{"gene":"CCNF","run_date":"2026-04-28T17:28:52","timeline":{"discoveries":[{"year":1994,"finding":"CCNF encodes cyclin F, a novel member of the cyclin family related to A- and B-type cyclins by sequence, located on chromosome 16p13.3.","method":"cDNA sequencing, Northern blot analysis, exon-intron boundary determination","journal":"Genomics","confidence":"Medium","confidence_rationale":"Tier 2 — original characterization of gene/transcript; sequence homology to cyclins established but function unknown at time of publication","pmids":["7896286"],"is_preprint":false},{"year":2016,"finding":"Cyclin F (CCNF) functions as the substrate-binding subunit of an SCF (SKP1-CUL1-F-box) E3 ubiquitin-protein ligase complex (SCF^CyclinF); ALS/FTD-associated missense mutations cause abnormal ubiquitination and accumulation of ubiquitinated proteins including TDP-43 and an SCF^CyclinF substrate in neuronal cells.","method":"Whole-exome sequencing for mutation identification; transfection of mutant CCNF in neuronal cells with ubiquitination assays and western blot for substrate accumulation","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 — highly cited foundational study (178 citations), multiple orthogonal methods, functional assays in neuronal cells","pmids":["27080313"],"is_preprint":false},{"year":2017,"finding":"The ALS/FTD-linked cyclin F p.S621G mutation causes elevated Lys48-linked ubiquitylation of proteins in neuronal cells, and proteomic analysis identified autophagy pathway proteins (p62/SQSTM1, heat shock proteins, chaperonin components) as targets; mutant cyclin F impairs autophagosomal-lysosome fusion, and cyclin F physically interacts with p62.","method":"Transfection of mutant CCNF (S621G) in Neuro-2A and SH-SY5Y cells; K48-linkage-specific ubiquitin immunoprecipitation followed by mass spectrometry proteomics; autophagy marker analysis (p62, LC3, Lamp2) by immunoblot and immunofluorescence","journal":"Cellular and molecular life sciences : CMLS","confidence":"High","confidence_rationale":"Tier 2 — multiple orthogonal methods (MS proteomics, specific K48-Ub IP, autophagy flux markers, Co-IP of cyclin F with p62)","pmids":["28852778"],"is_preprint":false},{"year":2017,"finding":"Cyclin F (CCNF) interacts with HIV-1 Vif protein; Vif is a substrate of the SCF^CyclinF E3 ligase complex, which mediates K48-linked ubiquitination and proteasomal degradation of Vif, thereby restoring APOBEC3G levels and restricting viral infectivity. A cyclin F-specific amino acid motif in the C-terminal region of Vif is required for this interaction.","method":"Co-immunoprecipitation, overexpression and knockdown studies, mutational analysis of Vif degron motif, ubiquitination assays, proteasome inhibitor experiments, APOBEC3G expression analysis","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1–2 — reciprocal Co-IP, mutagenesis of degron, ubiquitination assay, multiple orthogonal methods in a single study","pmids":["28184007"],"is_preprint":false},{"year":2017,"finding":"Expression of ALS-linked mutant CCNF in zebrafish embryos causes increased caspase-3 activation and cell death in the spinal cord, motor neuron axonopathy (shortened primary motor axons, aberrant branching), and reduced photomotor response; proteomic analysis of in vitro models identified disruption of caspase-3-mediated cell death pathways.","method":"Transient overexpression of human mutant CCNF in zebrafish embryos; immunostaining for cleaved caspase-3; motor response (photomotor response) assay; label-free quantitative proteomics of in vitro models","journal":"Human molecular genetics","confidence":"Medium","confidence_rationale":"Tier 2 — in vivo zebrafish model with quantitative phenotypic readouts and proteomics; single lab","pmids":["28444311"],"is_preprint":false},{"year":2019,"finding":"Cyclin F physically binds to VCP (valosin-containing protein, also an ALS gene) via the N-terminal region of Cyclin F, and the two proteins colocalize in the nucleus. Cyclin F enhances VCP ATPase activity in vitro. ALS-associated CCNF mutations increase Cyclin F binding to VCP and further elevate VCP ATPase activity while causing cytoplasmic mislocalization of Cyclin F. Elevated VCP ATPase activity promotes cytoplasmic TDP-43 aggregation.","method":"Co-immunoprecipitation and colocalization experiments; domain-mapping pulldowns; in vitro ATPase activity assay; overexpression of mutant CCNF in transfected cells; TDP-43 aggregation assay","journal":"Human molecular genetics","confidence":"High","confidence_rationale":"Tier 1–2 — direct in vitro ATPase assay, domain-mapping Co-IP, and functional consequence (TDP-43 aggregation) measured with multiple methods in one study","pmids":["31577344"],"is_preprint":false},{"year":2021,"finding":"Label-free quantitative proteomics of HEK293 cells expressing multiple ALS-associated CCNF mutations (K97R, S195R, S509P, R574Q, S621G) bioinformatically predicted and immunoblot-validated activation of neuronal apoptosis pathways; iPSC-derived cells from S621G patients showed the same pathway activation.","method":"Label-free quantitative proteomics of transfected HEK293 cells; pathway bioinformatics; immunoblot validation; iPSC-derived patient cell proteomics","journal":"Frontiers in molecular neuroscience","confidence":"Medium","confidence_rationale":"Tier 2–3 — proteomics with immunoblot validation across multiple mutations and patient-derived iPSCs, single lab","pmids":["33986643"],"is_preprint":false},{"year":2021,"finding":"CCNF protein is targeted for degradation by the E3 ligases FBXL8 and FZR1 (demonstrated by Co-IP pulldown); double knockdown of FBXL8 and FZR1 causes CCNF accumulation. CCNF itself pulls down RRM2 (ribonucleotide reductase subunit 2) and CCNF overexpression reduces RRM2 levels, indicating RRM2 is a substrate of SCF^CyclinF.","method":"Co-immunoprecipitation (FBXL8 and FZR1 pulldown of CCNF); double knockdown experiments; CCNF overexpression with RRM2 protein level measurement","journal":"Cancers","confidence":"Medium","confidence_rationale":"Tier 3 — Co-IP and overexpression/knockdown; single lab, moderate orthogonality","pmids":["34201347"],"is_preprint":false},{"year":2023,"finding":"FBXO1 (Cyclin F) directly binds E2F1, E2F2, and E2F3a transcription factors through Arg/Ile and Arg/Val degron motifs in their dimerization domains, mediating K48-linked ubiquitination and proteasomal degradation of E2Fs. MEK/ERK-dependent phosphorylation of threonine residues near these degron motifs regulates FBXO1-E2F interaction and E2F protein stability. Knockdown of FBXO1 elevated E2F levels and delayed G1/S cell cycle transition, inhibiting cancer cell proliferation.","method":"Co-immunoprecipitation; domain/motif mutation analysis (RI/AA, RV/AA); ubiquitination assays; cycloheximide chase for half-life; specific kinase inhibitors; FBXO1 knockdown with cell cycle analysis","journal":"Archives of pharmacal research","confidence":"High","confidence_rationale":"Tier 1–2 — reciprocal Co-IP, mutagenesis of degron motifs, ubiquitination assay, half-life measurement, and cell cycle functional readout in one study","pmids":["36607545"],"is_preprint":false},{"year":2023,"finding":"The ALS-associated CCNF S621G variant causes ubiquitin-proteasome system (UPS) dysfunction in iPSC-derived motor neurons, with increased ubiquitinated protein abundance and altered ubiquitination of key UPS components. Overexpression of CCNF in NSC-34 cells alters free ubiquitin levels; double mutations that reduce CCNF's ability to form an active E3 ligase complex improved UPS function and increased free monomeric ubiquitin, establishing that the E3 ligase activity of CCNF is central to its role in ubiquitin homeostasis.","method":"iPSC-derived motor neurons from CCNF S621G patients; ubiquitin abundance and UPS component ubiquitination analysis; overexpression in NSC-34 cells with free ubiquitin measurement; active-site double-mutant analysis with UPS functional assay","journal":"Human molecular genetics","confidence":"High","confidence_rationale":"Tier 1–2 — patient-derived iPSC neurons, active-site mutagenesis with functional rescue, multiple orthogonal methods in one study","pmids":["37220877"],"is_preprint":false},{"year":2024,"finding":"CRISPR/Cas9-mediated loss of ccnf in zebrafish causes abnormal motor neuron development and axonal outgrowth defects, and ccnf-deficient zebrafish show selective sensitization to endoplasmic reticulum stress but not oxidative stress, establishing a direct role for CCNF in motor neuron axonal maintenance in vivo.","method":"CRISPR/Cas9 genome editing in zebrafish to generate ccnf knockout; motor neuron morphology and axonal outgrowth analysis; pharmacological stress (ER stress, oxidative stress) with motor response readout","journal":"Cells","confidence":"Medium","confidence_rationale":"Tier 2 — clean genetic knockout in vivo with defined morphological and stress-response phenotypes; single lab","pmids":["38474336"],"is_preprint":false}],"current_model":"Cyclin F (CCNF/FBXO1) is the substrate-recognition subunit of the SCF^CyclinF E3 ubiquitin ligase complex that mediates K48-linked ubiquitination and proteasomal degradation of substrates including E2F transcription factors, HIV-1 Vif, and RRM2; it physically interacts with and stimulates the ATPase activity of VCP; ALS/FTD-associated CCNF mutations dysregulate these activities, causing elevated K48-ubiquitylation, impaired autophagosome-lysosome fusion, cytoplasmic VCP hyperactivation driving TDP-43 aggregation, and disruption of free ubiquitin homeostasis in motor neurons."},"narrative":{"teleology":[{"year":1994,"claim":"Identification of CCNF as a novel cyclin-family member related to A- and B-type cyclins established the gene as a cell-cycle-associated factor, but its specific biochemical function was unknown.","evidence":"cDNA sequencing and Northern blot analysis of human transcript","pmids":["7896286"],"confidence":"Medium","gaps":["No enzymatic or functional activity defined","No interacting partners identified","Cyclin-box homology alone did not establish CDK dependency"]},{"year":2016,"claim":"Discovery that cyclin F is the substrate-recognition (F-box) subunit of an SCF E3 ubiquitin ligase, and that ALS/FTD-associated mutations cause accumulation of ubiquitinated proteins including TDP-43 in neuronal cells, established CCNF as a disease-linked ubiquitin ligase component.","evidence":"Whole-exome sequencing of ALS/FTD families; mutant CCNF expression in neuronal cells with ubiquitination assays and immunoblot","pmids":["27080313"],"confidence":"High","gaps":["Endogenous substrates beyond TDP-43 not identified","Mechanism by which mutations alter ligase activity undefined","No in vivo validation of motor neuron phenotype"]},{"year":2017,"claim":"Characterization of the S621G mutant revealed that disease-linked cyclin F elevates Lys48-ubiquitylation of autophagy-related proteins, physically interacts with p62/SQSTM1, and impairs autophagosome–lysosome fusion, connecting SCF^CyclinF dysfunction to proteostasis failure via autophagy.","evidence":"K48-linkage-specific ubiquitin IP with mass spectrometry; autophagy flux markers (LC3, Lamp2) in Neuro-2A and SH-SY5Y cells; Co-IP of cyclin F and p62","pmids":["28852778"],"confidence":"High","gaps":["Whether p62 is a direct ubiquitination substrate or an adaptor not resolved","Autophagy defect not confirmed in patient-derived neurons at this stage"]},{"year":2017,"claim":"Demonstration that SCF^CyclinF targets HIV-1 Vif for K48-linked ubiquitination and proteasomal degradation — via a cyclin F-specific degron motif in Vif — expanded the substrate repertoire to viral proteins and revealed host antiviral restriction function.","evidence":"Reciprocal Co-IP, degron mutagenesis, ubiquitination assays, proteasome inhibitor rescue, APOBEC3G restoration upon Vif degradation","pmids":["28184007"],"confidence":"High","gaps":["Physiological relevance in HIV-infected patients not tested","Degron specificity for other viral substrates unknown"]},{"year":2017,"claim":"Expression of ALS-linked CCNF mutants in zebrafish embryos caused motor neuron axonopathy, spinal cord apoptosis, and impaired motor responses, providing the first in vivo evidence that mutant CCNF is neurotoxic.","evidence":"Transient overexpression of mutant human CCNF in zebrafish; caspase-3 immunostaining; photomotor response assay","pmids":["28444311"],"confidence":"Medium","gaps":["Overexpression model does not distinguish gain-of-function from dominant-negative effects","Endogenous loss-of-function phenotype not assessed","No mammalian in vivo model"]},{"year":2019,"claim":"Discovery that cyclin F directly binds and stimulates VCP ATPase activity, and that ALS mutations enhance this interaction causing cytoplasmic mislocalization and VCP hyperactivation that drives TDP-43 aggregation, provided a concrete pathogenic mechanism linking two ALS genes.","evidence":"Co-IP and colocalization; domain-mapping pulldowns; in vitro VCP ATPase activity assay; TDP-43 aggregation assay in transfected cells","pmids":["31577344"],"confidence":"High","gaps":["Structural basis of cyclin F–VCP interface unresolved","Whether VCP is itself a ubiquitination substrate of SCF^CyclinF unknown","TDP-43 aggregation not validated in patient-derived neurons"]},{"year":2021,"claim":"Identification of RRM2 as an SCF^CyclinF substrate and of FBXL8/FZR1 as E3 ligases that target CCNF itself for degradation established bidirectional regulation of cyclin F abundance and a new substrate relevant to DNA replication.","evidence":"Co-IP of FBXL8/FZR1 with CCNF; double knockdown causing CCNF accumulation; CCNF overexpression reducing RRM2 levels","pmids":["34201347"],"confidence":"Medium","gaps":["Direct ubiquitination of RRM2 by SCF^CyclinF not demonstrated with purified components","Physiological contexts (cell cycle phase) not resolved","Single-lab finding awaiting independent replication"]},{"year":2023,"claim":"Identification of E2F1/E2F2/E2F3a as direct SCF^CyclinF substrates, degraded via specific Arg/Ile and Arg/Val degron motifs regulated by MEK/ERK phosphorylation, established cyclin F as a master regulator of G1/S progression through E2F turnover.","evidence":"Reciprocal Co-IP; degron motif mutagenesis (RI/AA, RV/AA); ubiquitination and cycloheximide chase assays; MEK/ERK inhibitor treatment; FBXO1 knockdown with cell-cycle analysis","pmids":["36607545"],"confidence":"High","gaps":["In vivo cell-cycle phenotype of CCNF loss in mammals not examined","Whether all three E2Fs are targeted simultaneously or in different contexts is unclear"]},{"year":2023,"claim":"Patient iPSC-derived motor neurons carrying CCNF S621G showed increased ubiquitinated protein loads and altered UPS component ubiquitination; active-site double mutations restoring ligase function rescued free ubiquitin levels, proving that SCF^CyclinF E3 ligase activity directly controls ubiquitin homeostasis in motor neurons.","evidence":"iPSC-derived motor neurons from CCNF S621G patients; ubiquitin profiling; active-site double-mutant rescue in NSC-34 cells","pmids":["37220877"],"confidence":"High","gaps":["Specific ubiquitin-modified substrates driving motor neuron vulnerability not identified","Rescue not performed in patient neurons","Relationship between UPS dysfunction and autophagy defects not integrated"]},{"year":2024,"claim":"CRISPR knockout of ccnf in zebrafish confirmed a direct requirement for CCNF in motor neuron axonal outgrowth and revealed selective vulnerability to ER stress but not oxidative stress, distinguishing the stress-response pathways downstream of CCNF loss.","evidence":"CRISPR/Cas9 ccnf knockout zebrafish; motor neuron morphology; pharmacological ER and oxidative stress challenges","pmids":["38474336"],"confidence":"Medium","gaps":["Mechanism linking CCNF loss to ER stress sensitivity unknown","Mammalian knockout model still lacking","Transcriptomic/proteomic characterization of knockout neurons not performed"]},{"year":null,"claim":"Key unresolved questions include the structural basis of SCF^CyclinF substrate recognition and VCP interaction, the full catalogue of physiological substrates in motor neurons, the relative contributions of UPS dysfunction versus autophagy impairment versus VCP hyperactivation to ALS/FTD pathogenesis, and validation of these mechanisms in mammalian in vivo models.","evidence":"","pmids":[],"confidence":"Low","gaps":["No crystal or cryo-EM structure of SCF^CyclinF with any substrate","No mammalian in vivo conditional knockout model","Integration of UPS, autophagy, and VCP pathways into unified pathogenic cascade not achieved"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[1,2,3,8,9]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[5,8]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[5]}],"pathway":[{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[1,2,3,8,9]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[8]},{"term_id":"R-HSA-9612973","term_label":"Autophagy","supporting_discovery_ids":[2]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[4,6]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[1,5,9]}],"complexes":["SCF^CyclinF (SKP1-CUL1-CyclinF)"],"partners":["SKP1","CUL1","VCP","SQSTM1","E2F1","E2F2","E2F3","RRM2"],"other_free_text":[]},"mechanistic_narrative":"Cyclin F (CCNF/FBXO1) is the substrate-recognition subunit of the SCF^CyclinF E3 ubiquitin ligase complex, which mediates Lys48-linked ubiquitination and proteasomal degradation of key cell-cycle and homeostatic substrates including E2F1/E2F2/E2F3a transcription factors, RRM2, and HIV-1 Vif [PMID:36607545, PMID:34201347, PMID:28184007]. Through recognition of specific degron motifs in substrates — regulated by MEK/ERK-dependent phosphorylation in the case of E2Fs — SCF^CyclinF controls G1/S cell-cycle progression, ubiquitin-proteasome system homeostasis, and autophagosome–lysosome fusion, and it modulates VCP ATPase activity via direct physical interaction [PMID:36607545, PMID:37220877, PMID:28852778, PMID:31577344]. Missense mutations in CCNF cause familial ALS/FTD by elevating aberrant Lys48-ubiquitylation, disrupting free ubiquitin pools in motor neurons, hyperactivating cytoplasmic VCP to promote TDP-43 aggregation, and impairing autophagic clearance [PMID:27080313, PMID:37220877, PMID:31577344, PMID:28852778]. Loss of CCNF in zebrafish produces motor neuron axonal outgrowth defects and selective vulnerability to endoplasmic reticulum stress, confirming a direct requirement for CCNF in motor neuron maintenance [PMID:38474336]."},"prefetch_data":{"uniprot":{"accession":"P41002","full_name":"Cyclin-F","aliases":["F-box only protein 1"],"length_aa":786,"mass_kda":87.6,"function":"Substrate recognition component of a SCF (SKP1-CUL1-F-box protein) E3 ubiquitin-protein ligase complex which mediates the ubiquitination and subsequent proteasomal degradation of target proteins (PubMed:20596027, PubMed:22632967, PubMed:26818844, PubMed:27080313, PubMed:27653696, PubMed:28852778). The SCF(CCNF) E3 ubiquitin-protein ligase complex is an integral component of the ubiquitin proteasome system (UPS) and links proteasome degradation to the cell cycle (PubMed:20596027, PubMed:26818844, PubMed:27653696, PubMed:8706131). Mediates the substrate recognition and the proteasomal degradation of various target proteins involved in the regulation of cell cycle progression and in the maintenance of genome stability (PubMed:20596027, PubMed:22632967, PubMed:26818844, PubMed:27653696). Mediates the ubiquitination and proteasomal degradation of CP110 during G2 phase, thereby acting as an inhibitor of centrosome reduplication (PubMed:20596027). In G2, mediates the ubiquitination and subsequent degradation of ribonucleotide reductase RRM2, thereby maintaining a balanced pool of dNTPs and genome integrity (PubMed:22632967). In G2, mediates the ubiquitination and proteasomal degradation of CDC6, thereby suppressing DNA re-replication and preventing genome instability (PubMed:26818844). Involved in the ubiquitination and degradation of the substrate adapter CDH1 of the anaphase-promoting complex (APC/C), thereby acting as an antagonist of APC/C in regulating G1 progression and S phase entry (PubMed:27653696). May play a role in the G2 cell cycle checkpoint control after DNA damage, possibly by promoting the ubiquitination of MYBL2/BMYB (PubMed:25557911)","subcellular_location":"Nucleus; Cytoplasm, perinuclear region; Cytoplasm, cytoskeleton, microtubule organizing center, centrosome, centriole","url":"https://www.uniprot.org/uniprotkb/P41002/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/CCNF","classification":"Not Classified","n_dependent_lines":104,"n_total_lines":1208,"dependency_fraction":0.08609271523178808},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/CCNF","total_profiled":1310},"omim":[{"mim_id":"619141","title":"FRONTOTEMPORAL DEMENTIA AND/OR AMYOTROPHIC LATERAL SCLEROSIS 5; FTDALS5","url":"https://www.omim.org/entry/619141"},{"mim_id":"607112","title":"F-BOX ONLY PROTEIN 2; FBXO2","url":"https://www.omim.org/entry/607112"},{"mim_id":"605657","title":"LYSINE DEMETHYLASE 2A; KDM2A","url":"https://www.omim.org/entry/605657"},{"mim_id":"605656","title":"F-BOX AND LEUCINE-RICH REPEAT PROTEIN 7; FBXL7","url":"https://www.omim.org/entry/605656"},{"mim_id":"605655","title":"F-BOX AND LEUCINE-RICH REPEAT PROTEIN 5; FBXL5","url":"https://www.omim.org/entry/605655"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nucleoplasm","reliability":"Supported"},{"location":"Centrosome","reliability":"Additional"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"lymphoid tissue","ntpm":14.1}],"url":"https://www.proteinatlas.org/search/CCNF"},"hgnc":{"alias_symbol":["FBX1","FBXO1"],"prev_symbol":[]},"alphafold":{"accession":"P41002","domains":[{"cath_id":"-","chopping":"79-225","consensus_level":"high","plddt":90.1878,"start":79,"end":225},{"cath_id":"-","chopping":"237-283","consensus_level":"medium","plddt":86.5538,"start":237,"end":283},{"cath_id":"1.10.472.10","chopping":"291-408","consensus_level":"medium","plddt":94.0562,"start":291,"end":408},{"cath_id":"1.10.472.10","chopping":"409-544","consensus_level":"medium","plddt":94.0223,"start":409,"end":544}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P41002","model_url":"https://alphafold.ebi.ac.uk/files/AF-P41002-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P41002-F1-predicted_aligned_error_v6.png","plddt_mean":71.56},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=CCNF","jax_strain_url":"https://www.jax.org/strain/search?query=CCNF"},"sequence":{"accession":"P41002","fasta_url":"https://rest.uniprot.org/uniprotkb/P41002.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P41002/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P41002"}},"corpus_meta":[{"pmid":"27080313","id":"PMC_27080313","title":"CCNF mutations in amyotrophic lateral sclerosis and frontotemporal dementia.","date":"2016","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/27080313","citation_count":178,"is_preprint":false},{"pmid":"28852778","id":"PMC_28852778","title":"Pathogenic mutation in the ALS/FTD gene, CCNF, causes elevated Lys48-linked ubiquitylation and defective autophagy.","date":"2017","source":"Cellular and molecular life sciences : CMLS","url":"https://pubmed.ncbi.nlm.nih.gov/28852778","citation_count":43,"is_preprint":false},{"pmid":"28444311","id":"PMC_28444311","title":"Expression of ALS/FTD-linked mutant CCNF in zebrafish leads to increased cell death in the spinal cord and an aberrant motor phenotype.","date":"2017","source":"Human molecular genetics","url":"https://pubmed.ncbi.nlm.nih.gov/28444311","citation_count":40,"is_preprint":false},{"pmid":"31577344","id":"PMC_31577344","title":"Pathogenic mutations in the ALS gene CCNF cause cytoplasmic mislocalization of Cyclin F and elevated VCP ATPase activity.","date":"2019","source":"Human molecular genetics","url":"https://pubmed.ncbi.nlm.nih.gov/31577344","citation_count":28,"is_preprint":false},{"pmid":"28184007","id":"PMC_28184007","title":"Cyclin F/FBXO1 Interacts with HIV-1 Viral Infectivity Factor (Vif) and Restricts Progeny Virion Infectivity by Ubiquitination and Proteasomal Degradation of Vif Protein through SCFcyclin F E3 Ligase Machinery.","date":"2017","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/28184007","citation_count":25,"is_preprint":false},{"pmid":"28281833","id":"PMC_28281833","title":"Mutations of CCNF gene is rare in patients with amyotrophic lateral sclerosis and frontotemporal dementia from Mainland China.","date":"2017","source":"Amyotrophic lateral sclerosis & frontotemporal degeneration","url":"https://pubmed.ncbi.nlm.nih.gov/28281833","citation_count":17,"is_preprint":false},{"pmid":"34201347","id":"PMC_34201347","title":"A Novel Signature of CCNF-Associated E3 Ligases Collaborate and Counter Each Other in Breast Cancer.","date":"2021","source":"Cancers","url":"https://pubmed.ncbi.nlm.nih.gov/34201347","citation_count":17,"is_preprint":false},{"pmid":"29102476","id":"PMC_29102476","title":"Investigating CCNF mutations in a Taiwanese cohort with amyotrophic lateral sclerosis.","date":"2017","source":"Neurobiology of aging","url":"https://pubmed.ncbi.nlm.nih.gov/29102476","citation_count":17,"is_preprint":false},{"pmid":"36898988","id":"PMC_36898988","title":"The Skp1-Cullin1-FBXO1 complex is a pleiotropic regulator required for the formation of gametes and motile forms in Plasmodium berghei.","date":"2023","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/36898988","citation_count":15,"is_preprint":false},{"pmid":"33986643","id":"PMC_33986643","title":"Unbiased Label-Free Quantitative Proteomics of Cells Expressing Amyotrophic Lateral Sclerosis (ALS) Mutations in CCNF Reveals Activation of the Apoptosis Pathway: A Workflow to Screen Pathogenic Gene Mutations.","date":"2021","source":"Frontiers in molecular neuroscience","url":"https://pubmed.ncbi.nlm.nih.gov/33986643","citation_count":14,"is_preprint":false},{"pmid":"7896286","id":"PMC_7896286","title":"A novel cyclin gene (CCNF) in the region of the polycystic kidney disease gene (PKD1).","date":"1994","source":"Genomics","url":"https://pubmed.ncbi.nlm.nih.gov/7896286","citation_count":14,"is_preprint":false},{"pmid":"31445393","id":"PMC_31445393","title":"Generation and characterization of a human induced pluripotent stem cell line UOWi005-A from dermal fibroblasts derived from a CCNFS621G familial amyotrophic lateral sclerosis patient using mRNA reprogramming.","date":"2019","source":"Stem cell research","url":"https://pubmed.ncbi.nlm.nih.gov/31445393","citation_count":8,"is_preprint":false},{"pmid":"36607545","id":"PMC_36607545","title":"MEKs/ERKs-mediated FBXO1/E2Fs interaction interference modulates G1/S cell cycle transition 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Patients.","date":"2024","source":"Genes","url":"https://pubmed.ncbi.nlm.nih.gov/39766833","citation_count":0,"is_preprint":false},{"pmid":"41069067","id":"PMC_41069067","title":"Expanding the genetic spectrum of corticobasal syndrome: novel CCNF p.M394L variant from a South Asian cohort.","date":"2025","source":"Neurocase","url":"https://pubmed.ncbi.nlm.nih.gov/41069067","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.09.14.676084","title":"Cell type specific analysis of ALS-associated proteins reveals immune-regulated targets","date":"2025-09-17","source":"bioRxiv","url":"https://doi.org/10.1101/2025.09.14.676084","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":12741,"output_tokens":3157,"usd":0.042789},"stage2":{"model":"claude-opus-4-6","input_tokens":6492,"output_tokens":3063,"usd":0.163552},"total_usd":0.206341,"stage1_batch_id":"msgbatch_011kfMfE81o6CTcyhVUqumuW","stage2_batch_id":"msgbatch_01DTputS6WNkH745x1nVSoBo","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1994,\n      \"finding\": \"CCNF encodes cyclin F, a novel member of the cyclin family related to A- and B-type cyclins by sequence, located on chromosome 16p13.3.\",\n      \"method\": \"cDNA sequencing, Northern blot analysis, exon-intron boundary determination\",\n      \"journal\": \"Genomics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — original characterization of gene/transcript; sequence homology to cyclins established but function unknown at time of publication\",\n      \"pmids\": [\"7896286\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Cyclin F (CCNF) functions as the substrate-binding subunit of an SCF (SKP1-CUL1-F-box) E3 ubiquitin-protein ligase complex (SCF^CyclinF); ALS/FTD-associated missense mutations cause abnormal ubiquitination and accumulation of ubiquitinated proteins including TDP-43 and an SCF^CyclinF substrate in neuronal cells.\",\n      \"method\": \"Whole-exome sequencing for mutation identification; transfection of mutant CCNF in neuronal cells with ubiquitination assays and western blot for substrate accumulation\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — highly cited foundational study (178 citations), multiple orthogonal methods, functional assays in neuronal cells\",\n      \"pmids\": [\"27080313\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"The ALS/FTD-linked cyclin F p.S621G mutation causes elevated Lys48-linked ubiquitylation of proteins in neuronal cells, and proteomic analysis identified autophagy pathway proteins (p62/SQSTM1, heat shock proteins, chaperonin components) as targets; mutant cyclin F impairs autophagosomal-lysosome fusion, and cyclin F physically interacts with p62.\",\n      \"method\": \"Transfection of mutant CCNF (S621G) in Neuro-2A and SH-SY5Y cells; K48-linkage-specific ubiquitin immunoprecipitation followed by mass spectrometry proteomics; autophagy marker analysis (p62, LC3, Lamp2) by immunoblot and immunofluorescence\",\n      \"journal\": \"Cellular and molecular life sciences : CMLS\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal methods (MS proteomics, specific K48-Ub IP, autophagy flux markers, Co-IP of cyclin F with p62)\",\n      \"pmids\": [\"28852778\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Cyclin F (CCNF) interacts with HIV-1 Vif protein; Vif is a substrate of the SCF^CyclinF E3 ligase complex, which mediates K48-linked ubiquitination and proteasomal degradation of Vif, thereby restoring APOBEC3G levels and restricting viral infectivity. A cyclin F-specific amino acid motif in the C-terminal region of Vif is required for this interaction.\",\n      \"method\": \"Co-immunoprecipitation, overexpression and knockdown studies, mutational analysis of Vif degron motif, ubiquitination assays, proteasome inhibitor experiments, APOBEC3G expression analysis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — reciprocal Co-IP, mutagenesis of degron, ubiquitination assay, multiple orthogonal methods in a single study\",\n      \"pmids\": [\"28184007\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Expression of ALS-linked mutant CCNF in zebrafish embryos causes increased caspase-3 activation and cell death in the spinal cord, motor neuron axonopathy (shortened primary motor axons, aberrant branching), and reduced photomotor response; proteomic analysis of in vitro models identified disruption of caspase-3-mediated cell death pathways.\",\n      \"method\": \"Transient overexpression of human mutant CCNF in zebrafish embryos; immunostaining for cleaved caspase-3; motor response (photomotor response) assay; label-free quantitative proteomics of in vitro models\",\n      \"journal\": \"Human molecular genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — in vivo zebrafish model with quantitative phenotypic readouts and proteomics; single lab\",\n      \"pmids\": [\"28444311\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Cyclin F physically binds to VCP (valosin-containing protein, also an ALS gene) via the N-terminal region of Cyclin F, and the two proteins colocalize in the nucleus. Cyclin F enhances VCP ATPase activity in vitro. ALS-associated CCNF mutations increase Cyclin F binding to VCP and further elevate VCP ATPase activity while causing cytoplasmic mislocalization of Cyclin F. Elevated VCP ATPase activity promotes cytoplasmic TDP-43 aggregation.\",\n      \"method\": \"Co-immunoprecipitation and colocalization experiments; domain-mapping pulldowns; in vitro ATPase activity assay; overexpression of mutant CCNF in transfected cells; TDP-43 aggregation assay\",\n      \"journal\": \"Human molecular genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — direct in vitro ATPase assay, domain-mapping Co-IP, and functional consequence (TDP-43 aggregation) measured with multiple methods in one study\",\n      \"pmids\": [\"31577344\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Label-free quantitative proteomics of HEK293 cells expressing multiple ALS-associated CCNF mutations (K97R, S195R, S509P, R574Q, S621G) bioinformatically predicted and immunoblot-validated activation of neuronal apoptosis pathways; iPSC-derived cells from S621G patients showed the same pathway activation.\",\n      \"method\": \"Label-free quantitative proteomics of transfected HEK293 cells; pathway bioinformatics; immunoblot validation; iPSC-derived patient cell proteomics\",\n      \"journal\": \"Frontiers in molecular neuroscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 — proteomics with immunoblot validation across multiple mutations and patient-derived iPSCs, single lab\",\n      \"pmids\": [\"33986643\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"CCNF protein is targeted for degradation by the E3 ligases FBXL8 and FZR1 (demonstrated by Co-IP pulldown); double knockdown of FBXL8 and FZR1 causes CCNF accumulation. CCNF itself pulls down RRM2 (ribonucleotide reductase subunit 2) and CCNF overexpression reduces RRM2 levels, indicating RRM2 is a substrate of SCF^CyclinF.\",\n      \"method\": \"Co-immunoprecipitation (FBXL8 and FZR1 pulldown of CCNF); double knockdown experiments; CCNF overexpression with RRM2 protein level measurement\",\n      \"journal\": \"Cancers\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — Co-IP and overexpression/knockdown; single lab, moderate orthogonality\",\n      \"pmids\": [\"34201347\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"FBXO1 (Cyclin F) directly binds E2F1, E2F2, and E2F3a transcription factors through Arg/Ile and Arg/Val degron motifs in their dimerization domains, mediating K48-linked ubiquitination and proteasomal degradation of E2Fs. MEK/ERK-dependent phosphorylation of threonine residues near these degron motifs regulates FBXO1-E2F interaction and E2F protein stability. Knockdown of FBXO1 elevated E2F levels and delayed G1/S cell cycle transition, inhibiting cancer cell proliferation.\",\n      \"method\": \"Co-immunoprecipitation; domain/motif mutation analysis (RI/AA, RV/AA); ubiquitination assays; cycloheximide chase for half-life; specific kinase inhibitors; FBXO1 knockdown with cell cycle analysis\",\n      \"journal\": \"Archives of pharmacal research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — reciprocal Co-IP, mutagenesis of degron motifs, ubiquitination assay, half-life measurement, and cell cycle functional readout in one study\",\n      \"pmids\": [\"36607545\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"The ALS-associated CCNF S621G variant causes ubiquitin-proteasome system (UPS) dysfunction in iPSC-derived motor neurons, with increased ubiquitinated protein abundance and altered ubiquitination of key UPS components. Overexpression of CCNF in NSC-34 cells alters free ubiquitin levels; double mutations that reduce CCNF's ability to form an active E3 ligase complex improved UPS function and increased free monomeric ubiquitin, establishing that the E3 ligase activity of CCNF is central to its role in ubiquitin homeostasis.\",\n      \"method\": \"iPSC-derived motor neurons from CCNF S621G patients; ubiquitin abundance and UPS component ubiquitination analysis; overexpression in NSC-34 cells with free ubiquitin measurement; active-site double-mutant analysis with UPS functional assay\",\n      \"journal\": \"Human molecular genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — patient-derived iPSC neurons, active-site mutagenesis with functional rescue, multiple orthogonal methods in one study\",\n      \"pmids\": [\"37220877\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"CRISPR/Cas9-mediated loss of ccnf in zebrafish causes abnormal motor neuron development and axonal outgrowth defects, and ccnf-deficient zebrafish show selective sensitization to endoplasmic reticulum stress but not oxidative stress, establishing a direct role for CCNF in motor neuron axonal maintenance in vivo.\",\n      \"method\": \"CRISPR/Cas9 genome editing in zebrafish to generate ccnf knockout; motor neuron morphology and axonal outgrowth analysis; pharmacological stress (ER stress, oxidative stress) with motor response readout\",\n      \"journal\": \"Cells\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — clean genetic knockout in vivo with defined morphological and stress-response phenotypes; single lab\",\n      \"pmids\": [\"38474336\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"Cyclin F (CCNF/FBXO1) is the substrate-recognition subunit of the SCF^CyclinF E3 ubiquitin ligase complex that mediates K48-linked ubiquitination and proteasomal degradation of substrates including E2F transcription factors, HIV-1 Vif, and RRM2; it physically interacts with and stimulates the ATPase activity of VCP; ALS/FTD-associated CCNF mutations dysregulate these activities, causing elevated K48-ubiquitylation, impaired autophagosome-lysosome fusion, cytoplasmic VCP hyperactivation driving TDP-43 aggregation, and disruption of free ubiquitin homeostasis in motor neurons.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"Cyclin F (CCNF/FBXO1) is the substrate-recognition subunit of the SCF^CyclinF E3 ubiquitin ligase complex, which mediates Lys48-linked ubiquitination and proteasomal degradation of key cell-cycle and homeostatic substrates including E2F1/E2F2/E2F3a transcription factors, RRM2, and HIV-1 Vif [PMID:36607545, PMID:34201347, PMID:28184007]. Through recognition of specific degron motifs in substrates — regulated by MEK/ERK-dependent phosphorylation in the case of E2Fs — SCF^CyclinF controls G1/S cell-cycle progression, ubiquitin-proteasome system homeostasis, and autophagosome–lysosome fusion, and it modulates VCP ATPase activity via direct physical interaction [PMID:36607545, PMID:37220877, PMID:28852778, PMID:31577344]. Missense mutations in CCNF cause familial ALS/FTD by elevating aberrant Lys48-ubiquitylation, disrupting free ubiquitin pools in motor neurons, hyperactivating cytoplasmic VCP to promote TDP-43 aggregation, and impairing autophagic clearance [PMID:27080313, PMID:37220877, PMID:31577344, PMID:28852778]. Loss of CCNF in zebrafish produces motor neuron axonal outgrowth defects and selective vulnerability to endoplasmic reticulum stress, confirming a direct requirement for CCNF in motor neuron maintenance [PMID:38474336].\",\n  \"teleology\": [\n    {\n      \"year\": 1994,\n      \"claim\": \"Identification of CCNF as a novel cyclin-family member related to A- and B-type cyclins established the gene as a cell-cycle-associated factor, but its specific biochemical function was unknown.\",\n      \"evidence\": \"cDNA sequencing and Northern blot analysis of human transcript\",\n      \"pmids\": [\"7896286\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No enzymatic or functional activity defined\", \"No interacting partners identified\", \"Cyclin-box homology alone did not establish CDK dependency\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Discovery that cyclin F is the substrate-recognition (F-box) subunit of an SCF E3 ubiquitin ligase, and that ALS/FTD-associated mutations cause accumulation of ubiquitinated proteins including TDP-43 in neuronal cells, established CCNF as a disease-linked ubiquitin ligase component.\",\n      \"evidence\": \"Whole-exome sequencing of ALS/FTD families; mutant CCNF expression in neuronal cells with ubiquitination assays and immunoblot\",\n      \"pmids\": [\"27080313\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Endogenous substrates beyond TDP-43 not identified\", \"Mechanism by which mutations alter ligase activity undefined\", \"No in vivo validation of motor neuron phenotype\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Characterization of the S621G mutant revealed that disease-linked cyclin F elevates Lys48-ubiquitylation of autophagy-related proteins, physically interacts with p62/SQSTM1, and impairs autophagosome–lysosome fusion, connecting SCF^CyclinF dysfunction to proteostasis failure via autophagy.\",\n      \"evidence\": \"K48-linkage-specific ubiquitin IP with mass spectrometry; autophagy flux markers (LC3, Lamp2) in Neuro-2A and SH-SY5Y cells; Co-IP of cyclin F and p62\",\n      \"pmids\": [\"28852778\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether p62 is a direct ubiquitination substrate or an adaptor not resolved\", \"Autophagy defect not confirmed in patient-derived neurons at this stage\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Demonstration that SCF^CyclinF targets HIV-1 Vif for K48-linked ubiquitination and proteasomal degradation — via a cyclin F-specific degron motif in Vif — expanded the substrate repertoire to viral proteins and revealed host antiviral restriction function.\",\n      \"evidence\": \"Reciprocal Co-IP, degron mutagenesis, ubiquitination assays, proteasome inhibitor rescue, APOBEC3G restoration upon Vif degradation\",\n      \"pmids\": [\"28184007\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological relevance in HIV-infected patients not tested\", \"Degron specificity for other viral substrates unknown\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Expression of ALS-linked CCNF mutants in zebrafish embryos caused motor neuron axonopathy, spinal cord apoptosis, and impaired motor responses, providing the first in vivo evidence that mutant CCNF is neurotoxic.\",\n      \"evidence\": \"Transient overexpression of mutant human CCNF in zebrafish; caspase-3 immunostaining; photomotor response assay\",\n      \"pmids\": [\"28444311\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Overexpression model does not distinguish gain-of-function from dominant-negative effects\", \"Endogenous loss-of-function phenotype not assessed\", \"No mammalian in vivo model\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Discovery that cyclin F directly binds and stimulates VCP ATPase activity, and that ALS mutations enhance this interaction causing cytoplasmic mislocalization and VCP hyperactivation that drives TDP-43 aggregation, provided a concrete pathogenic mechanism linking two ALS genes.\",\n      \"evidence\": \"Co-IP and colocalization; domain-mapping pulldowns; in vitro VCP ATPase activity assay; TDP-43 aggregation assay in transfected cells\",\n      \"pmids\": [\"31577344\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of cyclin F–VCP interface unresolved\", \"Whether VCP is itself a ubiquitination substrate of SCF^CyclinF unknown\", \"TDP-43 aggregation not validated in patient-derived neurons\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Identification of RRM2 as an SCF^CyclinF substrate and of FBXL8/FZR1 as E3 ligases that target CCNF itself for degradation established bidirectional regulation of cyclin F abundance and a new substrate relevant to DNA replication.\",\n      \"evidence\": \"Co-IP of FBXL8/FZR1 with CCNF; double knockdown causing CCNF accumulation; CCNF overexpression reducing RRM2 levels\",\n      \"pmids\": [\"34201347\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct ubiquitination of RRM2 by SCF^CyclinF not demonstrated with purified components\", \"Physiological contexts (cell cycle phase) not resolved\", \"Single-lab finding awaiting independent replication\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Identification of E2F1/E2F2/E2F3a as direct SCF^CyclinF substrates, degraded via specific Arg/Ile and Arg/Val degron motifs regulated by MEK/ERK phosphorylation, established cyclin F as a master regulator of G1/S progression through E2F turnover.\",\n      \"evidence\": \"Reciprocal Co-IP; degron motif mutagenesis (RI/AA, RV/AA); ubiquitination and cycloheximide chase assays; MEK/ERK inhibitor treatment; FBXO1 knockdown with cell-cycle analysis\",\n      \"pmids\": [\"36607545\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo cell-cycle phenotype of CCNF loss in mammals not examined\", \"Whether all three E2Fs are targeted simultaneously or in different contexts is unclear\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Patient iPSC-derived motor neurons carrying CCNF S621G showed increased ubiquitinated protein loads and altered UPS component ubiquitination; active-site double mutations restoring ligase function rescued free ubiquitin levels, proving that SCF^CyclinF E3 ligase activity directly controls ubiquitin homeostasis in motor neurons.\",\n      \"evidence\": \"iPSC-derived motor neurons from CCNF S621G patients; ubiquitin profiling; active-site double-mutant rescue in NSC-34 cells\",\n      \"pmids\": [\"37220877\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Specific ubiquitin-modified substrates driving motor neuron vulnerability not identified\", \"Rescue not performed in patient neurons\", \"Relationship between UPS dysfunction and autophagy defects not integrated\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"CRISPR knockout of ccnf in zebrafish confirmed a direct requirement for CCNF in motor neuron axonal outgrowth and revealed selective vulnerability to ER stress but not oxidative stress, distinguishing the stress-response pathways downstream of CCNF loss.\",\n      \"evidence\": \"CRISPR/Cas9 ccnf knockout zebrafish; motor neuron morphology; pharmacological ER and oxidative stress challenges\",\n      \"pmids\": [\"38474336\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism linking CCNF loss to ER stress sensitivity unknown\", \"Mammalian knockout model still lacking\", \"Transcriptomic/proteomic characterization of knockout neurons not performed\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"Key unresolved questions include the structural basis of SCF^CyclinF substrate recognition and VCP interaction, the full catalogue of physiological substrates in motor neurons, the relative contributions of UPS dysfunction versus autophagy impairment versus VCP hyperactivation to ALS/FTD pathogenesis, and validation of these mechanisms in mammalian in vivo models.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No crystal or cryo-EM structure of SCF^CyclinF with any substrate\", \"No mammalian in vivo conditional knockout model\", \"Integration of UPS, autophagy, and VCP pathways into unified pathogenic cascade not achieved\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [1, 2, 3, 8, 9]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [5, 8]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [5]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [1, 2, 3, 8, 9]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [8]},\n      {\"term_id\": \"R-HSA-9612973\", \"supporting_discovery_ids\": [2]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [4, 6]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [1, 5, 9]}\n    ],\n    \"complexes\": [\n      \"SCF^CyclinF (SKP1-CUL1-CyclinF)\"\n    ],\n    \"partners\": [\n      \"SKP1\",\n      \"CUL1\",\n      \"VCP\",\n      \"SQSTM1\",\n      \"E2F1\",\n      \"E2F2\",\n      \"E2F3\",\n      \"RRM2\"\n    ],\n    \"other_free_text\": []\n  }\n}\n```"}