{"gene":"FBXL6","run_date":"2026-06-09T23:54:43","timeline":{"discoveries":[{"year":2020,"finding":"FBXL6 acts as a substrate recognition subunit of the SCF (Skp1-Cul1-F-box protein) E3 ubiquitin ligase complex and directly interacts with HSP90AA1, promoting its K63-linked polyubiquitination, which stabilizes (rather than degrades) HSP90AA1. Stabilized HSP90AA1 in turn prevents c-MYC degradation, and c-MYC transcriptionally induces FBXL6 expression by binding its promoter, forming a positive feedback loop.","method":"Co-immunoprecipitation, IP/Mass Spectrometry, in vivo ubiquitination assay, luciferase reporter assay, ChIP assay, shRNA knockdown in HCC cells and xenograft mouse model","journal":"Cell communication and signaling : CCS","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, IP/MS, in vivo ubiquitination assay, and transcriptional assays in one study with multiple orthogonal methods","pmids":["32576198"],"is_preprint":false},{"year":2021,"finding":"FBXL6 directly targets phosphorylated p53 (phospho-Ser315) to mediate its K48-linked polyubiquitination and proteasomal degradation, thereby suppressing p53 signaling. Conversely, p53 transcriptionally represses FBXL6 expression by binding its core promoter, forming a negative feedback loop.","method":"Co-immunoprecipitation, ubiquitination assay, proteasome inhibitor rescue experiments, luciferase reporter assay, shRNA knockdown with cell cycle and apoptosis readouts","journal":"Cell death and differentiation","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, in vivo ubiquitination assay, and transcriptional assays with multiple orthogonal methods in one rigorous study","pmids":["33568778"],"is_preprint":false},{"year":2023,"finding":"FBXL6 promotes polyubiquitination of both wild-type KRAS and mutant KRASG12D specifically at lysine 128, leading to their activation and enhanced binding to RAF kinase, thereby activating MEK/ERK/mTOR signaling. The oncogenic output of this axis depends on PRELID2-induced ROS generation.","method":"Co-immunoprecipitation, ubiquitination assay, RAS activity detection assay, transgenic mouse models (LC, KC, KLC), multiomics, pharmacological inhibitors","journal":"Military Medical Research","confidence":"High","confidence_rationale":"Tier 2 / Strong — Co-IP, ubiquitination assay, site-specific mutagenesis (K128), RAS activity assay, and transgenic mouse models with multiple orthogonal methods","pmids":["38124228"],"is_preprint":false},{"year":2023,"finding":"VRK2 kinase phosphorylates TKT (transketolase) at Thr287, which recruits FBXL6 to promote TKT ubiquitination and activation. Activated TKT drives ROS-mTOR signaling to upregulate PD-L1 and VRK2, leading to immune evasion and HCC metastasis.","method":"Co-immunoprecipitation, ubiquitination assay, phosphorylation assays, knockdown/overexpression in vitro, transgenic mouse models, in vivo xenograft","journal":"Experimental & molecular medicine","confidence":"High","confidence_rationale":"Tier 2 / Strong — Co-IP, ubiquitination assay, phosphorylation site identification, and in vivo transgenic mouse models with multiple orthogonal methods","pmids":["37653031"],"is_preprint":false},{"year":2023,"finding":"FBXL6 functions as an E3 ubiquitin ligase that specifically ubiquitinates newly synthesized (cytosolically translated) mitochondrial ribosomal proteins to control their quality. FBXL6 physically associates with chaperones involved in folding/trafficking of newly synthesized peptides and with ribosomal-associated quality control proteins; deletion of these interacting partners abolishes FBXL6-substrate interactions. FBXL6-knockout cells fail to degrade mistranslated mitochondrial ribosomal proteins, accumulate mitochondrial ribosomal protein aggregates, display altered mitochondrial metabolism, and exhibit inhibited cell cycle in oxidative conditions.","method":"Co-immunoprecipitation, ubiquitination assays, FBXL6 knockout cells, mitochondrial functional assays, proteomics, cell cycle analysis","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (Co-IP, ubiquitination assay, KO phenotyping, mitochondrial functional assays) in one rigorous study","pmids":["37267103"],"is_preprint":false},{"year":2022,"finding":"FBXL6 promotes keloid fibroblast proliferation by inducing c-MYC expression; FBXL6 knockdown reduces c-MYC levels along with cyclins A1, D2, E1, and Collagen I, and c-MYC overexpression rescues the proliferative defect caused by FBXL6 depletion, placing c-MYC downstream of FBXL6 in this pathway.","method":"shRNA knockdown, overexpression plasmid transfection, rescue experiment with c-MYC overexpression, cell viability assay (CCK-8), Western blot, RT-PCR","journal":"International wound journal","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — genetic epistasis via rescue experiment, Western blot, but no direct biochemical interaction or ubiquitination assay; single lab","pmids":["35606330"],"is_preprint":false},{"year":2025,"finding":"FBXL6 directly binds CDKN1C (p57Kip2) and promotes its polyubiquitination and proteasomal degradation, destabilizing this tumor suppressor. Rescue experiments confirmed that CDKN1C mediates the pro-tumorigenic effects of FBXL6 on lung adenocarcinoma cell proliferation and metastasis.","method":"Co-immunoprecipitation, ubiquitination assay, shRNA knockdown, overexpression, rescue assay, in vivo mouse xenograft","journal":"Experimental cell research","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — Co-IP and ubiquitination assay with rescue validation, but single lab and single study without independent replication","pmids":["41443404"],"is_preprint":false},{"year":2025,"finding":"FBXL6 promotes K63-linked polyubiquitination of ATAD3A (ATPase family AAA domain-containing protein 3A), stabilizing it and activating aerobic glycolysis to promote triple-negative breast cancer progression.","method":"Co-immunoprecipitation, ubiquitination assay, genetic depletion of ATAD3A and FBXL6, cell and animal models, Western blot, IHC","journal":"International journal of biological macromolecules","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — Co-IP and ubiquitination assay, but single lab and single study without independent replication","pmids":["40975350"],"is_preprint":false},{"year":2026,"finding":"FBXL6 directly interacts with ENO1 via its LRR domain (binding the C-terminal region of ENO1), promotes K63-linked polyubiquitination of ENO1, thereby stabilizing ENO1 and enhancing glycolytic activity to drive bladder cancer progression. ENO1 re-expression partially rescues the anti-proliferative and anti-migratory effects of FBXL6 knockdown.","method":"Co-immunoprecipitation, GST pull-down assay, ubiquitination assay, domain mapping, shRNA knockdown, ENO1 rescue experiments, transcriptomic and metabolic analyses","journal":"Cell death discovery","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — Co-IP and GST pull-down with domain mapping, ubiquitination assay, and rescue experiment; single lab, single study","pmids":["42091879"],"is_preprint":false}],"current_model":"FBXL6 is an F-box/LRR-domain E3 ubiquitin ligase (SCF complex subunit) that stabilizes multiple oncoproteins (HSP90AA1, KRAS/KRASG12D, ATAD3A, ENO1, TKT) via non-degradative K63-linked polyubiquitination while targeting tumor suppressors (p53-pS315, CDKN1C) for proteasomal degradation via K48-linked ubiquitination, and also ubiquitinates newly synthesized mitochondrial ribosomal proteins as part of a cotranslational quality control mechanism; collectively these activities promote cell proliferation, metabolic reprogramming, and oncogenesis across multiple cancer types."},"narrative":{"mechanistic_narrative":"FBXL6 is the substrate-recognition subunit of an SCF (Skp1-Cul1-F-box) E3 ubiquitin ligase complex that drives cell proliferation, metabolic reprogramming, and oncogenesis by deploying linkage-specific ubiquitination to opposite ends, stabilizing oncoproteins while degrading tumor suppressors [PMID:32576198, PMID:38124228]. Through non-degradative K63-linked polyubiquitination it stabilizes or activates a set of pro-tumorigenic clients including HSP90AA1, KRAS/KRAS-G12D (at Lys128, enhancing RAF binding and MEK/ERK/mTOR signaling), transketolase (recruited via VRK2-mediated Thr287 phosphorylation, driving ROS-mTOR/PD-L1 signaling), ATAD3A, and the glycolytic enzyme ENO1 (bound through the FBXL6 LRR domain) [PMID:32576198, PMID:38124228, PMID:37653031, PMID:40975350, PMID:42091879]. Several of these activities converge on glycolytic and ROS-mTOR metabolic rewiring across HCC, lung, breast, and bladder cancers [PMID:37653031, PMID:40975350, PMID:42091879]. In parallel, FBXL6 directs K48-linked polyubiquitination and proteasomal degradation of tumor suppressors, including Ser315-phosphorylated p53 and CDKN1C (p57Kip2) [PMID:33568778, PMID:41443404]. FBXL6 is embedded in transcriptional feedback circuits: stabilized HSP90AA1 prevents c-MYC degradation while c-MYC induces FBXL6, forming a positive loop, whereas p53 represses FBXL6 in a negative loop [PMID:32576198, PMID:33568778]. Distinct from its oncogenic substrate set, FBXL6 also performs cotranslational quality control by ubiquitinating newly synthesized, cytosolically translated mitochondrial ribosomal proteins in concert with chaperones and ribosome-associated quality control factors, preventing accumulation of mistranslated protein aggregates [PMID:37267103].","teleology":[{"year":2020,"claim":"Established that FBXL6 is an SCF substrate-recognition subunit and that, atypically for an F-box ligase, it can stabilize rather than degrade a substrate via K63-linked ubiquitination, embedding it in an oncogenic feedback loop.","evidence":"Reciprocal Co-IP, IP/MS, in vivo ubiquitination, luciferase and ChIP assays, shRNA knockdown in HCC cells and xenografts","pmids":["32576198"],"confidence":"High","gaps":["Linkage-specificity determinants on FBXL6 not mapped","Whether c-MYC-FBXL6 loop operates outside HCC unknown"]},{"year":2021,"claim":"Showed FBXL6 also acts as a classical degradative ligase, selectively recognizing phospho-Ser315 p53 for K48-linked degradation, defining a phospho-dependent route by which FBXL6 suppresses tumor-suppressor signaling.","evidence":"Co-IP, ubiquitination assay, proteasome inhibitor rescue, luciferase reporter, shRNA knockdown with cell-cycle/apoptosis readouts","pmids":["33568778"],"confidence":"High","gaps":["Kinase generating p53-pSer315 for FBXL6 recognition not identified","Reconciliation of degradative vs stabilizing modes by FBXL6 unresolved"]},{"year":2023,"claim":"Demonstrated FBXL6 ubiquitinates KRAS at Lys128 to activate it and engage RAF/MEK/ERK/mTOR signaling, extending FBXL6's stabilizing ubiquitination to a canonical oncogene and linking output to PRELID2/ROS.","evidence":"Co-IP, ubiquitination assay, site-specific K128 mutagenesis, RAS activity assay, transgenic mouse models, multiomics","pmids":["38124228"],"confidence":"High","gaps":["Mechanism by which K128 ubiquitination enhances RAF binding not structurally defined","Relationship between PRELID2-ROS and FBXL6 recruitment unclear"]},{"year":2023,"claim":"Connected FBXL6 to metabolic reprogramming and immune evasion by showing VRK2-phosphorylated TKT recruits FBXL6 for activating ubiquitination, driving ROS-mTOR-PD-L1 signaling and metastasis.","evidence":"Co-IP, ubiquitination and phosphorylation assays, knockdown/overexpression, transgenic mouse and xenograft models","pmids":["37653031"],"confidence":"High","gaps":["Ubiquitin linkage type on TKT not specified","Generality of the VRK2-TKT-FBXL6 axis beyond HCC untested"]},{"year":2023,"claim":"Revealed a non-oncogenic housekeeping role: FBXL6 ubiquitinates newly synthesized mitochondrial ribosomal proteins as cotranslational quality control, partnering with chaperones and RQC factors to prevent aggregate accumulation.","evidence":"Co-IP, ubiquitination assays, FBXL6-knockout cells, mitochondrial functional assays, proteomics, cell-cycle analysis","pmids":["37267103"],"confidence":"High","gaps":["Identity of specific chaperone/RQC partners not fully enumerated","Relationship between QC function and oncogenic substrate roles unknown"]},{"year":2022,"claim":"Placed c-MYC genetically downstream of FBXL6 in a non-cancer proliferative context (keloid fibroblasts), broadening FBXL6's proliferation-promoting role beyond tumors.","evidence":"shRNA knockdown, overexpression, c-MYC rescue, CCK-8 viability, Western blot, RT-PCR","pmids":["35606330"],"confidence":"Medium","gaps":["No direct biochemical interaction or ubiquitination assay in this system","Single lab, mechanism of c-MYC induction by FBXL6 not biochemically defined"]},{"year":2025,"claim":"Identified CDKN1C (p57Kip2) as a second degradative tumor-suppressor substrate of FBXL6, mediating its pro-tumorigenic effects in lung adenocarcinoma.","evidence":"Co-IP, ubiquitination assay, shRNA knockdown, overexpression, rescue, in vivo xenograft","pmids":["41443404"],"confidence":"Medium","gaps":["Single study without independent replication","Ubiquitin linkage and degron not defined"]},{"year":2025,"claim":"Extended FBXL6's stabilizing K63-ubiquitination to ATAD3A, linking FBXL6 to aerobic glycolysis in triple-negative breast cancer.","evidence":"Co-IP, ubiquitination assay, genetic depletion, cell and animal models, Western blot, IHC","pmids":["40975350"],"confidence":"Medium","gaps":["Single study, no independent replication","Mechanism linking ATAD3A stabilization to glycolysis not detailed"]},{"year":2026,"claim":"Mapped a direct LRR-domain interaction between FBXL6 and ENO1 driving K63-stabilizing ubiquitination and enhanced glycolysis in bladder cancer, providing domain-level evidence for substrate engagement.","evidence":"Co-IP, GST pull-down, domain mapping, ubiquitination assay, shRNA knockdown, ENO1 rescue, transcriptomic/metabolic analyses","pmids":["42091879"],"confidence":"Medium","gaps":["Single study without independent replication","Whether LRR-mediated binding generalizes to other FBXL6 substrates untested"]},{"year":null,"claim":"How FBXL6 selects between K63-linked stabilizing ubiquitination of oncoproteins and K48-linked degradative ubiquitination of tumor suppressors, and what dictates linkage choice per substrate, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structural model of FBXL6 substrate/E2 engagement","Linkage-determining cofactors or E2 enzymes not identified","Integration of QC and oncogenic functions unexplained"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,1,2,3,4,6,7,8]},{"term_id":"GO:0016874","term_label":"ligase activity","supporting_discovery_ids":[0,1,2,4]}],"localization":[],"pathway":[{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[0,1,4]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[0,1,2,6,7,8]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[2,3]},{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[3,7,8]}],"complexes":["SCF (Skp1-Cul1-F-box) E3 ubiquitin ligase"],"partners":["HSP90AA1","KRAS","TKT","CDKN1C","ATAD3A","ENO1","SKP1","CUL1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q8N531","full_name":"F-box/LRR-repeat protein 6","aliases":["F-box and leucine-rich repeat protein 6","F-box protein FBL6","FBL6A"],"length_aa":539,"mass_kda":58.6,"function":"Substrate-recognition component of the SCF (SKP1-CUL1-F-box protein)-type E3 ubiquitin ligase complex","subcellular_location":"","url":"https://www.uniprot.org/uniprotkb/Q8N531/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/FBXL6","classification":"Not Classified","n_dependent_lines":42,"n_total_lines":1208,"dependency_fraction":0.0347682119205298},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/FBXL6","total_profiled":1310},"omim":[{"mim_id":"609076","title":"F-BOX AND LEUCINE-RICH REPEAT PROTEIN 6; FBXL6","url":"https://www.omim.org/entry/609076"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Nucleoplasm","reliability":"Approved"},{"location":"Vesicles","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in many","driving_tissues":[],"url":"https://www.proteinatlas.org/search/FBXL6"},"hgnc":{"alias_symbol":["FBL6"],"prev_symbol":[]},"alphafold":{"accession":"Q8N531","domains":[{"cath_id":"-","chopping":"424-539","consensus_level":"medium","plddt":89.7234,"start":424,"end":539},{"cath_id":"1.10.10","chopping":"114-157","consensus_level":"medium","plddt":90.038,"start":114,"end":157}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8N531","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q8N531-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q8N531-F1-predicted_aligned_error_v6.png","plddt_mean":78.0},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=FBXL6","jax_strain_url":"https://www.jax.org/strain/search?query=FBXL6"},"sequence":{"accession":"Q8N531","fasta_url":"https://rest.uniprot.org/uniprotkb/Q8N531.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q8N531/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8N531"}},"corpus_meta":[{"pmid":"32576198","id":"PMC_32576198","title":"FBXL6 governs c-MYC to promote hepatocellular carcinoma through ubiquitination and stabilization of HSP90AA1.","date":"2020","source":"Cell communication and signaling : CCS","url":"https://pubmed.ncbi.nlm.nih.gov/32576198","citation_count":72,"is_preprint":false},{"pmid":"38124228","id":"PMC_38124228","title":"Elevated FBXL6 activates both wild-type KRAS and mutant KRASG12D and drives HCC tumorigenesis via the ERK/mTOR/PRELID2/ROS axis in mice.","date":"2023","source":"Military Medical Research","url":"https://pubmed.ncbi.nlm.nih.gov/38124228","citation_count":44,"is_preprint":false},{"pmid":"33568778","id":"PMC_33568778","title":"FBXL6 degrades phosphorylated p53 to promote tumor growth.","date":"2021","source":"Cell death and differentiation","url":"https://pubmed.ncbi.nlm.nih.gov/33568778","citation_count":30,"is_preprint":false},{"pmid":"37653031","id":"PMC_37653031","title":"Elevated FBXL6 expression in hepatocytes activates VRK2-transketolase-ROS-mTOR-mediated immune evasion and liver cancer metastasis in mice.","date":"2023","source":"Experimental & molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/37653031","citation_count":28,"is_preprint":false},{"pmid":"36183674","id":"PMC_36183674","title":"FBXL6 depletion restrains clear cell renal cell carcinoma progression.","date":"2022","source":"Translational oncology","url":"https://pubmed.ncbi.nlm.nih.gov/36183674","citation_count":9,"is_preprint":false},{"pmid":"35606330","id":"PMC_35606330","title":"FBXL6 is dysregulated in keloids and promotes keloid fibroblast growth by inducing c-Myc expression.","date":"2022","source":"International wound journal","url":"https://pubmed.ncbi.nlm.nih.gov/35606330","citation_count":8,"is_preprint":false},{"pmid":"37267103","id":"PMC_37267103","title":"The E3 ubiquitin ligase FBXL6 controls the quality of newly synthesized mitochondrial ribosomal proteins.","date":"2023","source":"Cell reports","url":"https://pubmed.ncbi.nlm.nih.gov/37267103","citation_count":5,"is_preprint":false},{"pmid":"40975350","id":"PMC_40975350","title":"Elevated FBXL6 activates ATAD3A through K63-linked polyubiquitination and promotes the malignant progression of TNBC via metabolic reprogramming.","date":"2025","source":"International journal of biological macromolecules","url":"https://pubmed.ncbi.nlm.nih.gov/40975350","citation_count":1,"is_preprint":false},{"pmid":"41443404","id":"PMC_41443404","title":"FBXL6 drives tumorigenesis in lung adenocarcinoma through ubiquitination and proteasomal degradation of CDKN1C.","date":"2025","source":"Experimental cell research","url":"https://pubmed.ncbi.nlm.nih.gov/41443404","citation_count":0,"is_preprint":false},{"pmid":"42091879","id":"PMC_42091879","title":"FBXL6 promotes bladder cancer progression by stabilizing ENO1 through K63-linked ubiquitination.","date":"2026","source":"Cell death discovery","url":"https://pubmed.ncbi.nlm.nih.gov/42091879","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":7643,"output_tokens":2544,"usd":0.030544,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9689,"output_tokens":3559,"usd":0.06871,"stage2_stop_reason":"end_turn"},"total_usd":0.099254,"stage1_batch_id":"msgbatch_01AsR7g8dGTEiEw9U83bJeWn","stage2_batch_id":"msgbatch_01YV5ZyxtFNUZEjxRcpd9jzb","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2020,\n      \"finding\": \"FBXL6 acts as a substrate recognition subunit of the SCF (Skp1-Cul1-F-box protein) E3 ubiquitin ligase complex and directly interacts with HSP90AA1, promoting its K63-linked polyubiquitination, which stabilizes (rather than degrades) HSP90AA1. Stabilized HSP90AA1 in turn prevents c-MYC degradation, and c-MYC transcriptionally induces FBXL6 expression by binding its promoter, forming a positive feedback loop.\",\n      \"method\": \"Co-immunoprecipitation, IP/Mass Spectrometry, in vivo ubiquitination assay, luciferase reporter assay, ChIP assay, shRNA knockdown in HCC cells and xenograft mouse model\",\n      \"journal\": \"Cell communication and signaling : CCS\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, IP/MS, in vivo ubiquitination assay, and transcriptional assays in one study with multiple orthogonal methods\",\n      \"pmids\": [\"32576198\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"FBXL6 directly targets phosphorylated p53 (phospho-Ser315) to mediate its K48-linked polyubiquitination and proteasomal degradation, thereby suppressing p53 signaling. Conversely, p53 transcriptionally represses FBXL6 expression by binding its core promoter, forming a negative feedback loop.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, proteasome inhibitor rescue experiments, luciferase reporter assay, shRNA knockdown with cell cycle and apoptosis readouts\",\n      \"journal\": \"Cell death and differentiation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, in vivo ubiquitination assay, and transcriptional assays with multiple orthogonal methods in one rigorous study\",\n      \"pmids\": [\"33568778\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"FBXL6 promotes polyubiquitination of both wild-type KRAS and mutant KRASG12D specifically at lysine 128, leading to their activation and enhanced binding to RAF kinase, thereby activating MEK/ERK/mTOR signaling. The oncogenic output of this axis depends on PRELID2-induced ROS generation.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, RAS activity detection assay, transgenic mouse models (LC, KC, KLC), multiomics, pharmacological inhibitors\",\n      \"journal\": \"Military Medical Research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — Co-IP, ubiquitination assay, site-specific mutagenesis (K128), RAS activity assay, and transgenic mouse models with multiple orthogonal methods\",\n      \"pmids\": [\"38124228\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"VRK2 kinase phosphorylates TKT (transketolase) at Thr287, which recruits FBXL6 to promote TKT ubiquitination and activation. Activated TKT drives ROS-mTOR signaling to upregulate PD-L1 and VRK2, leading to immune evasion and HCC metastasis.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, phosphorylation assays, knockdown/overexpression in vitro, transgenic mouse models, in vivo xenograft\",\n      \"journal\": \"Experimental & molecular medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — Co-IP, ubiquitination assay, phosphorylation site identification, and in vivo transgenic mouse models with multiple orthogonal methods\",\n      \"pmids\": [\"37653031\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"FBXL6 functions as an E3 ubiquitin ligase that specifically ubiquitinates newly synthesized (cytosolically translated) mitochondrial ribosomal proteins to control their quality. FBXL6 physically associates with chaperones involved in folding/trafficking of newly synthesized peptides and with ribosomal-associated quality control proteins; deletion of these interacting partners abolishes FBXL6-substrate interactions. FBXL6-knockout cells fail to degrade mistranslated mitochondrial ribosomal proteins, accumulate mitochondrial ribosomal protein aggregates, display altered mitochondrial metabolism, and exhibit inhibited cell cycle in oxidative conditions.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assays, FBXL6 knockout cells, mitochondrial functional assays, proteomics, cell cycle analysis\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (Co-IP, ubiquitination assay, KO phenotyping, mitochondrial functional assays) in one rigorous study\",\n      \"pmids\": [\"37267103\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"FBXL6 promotes keloid fibroblast proliferation by inducing c-MYC expression; FBXL6 knockdown reduces c-MYC levels along with cyclins A1, D2, E1, and Collagen I, and c-MYC overexpression rescues the proliferative defect caused by FBXL6 depletion, placing c-MYC downstream of FBXL6 in this pathway.\",\n      \"method\": \"shRNA knockdown, overexpression plasmid transfection, rescue experiment with c-MYC overexpression, cell viability assay (CCK-8), Western blot, RT-PCR\",\n      \"journal\": \"International wound journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — genetic epistasis via rescue experiment, Western blot, but no direct biochemical interaction or ubiquitination assay; single lab\",\n      \"pmids\": [\"35606330\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"FBXL6 directly binds CDKN1C (p57Kip2) and promotes its polyubiquitination and proteasomal degradation, destabilizing this tumor suppressor. Rescue experiments confirmed that CDKN1C mediates the pro-tumorigenic effects of FBXL6 on lung adenocarcinoma cell proliferation and metastasis.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, shRNA knockdown, overexpression, rescue assay, in vivo mouse xenograft\",\n      \"journal\": \"Experimental cell research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — Co-IP and ubiquitination assay with rescue validation, but single lab and single study without independent replication\",\n      \"pmids\": [\"41443404\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"FBXL6 promotes K63-linked polyubiquitination of ATAD3A (ATPase family AAA domain-containing protein 3A), stabilizing it and activating aerobic glycolysis to promote triple-negative breast cancer progression.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, genetic depletion of ATAD3A and FBXL6, cell and animal models, Western blot, IHC\",\n      \"journal\": \"International journal of biological macromolecules\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — Co-IP and ubiquitination assay, but single lab and single study without independent replication\",\n      \"pmids\": [\"40975350\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"FBXL6 directly interacts with ENO1 via its LRR domain (binding the C-terminal region of ENO1), promotes K63-linked polyubiquitination of ENO1, thereby stabilizing ENO1 and enhancing glycolytic activity to drive bladder cancer progression. ENO1 re-expression partially rescues the anti-proliferative and anti-migratory effects of FBXL6 knockdown.\",\n      \"method\": \"Co-immunoprecipitation, GST pull-down assay, ubiquitination assay, domain mapping, shRNA knockdown, ENO1 rescue experiments, transcriptomic and metabolic analyses\",\n      \"journal\": \"Cell death discovery\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — Co-IP and GST pull-down with domain mapping, ubiquitination assay, and rescue experiment; single lab, single study\",\n      \"pmids\": [\"42091879\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"FBXL6 is an F-box/LRR-domain E3 ubiquitin ligase (SCF complex subunit) that stabilizes multiple oncoproteins (HSP90AA1, KRAS/KRASG12D, ATAD3A, ENO1, TKT) via non-degradative K63-linked polyubiquitination while targeting tumor suppressors (p53-pS315, CDKN1C) for proteasomal degradation via K48-linked ubiquitination, and also ubiquitinates newly synthesized mitochondrial ribosomal proteins as part of a cotranslational quality control mechanism; collectively these activities promote cell proliferation, metabolic reprogramming, and oncogenesis across multiple cancer types.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"FBXL6 is the substrate-recognition subunit of an SCF (Skp1-Cul1-F-box) E3 ubiquitin ligase complex that drives cell proliferation, metabolic reprogramming, and oncogenesis by deploying linkage-specific ubiquitination to opposite ends, stabilizing oncoproteins while degrading tumor suppressors [#0, #2]. Through non-degradative K63-linked polyubiquitination it stabilizes or activates a set of pro-tumorigenic clients including HSP90AA1, KRAS/KRAS-G12D (at Lys128, enhancing RAF binding and MEK/ERK/mTOR signaling), transketolase (recruited via VRK2-mediated Thr287 phosphorylation, driving ROS-mTOR/PD-L1 signaling), ATAD3A, and the glycolytic enzyme ENO1 (bound through the FBXL6 LRR domain) [#0, #2, #3, #7, #8]. Several of these activities converge on glycolytic and ROS-mTOR metabolic rewiring across HCC, lung, breast, and bladder cancers [#3, #7, #8]. In parallel, FBXL6 directs K48-linked polyubiquitination and proteasomal degradation of tumor suppressors, including Ser315-phosphorylated p53 and CDKN1C (p57Kip2) [#1, #6]. FBXL6 is embedded in transcriptional feedback circuits: stabilized HSP90AA1 prevents c-MYC degradation while c-MYC induces FBXL6, forming a positive loop, whereas p53 represses FBXL6 in a negative loop [#0, #1]. Distinct from its oncogenic substrate set, FBXL6 also performs cotranslational quality control by ubiquitinating newly synthesized, cytosolically translated mitochondrial ribosomal proteins in concert with chaperones and ribosome-associated quality control factors, preventing accumulation of mistranslated protein aggregates [#4].\",\n  \"teleology\": [\n    {\n      \"year\": 2020,\n      \"claim\": \"Established that FBXL6 is an SCF substrate-recognition subunit and that, atypically for an F-box ligase, it can stabilize rather than degrade a substrate via K63-linked ubiquitination, embedding it in an oncogenic feedback loop.\",\n      \"evidence\": \"Reciprocal Co-IP, IP/MS, in vivo ubiquitination, luciferase and ChIP assays, shRNA knockdown in HCC cells and xenografts\",\n      \"pmids\": [\"32576198\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Linkage-specificity determinants on FBXL6 not mapped\", \"Whether c-MYC-FBXL6 loop operates outside HCC unknown\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Showed FBXL6 also acts as a classical degradative ligase, selectively recognizing phospho-Ser315 p53 for K48-linked degradation, defining a phospho-dependent route by which FBXL6 suppresses tumor-suppressor signaling.\",\n      \"evidence\": \"Co-IP, ubiquitination assay, proteasome inhibitor rescue, luciferase reporter, shRNA knockdown with cell-cycle/apoptosis readouts\",\n      \"pmids\": [\"33568778\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Kinase generating p53-pSer315 for FBXL6 recognition not identified\", \"Reconciliation of degradative vs stabilizing modes by FBXL6 unresolved\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Demonstrated FBXL6 ubiquitinates KRAS at Lys128 to activate it and engage RAF/MEK/ERK/mTOR signaling, extending FBXL6's stabilizing ubiquitination to a canonical oncogene and linking output to PRELID2/ROS.\",\n      \"evidence\": \"Co-IP, ubiquitination assay, site-specific K128 mutagenesis, RAS activity assay, transgenic mouse models, multiomics\",\n      \"pmids\": [\"38124228\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism by which K128 ubiquitination enhances RAF binding not structurally defined\", \"Relationship between PRELID2-ROS and FBXL6 recruitment unclear\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Connected FBXL6 to metabolic reprogramming and immune evasion by showing VRK2-phosphorylated TKT recruits FBXL6 for activating ubiquitination, driving ROS-mTOR-PD-L1 signaling and metastasis.\",\n      \"evidence\": \"Co-IP, ubiquitination and phosphorylation assays, knockdown/overexpression, transgenic mouse and xenograft models\",\n      \"pmids\": [\"37653031\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Ubiquitin linkage type on TKT not specified\", \"Generality of the VRK2-TKT-FBXL6 axis beyond HCC untested\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Revealed a non-oncogenic housekeeping role: FBXL6 ubiquitinates newly synthesized mitochondrial ribosomal proteins as cotranslational quality control, partnering with chaperones and RQC factors to prevent aggregate accumulation.\",\n      \"evidence\": \"Co-IP, ubiquitination assays, FBXL6-knockout cells, mitochondrial functional assays, proteomics, cell-cycle analysis\",\n      \"pmids\": [\"37267103\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Identity of specific chaperone/RQC partners not fully enumerated\", \"Relationship between QC function and oncogenic substrate roles unknown\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Placed c-MYC genetically downstream of FBXL6 in a non-cancer proliferative context (keloid fibroblasts), broadening FBXL6's proliferation-promoting role beyond tumors.\",\n      \"evidence\": \"shRNA knockdown, overexpression, c-MYC rescue, CCK-8 viability, Western blot, RT-PCR\",\n      \"pmids\": [\"35606330\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No direct biochemical interaction or ubiquitination assay in this system\", \"Single lab, mechanism of c-MYC induction by FBXL6 not biochemically defined\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Identified CDKN1C (p57Kip2) as a second degradative tumor-suppressor substrate of FBXL6, mediating its pro-tumorigenic effects in lung adenocarcinoma.\",\n      \"evidence\": \"Co-IP, ubiquitination assay, shRNA knockdown, overexpression, rescue, in vivo xenograft\",\n      \"pmids\": [\"41443404\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single study without independent replication\", \"Ubiquitin linkage and degron not defined\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Extended FBXL6's stabilizing K63-ubiquitination to ATAD3A, linking FBXL6 to aerobic glycolysis in triple-negative breast cancer.\",\n      \"evidence\": \"Co-IP, ubiquitination assay, genetic depletion, cell and animal models, Western blot, IHC\",\n      \"pmids\": [\"40975350\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single study, no independent replication\", \"Mechanism linking ATAD3A stabilization to glycolysis not detailed\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Mapped a direct LRR-domain interaction between FBXL6 and ENO1 driving K63-stabilizing ubiquitination and enhanced glycolysis in bladder cancer, providing domain-level evidence for substrate engagement.\",\n      \"evidence\": \"Co-IP, GST pull-down, domain mapping, ubiquitination assay, shRNA knockdown, ENO1 rescue, transcriptomic/metabolic analyses\",\n      \"pmids\": [\"42091879\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single study without independent replication\", \"Whether LRR-mediated binding generalizes to other FBXL6 substrates untested\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How FBXL6 selects between K63-linked stabilizing ubiquitination of oncoproteins and K48-linked degradative ubiquitination of tumor suppressors, and what dictates linkage choice per substrate, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structural model of FBXL6 substrate/E2 engagement\", \"Linkage-determining cofactors or E2 enzymes not identified\", \"Integration of QC and oncogenic functions unexplained\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 1, 2, 3, 4, 6, 7, 8]},\n      {\"term_id\": \"GO:0016874\", \"supporting_discovery_ids\": [0, 1, 2, 4]}\n    ],\n    \"localization\": [],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [0, 1, 4]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [0, 1, 2, 6, 7, 8]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [2, 3]},\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [3, 7, 8]}\n    ],\n    \"complexes\": [\"SCF (Skp1-Cul1-F-box) E3 ubiquitin ligase\"],\n    \"partners\": [\"HSP90AA1\", \"KRAS\", \"TKT\", \"CDKN1C\", \"ATAD3A\", \"ENO1\", \"SKP1\", \"CUL1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}