"method\": \"Co-immunoprecipitation; ubiquitination assays; ATRA treatment; RNA G-quadruplex assay; immunofluorescence/flow cytometry for CD8+ T cell infiltration; in vivo mouse models (nude and immunocompetent)\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and ubiquitination assays with in vivo mechanistic validation; single lab; DDX5 deubiquitination confirmed\",\n      \"pmids\": [\"40651961\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"OTUD6B binds the KH domain of FXR1 via its N-terminal region and removes K48-linked polyubiquitin chains from FXR1 in a catalysis-dependent manner, thereby stabilizing FXR1. Stabilized FXR1 binds and stabilizes MEK2 mRNA, increasing MEK2 expression and activating ERK signaling. FXR1 also upregulates OTUD6B expression, forming a feed-forward oncogenic loop driving CRC liver metastasis.\",\n      \"method\": \"Co-immunoprecipitation; K48-specific ubiquitination assays; domain mapping; mRNA stability assay for MEK2; immunoblot; in vivo liver metastasis models; catalytic mutant analysis\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — domain-specific Co-IP, K48-ubiquitination assays, catalytic mutant, and in vivo model; single lab\",\n      \"pmids\": [\"42056075\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"OTUD6B localizes to stress granules (SGs) and regulates both their early assembly and clearance, partially dependent on its enzymatic activity. Using proximity proteomics and interactomics, OTUD6B was found to associate with the ATPase VCP/p97 through disordered regions involved in biomolecular condensation. OTUD6B promotes coalescence of VCP into SGs, accelerating SG assembly and post-stress clearance; VCP knockdown or inhibition phenocopied OTUD6B silencing.\",\n      \"method\": \"Proximity proteomics (BioID); co-immunoprecipitation; immunofluorescence; VCP inhibitor/siRNA; catalytic mutant; SG dynamics assays\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — proximity proteomics confirmed by Co-IP and immunofluorescence, catalytic mutant analysis; single lab\",\n      \"pmids\": [\"41651815\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"OTUD6B is a catalytically active OTU-family deubiquitinase with multiple cellular substrates: it stabilizes pVHL (enzyme-independently, by scaffolding the VHL–elongin B/C complex), LIN28B (via K48-deubiquitination to drive MYC/G1-S progression), KIFC1 (protecting it from mitotic degradation to enable centrosome clustering), IRF3 (via K33-deubiquitination to enhance antiviral IFN signaling in humans), β-TrCP, RIPK1, PTK2, DDX5, and FXR1, while its two splice isoforms oppositely regulate 48S preinitiation complex activity and protein synthesis; loss-of-function causes defective 26S proteasome assembly, and OTUD6B also localizes to centrosomes, the mitotic spindle, and stress granules where it modulates VCP-dependent granule dynamics.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"OTUD6B is a catalytically active OTU-family deubiquitinase that controls the stability of diverse substrates to regulate cell proliferation, proteasome function, and stress responses [#0, #2]. Its catalytic cysteine is required for ubiquitin chain hydrolysis, and enforced expression arrests cells in G1 by downregulating cyclin D2 [#0]. Biallelic loss-of-function variants in humans impair incorporation of 19S subunits into the 26S proteasome and cause accumulation of ubiquitin-protein conjugates, defining a Mendelian disorder with congenital heart defects recapitulated in Otud6b knockout mice [#2]. A recurring mechanistic theme is catalysis-dependent removal of K48-linked polyubiquitin to stabilize pro-proliferative substrates: OTUD6B deubiquitinates LIN28B in a G1/S-restricted manner to drive MYC expression and S-phase entry [#5, #9], and stabilizes RIPK1, PTK2/FAK, and FXR1 to promote tumor proliferation and metastasis through downstream STAT3 and ERK signaling [#10, #11, #16]. Beyond catalysis, OTUD6B can act as a scaffold: it stabilizes pVHL independently of its enzymatic activity by coupling pVHL with elongin B/C into a more stable CBC-VHL E3 ligase complex, thereby restraining HIF accumulation [#3, #6]. OTUD6B also localizes to centrosomes and the mitotic spindle, where it protects the kinesin KIFC1/HSET from premature mitotic degradation to enable bipolar spindle formation in centrosome-amplified cancer cells [#14], and to stress granules, where it associates with the ATPase VCP/p97 to modulate granule assembly and clearance [#17]. Two splice isoforms exert opposing control over the 48S preinitiation complex and protein synthesis downstream of mTORC1 [#1].\",\n  \"teleology\": [\n    {\n      \"year\": 2011,\n      \"claim\": \"Established that OTUD6B is an enzymatically active deubiquitinase and linked its activity to cell-cycle control, defining the core biochemical identity of the protein.\",\n      \"evidence\": \"In vitro deubiquitinating assay with catalytic Cys mutant plus flow-cytometry cell-cycle analysis in Ba/F3 cells\",\n      \"pmids\": [\"21267069\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No physiological substrate identified at this stage\", \"Mechanism linking DUB activity to cyclin D2 downregulation unresolved\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Connected OTUD6B to translational control, showing its two splice isoforms oppositely regulate the 48S preinitiation complex and protein synthesis downstream of mTORC1.\",\n      \"evidence\": \"Co-IP with initiation complex components, [35S]-methionine and BrdU incorporation, isoform overexpression/knockdown in NSCLC cells\",\n      \"pmids\": [\"27864334\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct deubiquitination target within the initiation machinery not defined\", \"Structural basis of isoform-specific opposing activity unknown\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Defined OTUD6B as a human disease gene and tied it to proteasome biogenesis, showing loss-of-function impairs 26S assembly.\",\n      \"evidence\": \"Proteasome activity assays and native PAGE in patient PBMCs, ubiquitin-conjugate immunoblot, and Otud6b knockout mouse model\",\n      \"pmids\": [\"28343629\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular mechanism by which OTUD6B promotes 19S incorporation not established\", \"Substrate responsible for proteasome assembly phenotype unidentified\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Revealed a catalysis-independent scaffolding function, showing OTUD6B stabilizes pVHL by assembling the CBC-VHL E3 complex, expanding its mechanistic repertoire beyond deubiquitination.\",\n      \"evidence\": \"Reciprocal Co-IP, ubiquitylation assays, catalytic mutant analysis, HIF-1α reporter and ChIP in HCC cells\",\n      \"pmids\": [\"32328410\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural detail of how OTUD6B couples pVHL to elongin B/C not resolved\", \"Relationship between scaffolding and catalytic substrate pools unclear\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Identified a direct OTUD6B–OTUB1 DUB-DUB interaction and, in zebrafish, an antiviral signaling role, broadening the partner and pathway landscape.\",\n      \"evidence\": \"GFP-Trap Co-IP and AlphaScreen for OTUB1; Co-IP, K63-specific ubiquitination assays and zebrafish knockout for IRF3/IRF7 regulation\",\n      \"pmids\": [\"33421002\", \"34183367\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional consequence of OTUB1 interaction not established\", \"Zebrafish ortholog function (negative IFN regulation) conflicts with later human data\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Defined a cell-cycle-restricted substrate, LIN28B, establishing how OTUD6B drives MYC-dependent S-phase entry, and confirmed pVHL stabilization across tumor-derived mutants.\",\n      \"evidence\": \"DUB siRNA screen, reciprocal Co-IP, ubiquitination assays, cell-cycle synchronization and xenografts for LIN28B; Co-IP and migration assays across pVHL mutants in ccRCC\",\n      \"pmids\": [\"36059274\", \"35110537\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism restricting LIN28B deubiquitination to G1/S not defined\", \"Determinants of the pVHL I151 dependence for binding unresolved\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Resolved a species-distinct antiviral role and expanded the catalytic substrate set, showing human OTUD6B cleaves K33-linked chains on IRF3 and K48 chains on β-TrCP and LIN28B (with MCTS1 as a coactivator).\",\n      \"evidence\": \"Linkage- and site-specific ubiquitination assays with IRF3 Lys315 mutagenesis and in vivo mouse virus challenge; Co-IP and ATRA studies for β-TrCP; MCTS1 Co-IP and K48 assays for LIN28B\",\n      \"pmids\": [\"37650650\", \"37038094\", \"37634410\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Reason for opposing human vs zebrafish IFN regulation mechanistically unexplained\", \"How OTUD6B selects among K33 and K48 linkages on different substrates unknown\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Extended the substrate catalog to RIPK1 and PTK2/FAK in cancer and implicated OTUD6B in a Calpain-1/HIF-1α loop, linking it to proliferation, metastasis, and vascular pathology.\",\n      \"evidence\": \"IP-MS, Co-IP and ubiquitination assays with xenografts for RIPK1 and PTK2; proteomics and siRNA in a mouse PAH model with Calpain-1 KO\",\n      \"pmids\": [\"38880876\", \"39192576\", \"38878112\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct deubiquitination of Calpain-1 not demonstrated\", \"Whether RIPK1 and PTK2 stabilization share a common recognition mechanism unknown\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Localized OTUD6B to centrosomes and the mitotic spindle and to stress granules, establishing roles in spindle integrity via KIFC1 and in condensate dynamics via VCP.\",\n      \"evidence\": \"siRNA screen, Co-IP, ubiquitination assays, CRISPR KO, catalytic-mutant and KIFC1 overexpression rescue with imaging; proximity proteomics and Co-IP with VCP plus SG dynamics assays; Co-IP and G-quadruplex assays for DDX5\",\n      \"pmids\": [\"39789388\", \"41651815\", \"40651961\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How OTUD6B is recruited to centrosomes vs stress granules not defined\", \"Whether the proteasome-assembly defect connects to centrosome/spindle phenotypes unexplored\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Defined FXR1 as a domain-specific K48 substrate, linking OTUD6B to an FXR1–MEK2–ERK feed-forward oncogenic loop in colorectal liver metastasis.\",\n      \"evidence\": \"Domain-mapping Co-IP, K48-specific ubiquitination assays, MEK2 mRNA stability assays, catalytic mutant and in vivo liver metastasis models\",\n      \"pmids\": [\"42056075\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Generalizability of the N-terminal/KH-domain recognition mode to other substrates untested\", \"Direct structural basis of OTUD6B–FXR1 binding unresolved\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How OTUD6B achieves substrate and linkage selectivity (K48 vs K33), and how its catalytic, scaffolding, translational, and condensate functions are coordinated within a single protein, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structural model of substrate or linkage recognition\", \"Mechanism integrating proteasome-assembly, spindle, and stress-granule roles unknown\", \"Determinants switching between catalytic and scaffold-only modes undefined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 3, 5, 8, 10, 11, 14, 15, 16]},\n      {\"term_id\": \"GO:0016787\", \"supporting_discovery_ids\": [0, 4, 7]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [3]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [2]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005815\", \"supporting_discovery_ids\": [14]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [14]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [17]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [0, 5, 14]},\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [2, 3]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [4, 7]},\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [1, 16]},\n      {\"term_id\": \"R-HSA-8953897\", \"supporting_discovery_ids\": [17]}\n    ],\n    \"complexes\": [\"CBC-VHL E3 ligase complex\", \"48S preinitiation complex\"],\n    \"partners\": [\"VHL\", \"LIN28B\", \"KIFC1\", \"IRF3\", \"BTRC\", \"RIPK1\", \"DDX5\", \"FXR1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}