{"gene":"KCMF1","run_date":"2026-06-10T01:55:23","timeline":{"discoveries":[{"year":2015,"finding":"KCMF1 C-terminus binds directly to RAD6 (ubiquitin E2), while KCMF1 N-terminal domains interact with UBR4 and intracellular vesicle- and mitochondria-associated proteins, forming a RAD6-KCMF1-UBR4 E2-E3 complex. KCMF1 and RAD6 colocalize at late endosomes and lysosomes, and disruption of KCMF1 or RAD6 causes defects in late endosome vesicle dynamics. RAD6A point mutants (R7W and R11Q) found in X-linked intellectual disability specifically lose interaction with KCMF1 and UBR4.","method":"Affinity purification-mass spectrometry, NMR, in vivo and in vitro interaction mapping, colocalization imaging","journal":"Molecular & cellular proteomics : MCP","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — NMR direct binding, in vitro and in vivo interaction mapping, colocalization, multiple orthogonal methods in a single rigorous study","pmids":["25582440"],"is_preprint":false},{"year":2025,"finding":"KCMF1 functions as an autophagic N-recognin (ZZ/N-recognin) in the Arg/N-degron pathway: its ZZ-type zinc finger domain binds N-terminal arginine (Nt-Arg) and structurally related Nt-motifs, analogous to the ZZ domain of p62. Under oxidative/hypoxic stress, Nt-Cys is oxidized to Cys sulfonic acid and arginylated (Arg-CysO3 N-degron), which binds KCMF1 to induce assembly of K63-linked ubiquitin chains; p62-type autophagic receptors then bind via UBA domain to direct autophagic degradation. KCMF1 also undergoes N-degron-stimulated self-polymerization.","method":"Biochemical assays with synthetic N-degrons, in vitro ubiquitination assays, interaction mapping with ZZ domain mutants","journal":"Methods in enzymology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with synthetic N-degrons, mutagenesis of ZZ domain, multiple biochemical assays in a single focused mechanistic study","pmids":["40992840"],"is_preprint":false},{"year":2024,"finding":"Cryo-EM structure of the UBR4-KCMF1-CALM1 complex reveals a 1.3 MDa ring architecture with a central substrate-binding arena and flexibly attached catalytic units. UBR4 acts as an E4 ligase extending K48-specific ubiquitin chains; efficient substrate targeting requires both pre-ubiquitination and specific N-degrons. KCMF1 acts as a key substrate filter within this megacomplex.","method":"Cryo-EM structural analysis, in vitro ubiquitination reconstitution","journal":"bioRxiv","confidence":"High","confidence_rationale":"Tier 1 / Moderate — cryo-EM structure with functional validation of substrate targeting and chain specificity, multiple orthogonal approaches in one study","pmids":[],"is_preprint":true},{"year":2024,"finding":"The UBR4-KCMF1 ubiquitin ligase complex is required for efficient degradation of multiple unrelated orphan subunits (from chaperonin, proteasome cap, proteasome core, and protein targeting complex). Epistasis analysis and in vitro reconstitution show UBR4-KCMF1 acts downstream of a priming E3 ligase that first mono-ubiquitinates orphan substrates; UBR4 then recognizes both the orphan substrate and its mono-ubiquitin and builds K48-linked poly-ubiquitin degradation signals.","method":"Epistasis analysis in cells, in vitro reconstitution, loss-of-function cellular assays","journal":"bioRxiv","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — in vitro reconstitution plus cellular epistasis with multiple substrates, two orthogonal approaches establishing pathway position","pmids":[],"is_preprint":true},{"year":2026,"finding":"KCMF1 directly interacts with AMPKα and catalyzes its K48-linked polyubiquitination, promoting AMPKα proteasomal degradation and suppressing hepatic AMPK signaling. This was demonstrated by co-immunoprecipitation, GST pull-down, and biochemical ubiquitin-linkage specificity assays. Hepatocyte-specific KCMF1 deletion protected against MASLD in multiple mouse models, and AMPK pharmacological activation rescued KCMF1-driven pathology.","method":"Co-immunoprecipitation, GST pull-down, ubiquitination assay with K48-linkage specificity, hepatocyte-specific KO mouse models, pharmacological rescue","journal":"Metabolism: clinical and experimental","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — direct binding by pulldown and Co-IP, in vivo genetic and pharmacological rescue experiments, K48-linkage specificity biochemically established","pmids":["42162901"],"is_preprint":false},{"year":2026,"finding":"KCMF1 interacts with nucleoredoxin (NXN) and promotes its degradation through K63-linked ubiquitination. Silencing NXN facilitates cell proliferation, migration, and invasion through activating the β-catenin signaling pathway. Substrate was identified by IP-LC/MS and label-free proteomics.","method":"IP-LC/MS, label-free proteomics, Co-immunoprecipitation, in vitro ubiquitination assay, loss-of-function cell assays and xenograft","journal":"Cell cycle (Georgetown, Tex.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and proteomic substrate identification with K63-linkage specificity, single lab, supported by in vivo xenograft","pmids":["41721648"],"is_preprint":false},{"year":2025,"finding":"KCMF1 ubiquitinates HRI (heme-regulated inhibitor kinase), promoting its degradation. KCMF1 knockdown reduced HRI ubiquitination and led to increased eIF2α phosphorylation and upregulation of ATF4, ATF3, and sestrin 2, activating the integrated stress response (ISR). An ISR inhibitor reversed the effects of KCMF1 knockdown, demonstrating pathway dependency.","method":"Ni-NTA pull-down ubiquitination assay, western blot, immunohistochemistry, loss-of-function cell assays, pharmacological rescue","journal":"Biochemical pharmacology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct ubiquitination demonstrated by pull-down, pathway dependency confirmed by pharmacological rescue, single lab","pmids":["41391693"],"is_preprint":false},{"year":2025,"finding":"KCMF1 overexpression facilitates FUS nuclear translocation in renal cell carcinoma, enhancing FUS binding to CENPT mRNA and subsequent CENPT upregulation. KCMF1 physically interacts with FUS, as shown by Co-immunoprecipitation. This axis promotes abnormal chromosome segregation and genomic instability via JNK pathway activation.","method":"Co-immunoprecipitation, immunofluorescence, flow cytometry, loss-of-function and gain-of-function cell assays, xenograft","journal":"Journal of translational medicine","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — Co-IP for interaction, immunofluorescence for nuclear translocation, functional cellular and in vivo assays; single lab","pmids":["41184988"],"is_preprint":false},{"year":2013,"finding":"KCMF1 physically interacts with 14-3-3σ protein, as identified by yeast two-hybrid screen and confirmed by co-immunoprecipitation. Knockdown of either 14-3-3σ or KCMF1 significantly inhibited cell proliferation and colony formation of colon cancer stem cells.","method":"Yeast two-hybrid, co-immunoprecipitation, siRNA knockdown with proliferation/colony assays","journal":"World journal of gastroenterology","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — yeast two-hybrid confirmed by Co-IP, functional knockdown phenotype; single lab, two orthogonal interaction methods","pmids":["23840115"],"is_preprint":false},{"year":2010,"finding":"KCMF1 promotes cell proliferation, migration, and invasion in vitro and in the chicken chorioallantoic membrane model. KCMF1 knockdown in TGF-α transgenic mice reduced premalignant lesions and prevented pancreatic cancer formation, associated with decreased expression of cyclin D and CDK4. Nuclear KCMF1 localization was established in preneoplastic lesions.","method":"Cell culture proliferation/migration/invasion assays, CAM model, gene-trap knockdown mouse crossed to TGF-α transgenic model, immunohistochemistry for subcellular localization and downstream markers","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo genetic KO model with defined mechanistic downstream (cyclin D, CDK4), multiple assays in single lab","pmids":["20473331"],"is_preprint":false},{"year":2023,"finding":"Disruption of the RAD6-KCMF1-UBR4 ubiquitin ligase complex in CD8+ memory T cells from renal cell carcinoma patients impairs autophagy and reduces the survival and anti-tumor capacity of these cells.","method":"Flow cytometry for memory T cell subsets, expression analysis (cellular and molecular levels) in patient PBMCs, JC-1 staining for mitochondrial membrane potential, Annexin/PI apoptosis assay","journal":"Cancer letters","confidence":"Low","confidence_rationale":"Tier 3 / Weak — descriptive colocalization and expression in patient samples, no direct mechanistic reconstitution; single study","pmids":["37084875"],"is_preprint":false},{"year":2022,"finding":"KCMF1 and its associated proteins RAD6 and UBR4 co-localize in renal cell carcinoma tumor cells, with discrepancies in ubiquitin ligase complex formation and autophagosome assembly (LC3B, p62) observed in tumor vs. non-tumor tissue. Ionic concentrations of K+, Na+, and Zn2+ differ between tumor and non-tumor cells of RCC patients.","method":"Confocal microscopy co-localization, immunofluorescence staining, inductively coupled plasma mass spectrometry (ICPMS)","journal":"Journal of cancer research and clinical oncology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — descriptive co-localization and ionic analysis in patient tissue, no direct functional reconstitution; single study","pmids":["36515749"],"is_preprint":false},{"year":2006,"finding":"KCMF1 expression is suppressed by constitutively high CD99 levels in Ewing's sarcoma cells. Forced ectopic KCMF1 expression reduced migratory ability of ESFT cells, similar to CD99 silencing.","method":"RNAi-mediated CD99 suppression, ectopic KCMF1 overexpression, migration assays in ESFT cell lines","journal":"Oncogene","confidence":"Low","confidence_rationale":"Tier 3 / Weak — functional overexpression with migration assay, no direct molecular mechanism identified; single lab","pmids":["16314831"],"is_preprint":false},{"year":2026,"finding":"ABHD10 interacts with KCMF1, as shown by co-immunoprecipitation, suggesting the ABHD10-KCMF1 complex integrates mitochondrial quality control, lipid homeostasis, and redox balance in cochlear aging.","method":"Co-immunoprecipitation, GO/KEGG analysis, senescence assays in HEI-OC1 cells","journal":"Journal of cellular physiology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single Co-IP demonstrating interaction, functional consequences attributed to ABHD10 knockdown not directly to KCMF1; single lab","pmids":["42206676"],"is_preprint":false}],"current_model":"KCMF1 is a RING zinc-finger E3 ubiquitin ligase that acts as a ZZ-domain N-recognin, binding N-terminal arginine degrons to assemble K63-linked ubiquitin chains and direct substrates to lysosomal/autophagic degradation; it also forms a tripartite RAD6 (E2)–KCMF1–UBR4 complex that acts as an E4 chain-elongating ligase extending K48-linked chains on pre-ubiquitinated orphan protein subunits to target them for proteasomal degradation, with KCMF1 serving as a critical substrate filter; additional direct substrates include AMPKα (K48-linked degradation promoting MASLD), HRI kinase (suppressing the integrated stress response), and NXN (K63-linked degradation activating β-catenin signaling), and KCMF1 physically interacts with partners including 14-3-3σ, FUS, and ABHD10."},"narrative":{"mechanistic_narrative":"KCMF1 is a RING zinc-finger E3 ubiquitin ligase that operates within the Arg/N-degron protein quality-control machinery, coupling substrate recognition to either autophagic or proteasomal degradation [PMID:40992840]. Through its ZZ-type zinc finger it acts as an N-recognin, binding N-terminal arginine and oxidized/arginylated Nt-Cys degrons to assemble K63-linked ubiquitin chains that recruit p62-type autophagic receptors; under oxidative or hypoxic stress this directs substrates to autophagy, and KCMF1 itself undergoes N-degron-stimulated self-polymerization [PMID:40992840]. KCMF1 also nucleates a tripartite RAD6 (E2)–KCMF1–UBR4 complex: its C-terminus binds RAD6 while its N-terminal region binds UBR4, and the assembly localizes to late endosomes and lysosomes where it controls vesicle dynamics [PMID:25582440]. Within the resulting megadalton UBR4–KCMF1 machine, UBR4 functions as an E4 ligase extending K48-linked chains on pre-ubiquitinated orphan protein subunits, with KCMF1 serving as the substrate filter that gates entry; efficient targeting requires both prior mono-ubiquitination by a priming E3 and a specific N-degron. Beyond orphan quality control, KCMF1 directly ubiquitinates discrete substrates with defined chain specificity and physiological consequence: K48-linked degradation of AMPKα suppresses hepatic AMPK signaling and drives MASLD [PMID:42162901], degradation of the HRI kinase restrains the integrated stress response [PMID:41391693], and K63-linked degradation of nucleoredoxin (NXN) activates β-catenin signaling [PMID:41721648]. KCMF1 additionally engages 14-3-3σ and FUS, and promotes proliferation, migration, and invasion across several cancer models [PMID:41184988, PMID:23840115, PMID:20473331].","teleology":[{"year":2010,"claim":"Established KCMF1 as a pro-tumorigenic factor with an in vivo phenotype before any enzymatic activity was known, linking it to proliferation control.","evidence":"Cell proliferation/migration/invasion assays, CAM model, gene-trap knockdown crossed to TGF-α transgenic mice, IHC for cyclin D/CDK4","pmids":["20473331"],"confidence":"Medium","gaps":["No molecular activity or substrate identified","Mechanism linking KCMF1 to cyclin D/CDK4 unresolved","Nuclear localization not mechanistically explained"]},{"year":2013,"claim":"Identified 14-3-3σ as a physical partner supporting a proliferative role in cancer stem cells, expanding the KCMF1 interaction landscape.","evidence":"Yeast two-hybrid and co-immunoprecipitation with siRNA knockdown proliferation/colony assays in colon cancer stem cells","pmids":["23840115"],"confidence":"Medium","gaps":["Functional consequence of the interaction at the molecular level undefined","No ubiquitination link established","Single lab"]},{"year":2015,"claim":"Defined KCMF1 as the scaffolding hub of a RAD6–KCMF1–UBR4 E2–E3 complex localized to the endolysosomal system, establishing its architecture and a disease-relevant binding interface.","evidence":"Affinity purification–MS, NMR direct binding, in vitro/in vivo interaction mapping, colocalization imaging; RAD6A XLID mutants lose KCMF1/UBR4 binding","pmids":["25582440"],"confidence":"High","gaps":["Catalytic output and substrates of the complex not yet defined","Chain-type specificity unknown at this stage","Role in vesicle dynamics mechanistically incomplete"]},{"year":2024,"claim":"Resolved the structure and pathway logic of the UBR4–KCMF1 machine, showing it is an E4 chain-elongating ligase acting downstream of a priming E3 to clear orphan subunits, with KCMF1 as substrate filter.","evidence":"Cryo-EM of the 1.3 MDa UBR4–KCMF1–CALM1 ring (preprint), cellular epistasis, and in vitro reconstitution with multiple orphan substrates (preprint)","pmids":[],"confidence":"High","gaps":["Preprint, not peer-reviewed","Precise KCMF1 contribution to substrate selection vs. UBR4 not fully separated","Identity of priming E3 not defined"]},{"year":2025,"claim":"Demonstrated KCMF1 is an autophagic N-recognin that reads Nt-Arg and oxidized/arginylated Nt-Cys degrons to build K63 chains, connecting it to stress-induced autophagy.","evidence":"In vitro ubiquitination and binding assays with synthetic N-degrons and ZZ-domain mutants","pmids":["40992840"],"confidence":"High","gaps":["Endogenous physiological substrates of the autophagic branch not enumerated","Mechanism of self-polymerization unresolved","Interplay between K63-autophagic and K48-proteasomal branches unclear"]},{"year":2025,"claim":"Identified a non-degradative KCMF1–FUS axis in renal cell carcinoma, broadening KCMF1 function beyond canonical ubiquitin-ligase activity.","evidence":"Co-IP, immunofluorescence, flow cytometry, loss-/gain-of-function cell assays and xenograft","pmids":["41184988"],"confidence":"Medium","gaps":["Whether FUS is ubiquitinated by KCMF1 not established","Mechanism of FUS nuclear translocation unclear","Single lab"]},{"year":2026,"claim":"Established specific physiological substrates with defined chain linkages — AMPKα (K48), HRI (degradation), and NXN (K63) — linking KCMF1 to metabolic disease, stress signaling, and Wnt/β-catenin signaling.","evidence":"Co-IP/GST pull-down/Ni-NTA ubiquitination with linkage specificity, IP-LC/MS substrate ID, hepatocyte-specific KO mice, and pharmacological rescue (AMPK activator; ISR inhibitor)","pmids":["42162901","41391693","41721648"],"confidence":"Medium","gaps":["Whether these substrates share the N-degron recognition mode is untested","Tissue/context determinants of substrate choice unknown","AMPKα and NXN findings each from single labs"]},{"year":null,"claim":"How KCMF1 switches between K63-linked autophagic targeting and K48-linked proteasomal targeting, and how degron recognition selects among its diverse substrates, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified model linking N-degron recognition to chain-type choice","Regulatory inputs controlling branch selection unknown","Endogenous substrate repertoire incompletely mapped"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[1,2,3,4,5,6]},{"term_id":"GO:0016740","term_label":"transferase activity","supporting_discovery_ids":[1,4,5]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,2,3]}],"localization":[{"term_id":"GO:0005768","term_label":"endosome","supporting_discovery_ids":[0]},{"term_id":"GO:0005764","term_label":"lysosome","supporting_discovery_ids":[0]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[9]}],"pathway":[{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[1,2,3,4]},{"term_id":"R-HSA-9612973","term_label":"Autophagy","supporting_discovery_ids":[1]},{"term_id":"R-HSA-8953897","term_label":"Cellular responses to stimuli","supporting_discovery_ids":[1,6]}],"complexes":["RAD6–KCMF1–UBR4 ubiquitin ligase complex","UBR4–KCMF1–CALM1 megacomplex"],"partners":["RAD6","UBR4","AMPKΑ","NXN","HRI","FUS","14-3-3Σ","ABHD10"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9P0J7","full_name":"E3 ubiquitin-protein ligase KCMF1","aliases":["FGF-induced in gastric cancer","Potassium channel modulatory factor","PCMF","ZZ-type zinc finger-containing protein 1"],"length_aa":381,"mass_kda":41.9,"function":"E3 ubiquitin-protein ligase which accepts ubiquitin from an E2 ubiquitin-conjugating enzyme and then transfers it to targeted substrates, promoting their degradation by the proteasome (PubMed:15581609, PubMed:25582440, PubMed:34893540, PubMed:37891180, PubMed:38297121). Together with UBR4, component of the N-end rule pathway: ubiquitinates proteins bearing specific N-terminal residues that are destabilizing according to the N-end rule, leading to their degradation (PubMed:34893540, PubMed:37891180). Does not ubiquitinate proteins that are acetylated at the N-terminus (PubMed:37891180). Together with UBR4, part of a protein quality control pathway that catalyzes ubiquitination and degradation of proteins that have been oxidized in response to reactive oxygen species (ROS): recognizes proteins with an Arg-CysO3(H) degron at the N-terminus, and mediates assembly of heterotypic 'Lys-63'-/'Lys-27'-linked branched ubiquitin chains on oxidized proteins, leading to their degradation by autophagy (PubMed:34893540). Catalytic component of the SIFI complex, a multiprotein complex required to inhibit the mitochondrial stress response after a specific stress event has been resolved: ubiquitinates and degrades (1) components of the HRI-mediated signaling of the integrated stress response, such as DELE1 and EIF2AK1/HRI, as well as (2) unimported mitochondrial precursors (PubMed:38297121). Within the SIFI complex, UBR4 initiates ubiquitin chain that are further elongated or branched by KCMF1 (PubMed:38297121)","subcellular_location":"Cytoplasm; Late endosome; Lysosome","url":"https://www.uniprot.org/uniprotkb/Q9P0J7/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/KCMF1","classification":"Common Essential","n_dependent_lines":1028,"n_total_lines":1208,"dependency_fraction":0.8509933774834437},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/KCMF1","total_profiled":1310},"omim":[{"mim_id":"615741","title":"DAP3-BINDING CELL DEATH ENHANCER 1; DELE1","url":"https://www.omim.org/entry/615741"},{"mim_id":"614719","title":"POTASSIUM CHANNEL MODULATORY FACTOR 1; KCMF1","url":"https://www.omim.org/entry/614719"},{"mim_id":"613635","title":"EUKARYOTIC TRANSLATION INITIATION FACTOR 2-ALPHA KINASE 1; EIF2AK1","url":"https://www.omim.org/entry/613635"},{"mim_id":"609890","title":"UBIQUITIN PROTEIN LIGASE E3 COMPONENT N-RECOGNIN 4; UBR4","url":"https://www.omim.org/entry/609890"},{"mim_id":"114180","title":"CALMODULIN 1; CALM1","url":"https://www.omim.org/entry/114180"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Enhanced","locations":[{"location":"Cytosol","reliability":"Enhanced"},{"location":"Nucleoplasm","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/KCMF1"},"hgnc":{"alias_symbol":["DEBT91","PCMF","DKFZP434L1021","ZZZ1"],"prev_symbol":[]},"alphafold":{"accession":"Q9P0J7","domains":[{"cath_id":"-","chopping":"19-135","consensus_level":"medium","plddt":90.6086,"start":19,"end":135}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9P0J7","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9P0J7-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9P0J7-F1-predicted_aligned_error_v6.png","plddt_mean":71.0},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=KCMF1","jax_strain_url":"https://www.jax.org/strain/search?query=KCMF1"},"sequence":{"accession":"Q9P0J7","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9P0J7.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9P0J7/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9P0J7"}},"corpus_meta":[{"pmid":"16314831","id":"PMC_16314831","title":"Suppression of KCMF1 by constitutive high CD99 expression is involved in the migratory ability of Ewing's sarcoma cells.","date":"2006","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/16314831","citation_count":45,"is_preprint":false},{"pmid":"25582440","id":"PMC_25582440","title":"KCMF1 (potassium channel modulatory factor 1) Links RAD6 to UBR4 (ubiquitin N-recognin domain-containing E3 ligase 4) and lysosome-mediated degradation.","date":"2015","source":"Molecular & cellular proteomics : MCP","url":"https://pubmed.ncbi.nlm.nih.gov/25582440","citation_count":38,"is_preprint":false},{"pmid":"20473331","id":"PMC_20473331","title":"The zinc-finger protein KCMF1 is overexpressed during pancreatic cancer development and downregulation of KCMF1 inhibits pancreatic cancer development in mice.","date":"2010","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/20473331","citation_count":21,"is_preprint":false},{"pmid":"35032791","id":"PMC_35032791","title":"CircHIPK3 contributes to human villous trophoblast growth, migration and invasion via modulating the pathway of miR-346/KCMF1.","date":"2021","source":"Placenta","url":"https://pubmed.ncbi.nlm.nih.gov/35032791","citation_count":14,"is_preprint":false},{"pmid":"37084875","id":"PMC_37084875","title":"Disruption in networking of KCMF1 linked ubiquitin ligase impairs autophagy in CD8+ memory T cells of patients with renal cell carcinoma.","date":"2023","source":"Cancer letters","url":"https://pubmed.ncbi.nlm.nih.gov/37084875","citation_count":13,"is_preprint":false},{"pmid":"23840115","id":"PMC_23840115","title":"Interaction of 14-3-3σ with KCMF1 suppresses the proliferation and colony formation of human colon cancer stem cells.","date":"2013","source":"World journal of gastroenterology","url":"https://pubmed.ncbi.nlm.nih.gov/23840115","citation_count":11,"is_preprint":false},{"pmid":"12810064","id":"PMC_12810064","title":"Debt91, a putative zinc finger protein differentially expressed during epithelial morphogenesis.","date":"2003","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/12810064","citation_count":8,"is_preprint":false},{"pmid":"36515749","id":"PMC_36515749","title":"KCMF1 regulates autophagy and ion channels' function in renal cell carcinoma: a future therapeutic target.","date":"2022","source":"Journal of cancer research and clinical oncology","url":"https://pubmed.ncbi.nlm.nih.gov/36515749","citation_count":4,"is_preprint":false},{"pmid":"37832749","id":"PMC_37832749","title":"KCMF1-like suppresses white spot syndrome virus infection by promoting apoptosis in mud crab (Scylla paramamosain).","date":"2023","source":"Fish & shellfish immunology","url":"https://pubmed.ncbi.nlm.nih.gov/37832749","citation_count":1,"is_preprint":false},{"pmid":"41721648","id":"PMC_41721648","title":"KCMF1 promotes malignant progression by NXN ubiquitin-dependent degradation in ovarian cancer.","date":"2026","source":"Cell cycle (Georgetown, Tex.)","url":"https://pubmed.ncbi.nlm.nih.gov/41721648","citation_count":0,"is_preprint":false},{"pmid":"41938511","id":"PMC_41938511","title":"Integrated analysis uncovers KCMF1 genetic susceptibility and the SNRPD2 axis in renal cell carcinoma.","date":"2026","source":"International journal of medical sciences","url":"https://pubmed.ncbi.nlm.nih.gov/41938511","citation_count":0,"is_preprint":false},{"pmid":"41391693","id":"PMC_41391693","title":"KCMF1 regulates HRI ubiquitination to inhibit the integrated stress response in ovarian cancer.","date":"2025","source":"Biochemical pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/41391693","citation_count":0,"is_preprint":false},{"pmid":"41184988","id":"PMC_41184988","title":"Fused in Sarcoma (FUS) promotes renal cell carcinoma progression via the KCMF1/FUS/CENPT axis and activation of the JNK signaling pathway.","date":"2025","source":"Journal of translational 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ligase","date":"2024-08-08","source":"bioRxiv","url":"https://doi.org/10.1101/2024.08.07.607117","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":12623,"output_tokens":3852,"usd":0.047824,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":11543,"output_tokens":3180,"usd":0.068608,"stage2_stop_reason":"end_turn"},"total_usd":0.116432,"stage1_batch_id":"msgbatch_01DUPACZi7AAWwuZiUjseQF4","stage2_batch_id":"msgbatch_018yt7chE3f48PEFtDwtj6AE","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2015,\n      \"finding\": \"KCMF1 C-terminus binds directly to RAD6 (ubiquitin E2), while KCMF1 N-terminal domains interact with UBR4 and intracellular vesicle- and mitochondria-associated proteins, forming a RAD6-KCMF1-UBR4 E2-E3 complex. KCMF1 and RAD6 colocalize at late endosomes and lysosomes, and disruption of KCMF1 or RAD6 causes defects in late endosome vesicle dynamics. RAD6A point mutants (R7W and R11Q) found in X-linked intellectual disability specifically lose interaction with KCMF1 and UBR4.\",\n      \"method\": \"Affinity purification-mass spectrometry, NMR, in vivo and in vitro interaction mapping, colocalization imaging\",\n      \"journal\": \"Molecular & cellular proteomics : MCP\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — NMR direct binding, in vitro and in vivo interaction mapping, colocalization, multiple orthogonal methods in a single rigorous study\",\n      \"pmids\": [\"25582440\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"KCMF1 functions as an autophagic N-recognin (ZZ/N-recognin) in the Arg/N-degron pathway: its ZZ-type zinc finger domain binds N-terminal arginine (Nt-Arg) and structurally related Nt-motifs, analogous to the ZZ domain of p62. Under oxidative/hypoxic stress, Nt-Cys is oxidized to Cys sulfonic acid and arginylated (Arg-CysO3 N-degron), which binds KCMF1 to induce assembly of K63-linked ubiquitin chains; p62-type autophagic receptors then bind via UBA domain to direct autophagic degradation. KCMF1 also undergoes N-degron-stimulated self-polymerization.\",\n      \"method\": \"Biochemical assays with synthetic N-degrons, in vitro ubiquitination assays, interaction mapping with ZZ domain mutants\",\n      \"journal\": \"Methods in enzymology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with synthetic N-degrons, mutagenesis of ZZ domain, multiple biochemical assays in a single focused mechanistic study\",\n      \"pmids\": [\"40992840\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Cryo-EM structure of the UBR4-KCMF1-CALM1 complex reveals a 1.3 MDa ring architecture with a central substrate-binding arena and flexibly attached catalytic units. UBR4 acts as an E4 ligase extending K48-specific ubiquitin chains; efficient substrate targeting requires both pre-ubiquitination and specific N-degrons. KCMF1 acts as a key substrate filter within this megacomplex.\",\n      \"method\": \"Cryo-EM structural analysis, in vitro ubiquitination reconstitution\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — cryo-EM structure with functional validation of substrate targeting and chain specificity, multiple orthogonal approaches in one study\",\n      \"pmids\": [],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"The UBR4-KCMF1 ubiquitin ligase complex is required for efficient degradation of multiple unrelated orphan subunits (from chaperonin, proteasome cap, proteasome core, and protein targeting complex). Epistasis analysis and in vitro reconstitution show UBR4-KCMF1 acts downstream of a priming E3 ligase that first mono-ubiquitinates orphan substrates; UBR4 then recognizes both the orphan substrate and its mono-ubiquitin and builds K48-linked poly-ubiquitin degradation signals.\",\n      \"method\": \"Epistasis analysis in cells, in vitro reconstitution, loss-of-function cellular assays\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — in vitro reconstitution plus cellular epistasis with multiple substrates, two orthogonal approaches establishing pathway position\",\n      \"pmids\": [],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"KCMF1 directly interacts with AMPKα and catalyzes its K48-linked polyubiquitination, promoting AMPKα proteasomal degradation and suppressing hepatic AMPK signaling. This was demonstrated by co-immunoprecipitation, GST pull-down, and biochemical ubiquitin-linkage specificity assays. Hepatocyte-specific KCMF1 deletion protected against MASLD in multiple mouse models, and AMPK pharmacological activation rescued KCMF1-driven pathology.\",\n      \"method\": \"Co-immunoprecipitation, GST pull-down, ubiquitination assay with K48-linkage specificity, hepatocyte-specific KO mouse models, pharmacological rescue\",\n      \"journal\": \"Metabolism: clinical and experimental\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — direct binding by pulldown and Co-IP, in vivo genetic and pharmacological rescue experiments, K48-linkage specificity biochemically established\",\n      \"pmids\": [\"42162901\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"KCMF1 interacts with nucleoredoxin (NXN) and promotes its degradation through K63-linked ubiquitination. Silencing NXN facilitates cell proliferation, migration, and invasion through activating the β-catenin signaling pathway. Substrate was identified by IP-LC/MS and label-free proteomics.\",\n      \"method\": \"IP-LC/MS, label-free proteomics, Co-immunoprecipitation, in vitro ubiquitination assay, loss-of-function cell assays and xenograft\",\n      \"journal\": \"Cell cycle (Georgetown, Tex.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and proteomic substrate identification with K63-linkage specificity, single lab, supported by in vivo xenograft\",\n      \"pmids\": [\"41721648\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"KCMF1 ubiquitinates HRI (heme-regulated inhibitor kinase), promoting its degradation. KCMF1 knockdown reduced HRI ubiquitination and led to increased eIF2α phosphorylation and upregulation of ATF4, ATF3, and sestrin 2, activating the integrated stress response (ISR). An ISR inhibitor reversed the effects of KCMF1 knockdown, demonstrating pathway dependency.\",\n      \"method\": \"Ni-NTA pull-down ubiquitination assay, western blot, immunohistochemistry, loss-of-function cell assays, pharmacological rescue\",\n      \"journal\": \"Biochemical pharmacology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct ubiquitination demonstrated by pull-down, pathway dependency confirmed by pharmacological rescue, single lab\",\n      \"pmids\": [\"41391693\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"KCMF1 overexpression facilitates FUS nuclear translocation in renal cell carcinoma, enhancing FUS binding to CENPT mRNA and subsequent CENPT upregulation. KCMF1 physically interacts with FUS, as shown by Co-immunoprecipitation. This axis promotes abnormal chromosome segregation and genomic instability via JNK pathway activation.\",\n      \"method\": \"Co-immunoprecipitation, immunofluorescence, flow cytometry, loss-of-function and gain-of-function cell assays, xenograft\",\n      \"journal\": \"Journal of translational medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — Co-IP for interaction, immunofluorescence for nuclear translocation, functional cellular and in vivo assays; single lab\",\n      \"pmids\": [\"41184988\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"KCMF1 physically interacts with 14-3-3σ protein, as identified by yeast two-hybrid screen and confirmed by co-immunoprecipitation. Knockdown of either 14-3-3σ or KCMF1 significantly inhibited cell proliferation and colony formation of colon cancer stem cells.\",\n      \"method\": \"Yeast two-hybrid, co-immunoprecipitation, siRNA knockdown with proliferation/colony assays\",\n      \"journal\": \"World journal of gastroenterology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — yeast two-hybrid confirmed by Co-IP, functional knockdown phenotype; single lab, two orthogonal interaction methods\",\n      \"pmids\": [\"23840115\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"KCMF1 promotes cell proliferation, migration, and invasion in vitro and in the chicken chorioallantoic membrane model. KCMF1 knockdown in TGF-α transgenic mice reduced premalignant lesions and prevented pancreatic cancer formation, associated with decreased expression of cyclin D and CDK4. Nuclear KCMF1 localization was established in preneoplastic lesions.\",\n      \"method\": \"Cell culture proliferation/migration/invasion assays, CAM model, gene-trap knockdown mouse crossed to TGF-α transgenic model, immunohistochemistry for subcellular localization and downstream markers\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo genetic KO model with defined mechanistic downstream (cyclin D, CDK4), multiple assays in single lab\",\n      \"pmids\": [\"20473331\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Disruption of the RAD6-KCMF1-UBR4 ubiquitin ligase complex in CD8+ memory T cells from renal cell carcinoma patients impairs autophagy and reduces the survival and anti-tumor capacity of these cells.\",\n      \"method\": \"Flow cytometry for memory T cell subsets, expression analysis (cellular and molecular levels) in patient PBMCs, JC-1 staining for mitochondrial membrane potential, Annexin/PI apoptosis assay\",\n      \"journal\": \"Cancer letters\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — descriptive colocalization and expression in patient samples, no direct mechanistic reconstitution; single study\",\n      \"pmids\": [\"37084875\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"KCMF1 and its associated proteins RAD6 and UBR4 co-localize in renal cell carcinoma tumor cells, with discrepancies in ubiquitin ligase complex formation and autophagosome assembly (LC3B, p62) observed in tumor vs. non-tumor tissue. Ionic concentrations of K+, Na+, and Zn2+ differ between tumor and non-tumor cells of RCC patients.\",\n      \"method\": \"Confocal microscopy co-localization, immunofluorescence staining, inductively coupled plasma mass spectrometry (ICPMS)\",\n      \"journal\": \"Journal of cancer research and clinical oncology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — descriptive co-localization and ionic analysis in patient tissue, no direct functional reconstitution; single study\",\n      \"pmids\": [\"36515749\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"KCMF1 expression is suppressed by constitutively high CD99 levels in Ewing's sarcoma cells. Forced ectopic KCMF1 expression reduced migratory ability of ESFT cells, similar to CD99 silencing.\",\n      \"method\": \"RNAi-mediated CD99 suppression, ectopic KCMF1 overexpression, migration assays in ESFT cell lines\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — functional overexpression with migration assay, no direct molecular mechanism identified; single lab\",\n      \"pmids\": [\"16314831\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"ABHD10 interacts with KCMF1, as shown by co-immunoprecipitation, suggesting the ABHD10-KCMF1 complex integrates mitochondrial quality control, lipid homeostasis, and redox balance in cochlear aging.\",\n      \"method\": \"Co-immunoprecipitation, GO/KEGG analysis, senescence assays in HEI-OC1 cells\",\n      \"journal\": \"Journal of cellular physiology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single Co-IP demonstrating interaction, functional consequences attributed to ABHD10 knockdown not directly to KCMF1; single lab\",\n      \"pmids\": [\"42206676\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"KCMF1 is a RING zinc-finger E3 ubiquitin ligase that acts as a ZZ-domain N-recognin, binding N-terminal arginine degrons to assemble K63-linked ubiquitin chains and direct substrates to lysosomal/autophagic degradation; it also forms a tripartite RAD6 (E2)–KCMF1–UBR4 complex that acts as an E4 chain-elongating ligase extending K48-linked chains on pre-ubiquitinated orphan protein subunits to target them for proteasomal degradation, with KCMF1 serving as a critical substrate filter; additional direct substrates include AMPKα (K48-linked degradation promoting MASLD), HRI kinase (suppressing the integrated stress response), and NXN (K63-linked degradation activating β-catenin signaling), and KCMF1 physically interacts with partners including 14-3-3σ, FUS, and ABHD10.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"KCMF1 is a RING zinc-finger E3 ubiquitin ligase that operates within the Arg/N-degron protein quality-control machinery, coupling substrate recognition to either autophagic or proteasomal degradation [#1, #2]. Through its ZZ-type zinc finger it acts as an N-recognin, binding N-terminal arginine and oxidized/arginylated Nt-Cys degrons to assemble K63-linked ubiquitin chains that recruit p62-type autophagic receptors; under oxidative or hypoxic stress this directs substrates to autophagy, and KCMF1 itself undergoes N-degron-stimulated self-polymerization [#1]. KCMF1 also nucleates a tripartite RAD6 (E2)–KCMF1–UBR4 complex: its C-terminus binds RAD6 while its N-terminal region binds UBR4, and the assembly localizes to late endosomes and lysosomes where it controls vesicle dynamics [#0]. Within the resulting megadalton UBR4–KCMF1 machine, UBR4 functions as an E4 ligase extending K48-linked chains on pre-ubiquitinated orphan protein subunits, with KCMF1 serving as the substrate filter that gates entry; efficient targeting requires both prior mono-ubiquitination by a priming E3 and a specific N-degron [#2, #3]. Beyond orphan quality control, KCMF1 directly ubiquitinates discrete substrates with defined chain specificity and physiological consequence: K48-linked degradation of AMPKα suppresses hepatic AMPK signaling and drives MASLD [#4], degradation of the HRI kinase restrains the integrated stress response [#6], and K63-linked degradation of nucleoredoxin (NXN) activates β-catenin signaling [#5]. KCMF1 additionally engages 14-3-3σ and FUS, and promotes proliferation, migration, and invasion across several cancer models [#7, #8, #9].\",\n  \"teleology\": [\n    {\n      \"year\": 2010,\n      \"claim\": \"Established KCMF1 as a pro-tumorigenic factor with an in vivo phenotype before any enzymatic activity was known, linking it to proliferation control.\",\n      \"evidence\": \"Cell proliferation/migration/invasion assays, CAM model, gene-trap knockdown crossed to TGF-α transgenic mice, IHC for cyclin D/CDK4\",\n      \"pmids\": [\"20473331\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No molecular activity or substrate identified\", \"Mechanism linking KCMF1 to cyclin D/CDK4 unresolved\", \"Nuclear localization not mechanistically explained\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Identified 14-3-3σ as a physical partner supporting a proliferative role in cancer stem cells, expanding the KCMF1 interaction landscape.\",\n      \"evidence\": \"Yeast two-hybrid and co-immunoprecipitation with siRNA knockdown proliferation/colony assays in colon cancer stem cells\",\n      \"pmids\": [\"23840115\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional consequence of the interaction at the molecular level undefined\", \"No ubiquitination link established\", \"Single lab\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Defined KCMF1 as the scaffolding hub of a RAD6–KCMF1–UBR4 E2–E3 complex localized to the endolysosomal system, establishing its architecture and a disease-relevant binding interface.\",\n      \"evidence\": \"Affinity purification–MS, NMR direct binding, in vitro/in vivo interaction mapping, colocalization imaging; RAD6A XLID mutants lose KCMF1/UBR4 binding\",\n      \"pmids\": [\"25582440\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Catalytic output and substrates of the complex not yet defined\", \"Chain-type specificity unknown at this stage\", \"Role in vesicle dynamics mechanistically incomplete\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Resolved the structure and pathway logic of the UBR4–KCMF1 machine, showing it is an E4 chain-elongating ligase acting downstream of a priming E3 to clear orphan subunits, with KCMF1 as substrate filter.\",\n      \"evidence\": \"Cryo-EM of the 1.3 MDa UBR4–KCMF1–CALM1 ring (preprint), cellular epistasis, and in vitro reconstitution with multiple orphan substrates (preprint)\",\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Preprint, not peer-reviewed\", \"Precise KCMF1 contribution to substrate selection vs. UBR4 not fully separated\", \"Identity of priming E3 not defined\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Demonstrated KCMF1 is an autophagic N-recognin that reads Nt-Arg and oxidized/arginylated Nt-Cys degrons to build K63 chains, connecting it to stress-induced autophagy.\",\n      \"evidence\": \"In vitro ubiquitination and binding assays with synthetic N-degrons and ZZ-domain mutants\",\n      \"pmids\": [\"40992840\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Endogenous physiological substrates of the autophagic branch not enumerated\", \"Mechanism of self-polymerization unresolved\", \"Interplay between K63-autophagic and K48-proteasomal branches unclear\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Identified a non-degradative KCMF1–FUS axis in renal cell carcinoma, broadening KCMF1 function beyond canonical ubiquitin-ligase activity.\",\n      \"evidence\": \"Co-IP, immunofluorescence, flow cytometry, loss-/gain-of-function cell assays and xenograft\",\n      \"pmids\": [\"41184988\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether FUS is ubiquitinated by KCMF1 not established\", \"Mechanism of FUS nuclear translocation unclear\", \"Single lab\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Established specific physiological substrates with defined chain linkages — AMPKα (K48), HRI (degradation), and NXN (K63) — linking KCMF1 to metabolic disease, stress signaling, and Wnt/β-catenin signaling.\",\n      \"evidence\": \"Co-IP/GST pull-down/Ni-NTA ubiquitination with linkage specificity, IP-LC/MS substrate ID, hepatocyte-specific KO mice, and pharmacological rescue (AMPK activator; ISR inhibitor)\",\n      \"pmids\": [\"42162901\", \"41391693\", \"41721648\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether these substrates share the N-degron recognition mode is untested\", \"Tissue/context determinants of substrate choice unknown\", \"AMPKα and NXN findings each from single labs\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How KCMF1 switches between K63-linked autophagic targeting and K48-linked proteasomal targeting, and how degron recognition selects among its diverse substrates, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified model linking N-degron recognition to chain-type choice\", \"Regulatory inputs controlling branch selection unknown\", \"Endogenous substrate repertoire incompletely mapped\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [1, 2, 3, 4, 5, 6]},\n      {\"term_id\": \"GO:0016740\", \"supporting_discovery_ids\": [1, 4, 5]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0, 2, 3]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005768\", \"supporting_discovery_ids\": [0]},\n      {\"term_id\": \"GO:0005764\", \"supporting_discovery_ids\": [0]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [9]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [1, 2, 3, 4]},\n      {\"term_id\": \"R-HSA-9612973\", \"supporting_discovery_ids\": [1]},\n      {\"term_id\": \"R-HSA-8953897\", \"supporting_discovery_ids\": [1, 6]}\n    ],\n    \"complexes\": [\"RAD6–KCMF1–UBR4 ubiquitin ligase complex\", \"UBR4–KCMF1–CALM1 megacomplex\"],\n    \"partners\": [\"RAD6\", \"UBR4\", \"AMPKα\", \"NXN\", \"HRI\", \"FUS\", \"14-3-3σ\", \"ABHD10\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}