FBXL4 is an F-box and leucine-rich-repeat protein that serves as a substrate-recognition component of an SCF ubiquitin ligase at the mitochondrial outer membrane. By promoting turnover of the mitophagy receptors BNIP3 and BNIP3L/NIX, it restrains excessive mitochondrial removal and helps preserve mitochondrial content. PPTC7 assists receptor recruitment, while FBXL4 associates with the SKP1–CUL1 core. Biallelic pathogenic variants cause mitochondrial DNA depletion syndrome 13 (MTDPS13), an encephalomyopathic disorder associated with early-onset developmental impairment, hypotonia, lactic acidosis and mitochondrial oxidative-phosphorylation defects. A contextual role in JMJD2A/KDM4A turnover has also been reported in human cell studies.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0000151 ubiquitin ligase complex | TAS PMID:10531035 Identification of a family of human F-box proteins. | TOO GENERAL | Summary: FBXL4 belongs to the SCF ubiquitin ligase complex. Reason: The 1999 paper identifies Fbl4 among F-box family members by database searching; its representative interaction assays should not be confused with assays of the similarly named Fbx4. The family-level ubiquitin-ligase-complex inference is positive. Independent target co-immunoprecipitation, F-box perturbation and ubiquitination experiments in PMID36896912 and PMID37161784 establish the more informative SCF complex, already retained at GO:0019005. Official ontology ancestry confirms GO:0019005 is a descendant of GO:0000151. The generic complex term adds no separate complex role; this does not reject its TAS support. Supporting Evidence: PMID:10531035 In fact, F-box proteins are one of the four subunits of ubiquitin protein ligases called SCFs. PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. |
| GO:0000422 autophagy of mitochondrion | IEA GO_REF:0000117 | MODIFY | Summary: FBXL4 restricts mitophagy through turnover of the BNIP3 and NIX receptors. Reason: The actual ARBA rule predicts autophagy of mitochondrion, but the target experiments distinguish the regulator from the machinery that delivers and degrades mitochondria. In PMID:36896912, loss of FBXL4 increases BNIP3/NIX-dependent mitophagy and wild-type rescue restores restraint; SCF-dependent receptor ubiquitination and degradation supply the mechanism. PMID:37161784 independently supports receptor turnover and basal mitophagy suppression. Negative regulation of mitophagy is the appropriate role correction and is already supported by direct row 27. This is a lateral regulatory correction, not a strict is_a/part_of refinement or a claim that FBXL4 performs mitochondrial degradation. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: ROLE CONFLATION Sources checked: ARBA:ARBA00026840 · ARBA00026840 SOURCE WEAK OR INFERRED Actual current rule has eight alternative condition sets and predicts GO:0000422. Its full target FunFam condition match and historical model state were not independently replayed; target perturbation and receptor-turnover evidence determine the role correction. Proposed replacements: negative regulation of mitophagy Supporting Evidence: PMID:36896912 we found that knockout (KO) of FBXL4, a mitochondrial disease gene, hyperactivates mitophagy at basal conditions. PMID:37161784 Thus, we reveal a molecular mechanism whereby FBXL4 actively suppresses mitophagy by preventing NIX and BNIP3 accumulation. |
| GO:0000422 autophagy of mitochondrion | IMP PMID:32525278 FBXL4 deficiency increases mitochondrial removal by autophag... | MODIFY | Summary: FBXL4 limits excessive mitochondrial removal by autophagy. Reason: The actual patient-cell and mouse study measures increased lysosome-dependent mitochondrial turnover after FBXL4 deficiency, rather than showing that FBXL4 executes mitochondrial degradation. Its earlier LC3-related interpretation is bounded to those assays. Subsequent target studies resolve SCF-mediated turnover of BNIP3/NIX as a mechanism restraining mitophagy. Negative regulation of mitophagy (GO:1901525) states this inhibitory role more precisely while preserving the original acts_upstream_of_or_within source qualifier. Proposed replacements: negative regulation of mitophagy Supporting Evidence: PMID:32525278 the results we present here show that FBXL4 prevents mitochondrial removal via autophagy and that loss of FBXL4 leads to decreased mitochondrial content and mitochondrial disease PMID:36896912 we found that knockout (KO) of FBXL4, a mitochondrial disease gene, hyperactivates mitophagy at basal conditions. |
| GO:0005515 protein binding | IPI PMID:32814053 Interactome Mapping Provides a Network of Neurodegenerative ... | MODIFY | Summary: Human FBXL4 and TPP1 form a positive yeast two-hybrid pair. Reason: Human FBXL4–TPP1 binding is supported by the original Table S2 (NN1.0 row3486; NN1.0ext row7570) and exact positive IntAct records EBI-25838630, EBI-25995383 and EBI-26149530 joining Q9UKA2 to O14773. These report array, pooling and validated yeast two-hybrid evidence; the study used reporter non-autoactivation checks and pairwise retesting. GO:0002020 means binding to a protease or peptidase, and O14773 is tripeptidyl-peptidase 1. The refinement describes this measured pair, not protease regulation, SCF substrate recruitment or an established physiological interaction. No target-specific DULIP validation was recovered. Clone CCSB_992 and empty IntAct feature fields do not resolve insert boundaries or mature-versus-precursor state. The inherited abstract quotation identifies the study but does not itself support this pair; the original workbook and curated interaction records supply that support. Proposed replacements: protease binding Supporting Evidence: PMID:32814053 Here, we report on an interactome map that focuses on neurodegenerative disease (ND) |
| GO:0005515 protein binding | IPI PMID:32814053 Interactome Mapping Provides a Network of Neurodegenerative ... | MODIFY | Summary: Human FBXL4 and the source DNM2 isoform form a positive yeast two-hybrid pair. Reason: Human FBXL4–DNM2 binding is supported by the original Table S2 (NN1.0 row3485; NN1.0ext row7569) and exact positive IntAct records EBI-25845005, EBI-26001463 and EBI-26155610 joining Q9UKA2 to P50570-2. These report array, pooling and validated yeast two-hybrid evidence with pairwise retesting. GO:0051020 means binding to a GTPase; dynamin-2 is a GTPase (EC3.6.5.5). The definition does not require effector activity, GTP-bound-state specificity or regulation. Those activities, physiological membrane remodeling and target-specific mammalian validation are not established here. The gene-level workbook and curator-resolved P50570-2 identity remain distinct. Clone IOH28879 and empty feature fields leave insert boundaries unresolved. The inherited abstract quotation identifies the study, while the exact workbook and IntAct pairs support this binding refinement. Proposed replacements: GTPase binding Supporting Evidence: PMID:32814053 Here, we report on an interactome map that focuses on neurodegenerative disease (ND) |
| GO:0005634 nucleus | IEA GO_REF:0000044 | TOO GENERAL | Summary: A broad nuclear location is covered by the retained HPA nuclear-speck observation. Reason: Current UniProt explicitly retains SL-0191 nucleus but supplies no location-specific evidence tag for that field. Independently inspected HPA029140 immunofluorescence reports supported nuclear-speck staining in A-431, U-251MG and U2OS, retained as contextual row 22. Nuclear speck is a strict is_a/part_of descendant of nucleus. The broad term adds no separate resolved nuclear compartment here. PMID:23993194 reported no detectable nuclear FBXL4 in its assayed cell/fractionation conditions; that discrepancy and the single-antibody specificity limit remain explicit and do not justify a universal nuclear exclusion. The inherited mitochondrial quotes document the other location, not evidence against every nuclear pool. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: UniProtKB-SubCell:SL-0191 · Nucleus SUPPORTS TRANSFER The literal location remains in the official record. Independent HPA nuclear-speck staining supports the narrower retained compartment, without establishing a nuclear molecular function. Supporting Evidence: PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. PMID:37161784 localises to the mitochondrial outer membrane in unstressed cells and mediates the constitutive ubiquitylation and degradation of the mitophagy receptors NIX and BNIP3 to suppress basal levels of mitophagy |
| GO:0005737 cytoplasm | IEA GO_REF:0000044 | TOO GENERAL | Summary: Mitochondrial residence is compatible with the broad cytoplasm term. Reason: GO:0005737 includes organelles and is not synonymous with cytosol. The positively supported mitochondrial outer-membrane location in rows 8 and 9 is a strict is_a/part_of descendant of cytoplasm and provides the resolved location for this source. Current UniProt retains SL-0086 cytoplasm without a field-specific evidence tag. The broad term is redundant with that retained organellar location; it is not biologically false. This action does not convert the separate cytosol records into mitochondrial annotations. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: UniProtKB-SubCell:SL-0086 · Cytoplasm SUPPORTS TRANSFER Official location field was read. Its broad meaning includes the experimentally supported mitochondrial compartment. Supporting Evidence: PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. |
| GO:0005739 mitochondrion | HTP PMID:34800366 Quantitative high-confidence human mitochondrial proteome an... | ACCEPT | Summary: FBXL4 is directly identified in the high-confidence human mitochondrial proteome. Reason: The original MitoCoP workbooks identify the sole Q9UKA2/FBXL4 protein group: Table S1B row340, subtractive Table S2B row6990 class1, spatial S2E row7954 cluster1, and TOMM40-importomics S3A row6030 class1. The multi-cell-line S2G row9225 records mitochondrial clustering in HEK, HeLa and Huh7, with the U2OS mitochondrial-cluster cell blank. S3C row738 is assigned the ambiguous membrane-association cluster, so this HTP source is not recast as a precise outer-membrane assay. Retain the informative organelle location under the project organelle guidance; separate target experiments support the outer-membrane annotation. Supporting Evidence: PMID:34800366 We classified >8,000 proteins in mitochondrial preparations of human cells and defined a mitochondrial high-confidence proteome of >1,100 proteins (MitoCoP). PMID:23993193 We show that FBXL4 is an F-box protein that colocalizes with mitochondria |
| GO:0005741 mitochondrial outer membrane | IDA PMID:36896912 A mitochondrial SCF-FBXL4 ubiquitin E3 ligase complex degrad... | ACCEPT | Summary: FBXL4 is an integral mitochondrial outer-membrane component of the SCF ligase. Reason: The actual PMID36896912 Results and submitochondrial-localization Methods combine mitochondrial protease accessibility, alkaline extraction, microscopy and N-terminal targeting constructs with SCF association. Tagged FBXL4 is accessible in intact mitochondria and remains in the membrane pellet under the reported conditions. Independent PMID37161784 microscopy distinguishes TOM20-associated outer-membrane localization from TIM50 after mitochondrial swelling. These positive experiments support the active outer-membrane pool, without treating the conflicting 2013 protection/fractionation result as never performed or proving exclusive localization in every condition. Engineered targeting constructs are not natural products. Supporting Evidence: PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. PMID:37161784 localises to the mitochondrial outer membrane in unstressed cells and mediates the constitutive ubiquitylation and degradation of the mitophagy receptors NIX and BNIP3 to suppress basal levels of mitophagy |
| GO:0005741 mitochondrial outer membrane | IEA GO_REF:0000120 | ACCEPT | Summary: FBXL4 localizes to the mitochondrial outer membrane. Reason: The combined electronic source joins ARBA00088384 and UniProt SL-0172. The current UniProt outer-membrane field explicitly cites PMID:36896912. In that human HeLa study, tagged FBXL4 is accessible to protease in intact mitochondria and remains in the membrane pellet under alkaline extraction, with compartment controls. PMID:37161784 independently reports tagged FBXL4 localization relative to TOM20 and TIM50 and requires mitochondrial targeting for receptor turnover. These target results support the mapped location. They do not establish exclusive localization in every cell or condition, and engineered targeting constructs are not natural products. Propagation Review Root cause: NO FAILURE CORE Sources checked: ARBA:ARBA00088384 · ARBA00088384 SUPPORTS TRANSFER Actual rule predicts GO:0005741 through alternative FunFam/taxon conditions. All target rule matches were not replayed; independent target localization supports the consequence. UniProtKB-SubCell:SL-0172 · Mitochondrion outer membrane SUPPORTS TRANSFER Official target field has ECO:0000269 evidence from PMID:36896912; actual original localization results were read. Supporting Evidence: PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. |
| GO:0005758 mitochondrial intermembrane space | IDA PMID:23993194 Mutations in FBXL4, encoding a mitochondrial protein, cause ... | UNDECIDED | Summary: The original intermembrane-space assignment conflicts with later target topology experiments. Reason: The full 2013 original was recovered and read: tagged human FBXL4 was protected from trypsin in intact mitochondrial preparations and became accessible after digitonin treatment; separate mouse-liver fractionation supported a soluble mitochondrial pool. These are actual experiments, not an abstract-only inference. Later PMID36896912 reports protease accessibility and membrane association compatible with an outer-membrane pool. Preparation, expression and protease conditions differ; the 2013 Results and Methods also print different trypsin-concentration units. The evidence assessed here does not resolve that discrepancy or demonstrate which side each endogenous segment occupies. Preserve the positive historical observation and curator source while leaving precise intermembrane-space localization undecided. Supporting Evidence: PMID:23993194 We also demonstrate that FBXL4 is targeted to mitochondria and localizes in the intermembrane space, where it participates in an approximately 400 kDa protein complex. PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. |
| GO:0005829 cytosol | TAS Reactome:R-HSA-8952618 | MARK AS OVER ANNOTATED | Summary: The modeled cytosolic compartment is overextended to FBXL4 through a generic ligase set. Reason: This event reaches FBXL4 through F-box DefinedSet 976085. The target entity 8850899 itself explicitly assigns Q9UKA2 residues 1–621 to cytosol; the compartment is not merely a label on a parent complex. This modeled set membership does not establish a FBXL4-specific cytosolic pool. Primary experiments support an integral mitochondrial outer-membrane pool (PMID:36896912), and the historical fractionation study did not detect a postmitochondrial cytosolic pool under its tested conditions (PMID:23993194). Cytosolic exposure of a membrane protein does not establish localization in the organelle-excluding cytosol compartment. The source therefore overextends compartment evidence. This judgment does not exclude a conditional cytosolic pool or assert that outer membrane is a descendant of cytosol. The separate outer-membrane annotation remains supported. Propagation Review Root cause: SOURCE WEAK OR INFERRED Failure modes: SOURCE EVIDENCE WEAK COMPARTMENT OR COMPLEX MISMATCH Sources checked: Reactome:R-HSA-8952618 · AcM-UBE2M transfers NEDD8 to CRL1 E3 ubiquitin ligase complex SOURCE WEAK OR INFERRED Actual route: 8952618 -> output -> 8952593 -> hasComponent -> 8952588 -> hasComponent -> 976085 -> hasMember -> 8850899. Target membership and its explicit cytosol field are verified; the set-based compartment model lacks resolved target-specific localization support. SCF node976110, when present, is a Complex, not a F-box set. Supporting Evidence: PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. PMID:37161784 localises to the mitochondrial outer membrane in unstressed cells and mediates the constitutive ubiquitylation and degradation of the mitophagy receptors NIX and BNIP3 to suppress basal levels of mitophagy |
| GO:0005829 cytosol | TAS Reactome:R-HSA-8952620 | MARK AS OVER ANNOTATED | Summary: The modeled cytosolic compartment is overextended to FBXL4 through a generic ligase set. Reason: This event reaches FBXL4 through F-box DefinedSet 976085. The target entity 8850899 itself explicitly assigns Q9UKA2 residues 1–621 to cytosol; the compartment is not merely a label on a parent complex. This modeled set membership does not establish a FBXL4-specific cytosolic pool. Primary experiments support an integral mitochondrial outer-membrane pool (PMID:36896912), and the historical fractionation study did not detect a postmitochondrial cytosolic pool under its tested conditions (PMID:23993194). Cytosolic exposure of a membrane protein does not establish localization in the organelle-excluding cytosol compartment. The source therefore overextends compartment evidence. This judgment does not exclude a conditional cytosolic pool or assert that outer membrane is a descendant of cytosol. The separate outer-membrane annotation remains supported. Propagation Review Root cause: SOURCE WEAK OR INFERRED Failure modes: SOURCE EVIDENCE WEAK COMPARTMENT OR COMPLEX MISMATCH Sources checked: Reactome:R-HSA-8952620 · NEDD8:AcM-UBE2M binds CRL1 E3 ubiquitin ligase complex SOURCE WEAK OR INFERRED Actual route: 8952620 -> input -> 8955230 -> hasComponent -> 976110 -> hasComponent -> 976085 -> hasMember -> 8850899. Target membership and its explicit cytosol field are verified; the set-based compartment model lacks resolved target-specific localization support. SCF node976110, when present, is a Complex, not a F-box set. Supporting Evidence: PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. PMID:37161784 localises to the mitochondrial outer membrane in unstressed cells and mediates the constitutive ubiquitylation and degradation of the mitophagy receptors NIX and BNIP3 to suppress basal levels of mitophagy |
| GO:0005829 cytosol | TAS Reactome:R-HSA-8955241 | MARK AS OVER ANNOTATED | Summary: The modeled cytosolic compartment is overextended to FBXL4 through a generic ligase set. Reason: This event reaches FBXL4 through F-box DefinedSet 976085. The target entity 8850899 itself explicitly assigns Q9UKA2 residues 1–621 to cytosol; the compartment is not merely a label on a parent complex. This modeled set membership does not establish a FBXL4-specific cytosolic pool. Primary experiments support an integral mitochondrial outer-membrane pool (PMID:36896912), and the historical fractionation study did not detect a postmitochondrial cytosolic pool under its tested conditions (PMID:23993194). Cytosolic exposure of a membrane protein does not establish localization in the organelle-excluding cytosol compartment. The source therefore overextends compartment evidence. This judgment does not exclude a conditional cytosolic pool or assert that outer membrane is a descendant of cytosol. The separate outer-membrane annotation remains supported. Propagation Review Root cause: SOURCE WEAK OR INFERRED Failure modes: SOURCE EVIDENCE WEAK COMPARTMENT OR COMPLEX MISMATCH Sources checked: Reactome:R-HSA-8955241 · CAND1 binds cytosolic CRL E3 ubiquitin ligases SOURCE WEAK OR INFERRED Actual route: 8955241 -> input -> 8955222 -> hasMember -> 976110 -> hasComponent -> 976085 -> hasMember -> 8850899. Target membership and its explicit cytosol field are verified; the set-based compartment model lacks resolved target-specific localization support. SCF node976110, when present, is a Complex, not a F-box set. Supporting Evidence: PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. PMID:37161784 localises to the mitochondrial outer membrane in unstressed cells and mediates the constitutive ubiquitylation and degradation of the mitophagy receptors NIX and BNIP3 to suppress basal levels of mitophagy |
| GO:0005829 cytosol | TAS Reactome:R-HSA-8955289 | MARK AS OVER ANNOTATED | Summary: The modeled cytosolic compartment is overextended to FBXL4 through a generic ligase set. Reason: This event reaches FBXL4 through F-box DefinedSet 976085. The target entity 8850899 itself explicitly assigns Q9UKA2 residues 1–621 to cytosol; the compartment is not merely a label on a parent complex. This modeled set membership does not establish a FBXL4-specific cytosolic pool. Primary experiments support an integral mitochondrial outer-membrane pool (PMID:36896912), and the historical fractionation study did not detect a postmitochondrial cytosolic pool under its tested conditions (PMID:23993194). Cytosolic exposure of a membrane protein does not establish localization in the organelle-excluding cytosol compartment. The source therefore overextends compartment evidence. This judgment does not exclude a conditional cytosolic pool or assert that outer membrane is a descendant of cytosol. The separate outer-membrane annotation remains supported. Propagation Review Root cause: SOURCE WEAK OR INFERRED Failure modes: SOURCE EVIDENCE WEAK COMPARTMENT OR COMPLEX MISMATCH Sources checked: Reactome:R-HSA-8955289 · COMMDs displace CAND1 from cytosolic CRL E3 ubiquitin ligase complexes SOURCE WEAK OR INFERRED Actual route: 8955289 -> input -> 8955224 -> hasComponent -> 8955222 -> hasMember -> 976110 -> hasComponent -> 976085 -> hasMember -> 8850899. Target membership and its explicit cytosol field are verified; the set-based compartment model lacks resolved target-specific localization support. SCF node976110, when present, is a Complex, not a F-box set. Supporting Evidence: PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. PMID:37161784 localises to the mitochondrial outer membrane in unstressed cells and mediates the constitutive ubiquitylation and degradation of the mitophagy receptors NIX and BNIP3 to suppress basal levels of mitophagy |
| GO:0005829 cytosol | TAS Reactome:R-HSA-8956040 | MARK AS OVER ANNOTATED | Summary: The modeled cytosolic compartment is overextended to FBXL4 through a generic ligase set. Reason: This event reaches FBXL4 through F-box DefinedSet 976085. The target entity 8850899 itself explicitly assigns Q9UKA2 residues 1–621 to cytosol; the compartment is not merely a label on a parent complex. This modeled set membership does not establish a FBXL4-specific cytosolic pool. Primary experiments support an integral mitochondrial outer-membrane pool (PMID:36896912), and the historical fractionation study did not detect a postmitochondrial cytosolic pool under its tested conditions (PMID:23993194). Cytosolic exposure of a membrane protein does not establish localization in the organelle-excluding cytosol compartment. The source therefore overextends compartment evidence. This judgment does not exclude a conditional cytosolic pool or assert that outer membrane is a descendant of cytosol. The separate outer-membrane annotation remains supported. Propagation Review Root cause: SOURCE WEAK OR INFERRED Failure modes: SOURCE EVIDENCE WEAK COMPARTMENT OR COMPLEX MISMATCH Sources checked: Reactome:R-HSA-8956040 · COP9 signalosome deneddylates cytosolic CRL E3 ubiquitin ligase complexes SOURCE WEAK OR INFERRED Actual route: 8956040 -> input -> 8956021 -> hasMember -> 8952593 -> hasComponent -> 8952588 -> hasComponent -> 976085 -> hasMember -> 8850899. Target membership and its explicit cytosol field are verified; the set-based compartment model lacks resolved target-specific localization support. SCF node976110, when present, is a Complex, not a F-box set. Supporting Evidence: PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. PMID:37161784 localises to the mitochondrial outer membrane in unstressed cells and mediates the constitutive ubiquitylation and degradation of the mitophagy receptors NIX and BNIP3 to suppress basal levels of mitophagy |
| GO:0005829 cytosol | TAS Reactome:R-HSA-8956200 | MARK AS OVER ANNOTATED | Summary: The modeled cytosolic compartment is overextended to FBXL4 through a generic ligase set. Reason: This event reaches FBXL4 through F-box DefinedSet 976085. The target entity 8850899 itself explicitly assigns Q9UKA2 residues 1–621 to cytosol; the compartment is not merely a label on a parent complex. This modeled set membership does not establish a FBXL4-specific cytosolic pool. Primary experiments support an integral mitochondrial outer-membrane pool (PMID:36896912), and the historical fractionation study did not detect a postmitochondrial cytosolic pool under its tested conditions (PMID:23993194). Cytosolic exposure of a membrane protein does not establish localization in the organelle-excluding cytosol compartment. The source therefore overextends compartment evidence. This judgment does not exclude a conditional cytosolic pool or assert that outer membrane is a descendant of cytosol. The separate outer-membrane annotation remains supported. Propagation Review Root cause: SOURCE WEAK OR INFERRED Failure modes: SOURCE EVIDENCE WEAK COMPARTMENT OR COMPLEX MISMATCH Sources checked: Reactome:R-HSA-8956200 · MyrG-DCUN1D3 binds CRL1 E3 ubiquitin ligase complex SOURCE WEAK OR INFERRED Actual route: 8956200 -> input -> 8955230 -> hasComponent -> 976110 -> hasComponent -> 976085 -> hasMember -> 8850899. Target membership and its explicit cytosol field are verified; the set-based compartment model lacks resolved target-specific localization support. SCF node976110, when present, is a Complex, not a F-box set. Supporting Evidence: PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. PMID:37161784 localises to the mitochondrial outer membrane in unstressed cells and mediates the constitutive ubiquitylation and degradation of the mitophagy receptors NIX and BNIP3 to suppress basal levels of mitophagy |
| GO:0005829 cytosol | TAS Reactome:R-HSA-983140 | MARK AS OVER ANNOTATED | Summary: The modeled cytosolic compartment is overextended to FBXL4 through a generic ligase set. Reason: This event reaches FBXL4 through generic E3 CandidateSet 976075. The target entity 8850899 itself explicitly assigns Q9UKA2 residues 1–621 to cytosol; the compartment is not merely a label on a parent complex. This modeled set membership does not establish a FBXL4-specific cytosolic pool. Primary experiments support an integral mitochondrial outer-membrane pool (PMID:36896912), and the historical fractionation study did not detect a postmitochondrial cytosolic pool under its tested conditions (PMID:23993194). Cytosolic exposure of a membrane protein does not establish localization in the organelle-excluding cytosol compartment. The source therefore overextends compartment evidence. This judgment does not exclude a conditional cytosolic pool or assert that outer membrane is a descendant of cytosol. The separate outer-membrane annotation remains supported. Propagation Review Root cause: SOURCE WEAK OR INFERRED Failure modes: SOURCE EVIDENCE WEAK COMPARTMENT OR COMPLEX MISMATCH Sources checked: Reactome:R-HSA-983140 · Transfer of Ub from E2 to substrate and release of E2 SOURCE WEAK OR INFERRED Actual route: 983140 -> input -> 983126 -> hasComponent -> 976075 -> hasMember -> 8850899. Target membership and its explicit cytosol field are verified; the set-based compartment model lacks resolved target-specific localization support. SCF node976110, when present, is a Complex, not a F-box set. Supporting Evidence: PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. PMID:37161784 localises to the mitochondrial outer membrane in unstressed cells and mediates the constitutive ubiquitylation and degradation of the mitophagy receptors NIX and BNIP3 to suppress basal levels of mitophagy |
| GO:0005829 cytosol | TAS Reactome:R-HSA-983147 | MARK AS OVER ANNOTATED | Summary: The modeled cytosolic compartment is overextended to FBXL4 through a generic ligase set. Reason: This event reaches FBXL4 through generic E3 CandidateSet 976075. The target entity 8850899 itself explicitly assigns Q9UKA2 residues 1–621 to cytosol; the compartment is not merely a label on a parent complex. This modeled set membership does not establish a FBXL4-specific cytosolic pool. Primary experiments support an integral mitochondrial outer-membrane pool (PMID:36896912), and the historical fractionation study did not detect a postmitochondrial cytosolic pool under its tested conditions (PMID:23993194). Cytosolic exposure of a membrane protein does not establish localization in the organelle-excluding cytosol compartment. The source therefore overextends compartment evidence. This judgment does not exclude a conditional cytosolic pool or assert that outer membrane is a descendant of cytosol. The separate outer-membrane annotation remains supported. Propagation Review Root cause: SOURCE WEAK OR INFERRED Failure modes: SOURCE EVIDENCE WEAK COMPARTMENT OR COMPLEX MISMATCH Sources checked: Reactome:R-HSA-983147 · Release of E3 from polyubiquitinated substrate SOURCE WEAK OR INFERRED Actual route: 983147 -> output -> 976075 -> hasMember -> 8850899. Target membership and its explicit cytosol field are verified; the set-based compartment model lacks resolved target-specific localization support. SCF node976110, when present, is a Complex, not a F-box set. Supporting Evidence: PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. PMID:37161784 localises to the mitochondrial outer membrane in unstressed cells and mediates the constitutive ubiquitylation and degradation of the mitophagy receptors NIX and BNIP3 to suppress basal levels of mitophagy |
| GO:0005829 cytosol | TAS Reactome:R-HSA-983156 | MARK AS OVER ANNOTATED | Summary: The modeled cytosolic compartment is overextended to FBXL4 through a generic ligase set. Reason: This event reaches FBXL4 through generic E3 CandidateSet 976075. The target entity 8850899 itself explicitly assigns Q9UKA2 residues 1–621 to cytosol; the compartment is not merely a label on a parent complex. This modeled set membership does not establish a FBXL4-specific cytosolic pool. Primary experiments support an integral mitochondrial outer-membrane pool (PMID:36896912), and the historical fractionation study did not detect a postmitochondrial cytosolic pool under its tested conditions (PMID:23993194). Cytosolic exposure of a membrane protein does not establish localization in the organelle-excluding cytosol compartment. The source therefore overextends compartment evidence. This judgment does not exclude a conditional cytosolic pool or assert that outer membrane is a descendant of cytosol. The separate outer-membrane annotation remains supported. Propagation Review Root cause: SOURCE WEAK OR INFERRED Failure modes: SOURCE EVIDENCE WEAK COMPARTMENT OR COMPLEX MISMATCH Sources checked: Reactome:R-HSA-983156 · Polyubiquitination of substrate SOURCE WEAK OR INFERRED Actual route: 983156 -> input -> 983131 -> hasComponent -> 976075 -> hasMember -> 8850899. Target membership and its explicit cytosol field are verified; the set-based compartment model lacks resolved target-specific localization support. SCF node976110, when present, is a Complex, not a F-box set. Supporting Evidence: PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. PMID:37161784 localises to the mitochondrial outer membrane in unstressed cells and mediates the constitutive ubiquitylation and degradation of the mitophagy receptors NIX and BNIP3 to suppress basal levels of mitophagy |
| GO:0005829 cytosol | TAS Reactome:R-HSA-983157 | MARK AS OVER ANNOTATED | Summary: The modeled cytosolic compartment is overextended to FBXL4 through a generic ligase set. Reason: This event reaches FBXL4 through generic E3 CandidateSet 976075. The target entity 8850899 itself explicitly assigns Q9UKA2 residues 1–621 to cytosol; the compartment is not merely a label on a parent complex. This modeled set membership does not establish a FBXL4-specific cytosolic pool. Primary experiments support an integral mitochondrial outer-membrane pool (PMID:36896912), and the historical fractionation study did not detect a postmitochondrial cytosolic pool under its tested conditions (PMID:23993194). Cytosolic exposure of a membrane protein does not establish localization in the organelle-excluding cytosol compartment. The source therefore overextends compartment evidence. This judgment does not exclude a conditional cytosolic pool or assert that outer membrane is a descendant of cytosol. The separate outer-membrane annotation remains supported. Propagation Review Root cause: SOURCE WEAK OR INFERRED Failure modes: SOURCE EVIDENCE WEAK COMPARTMENT OR COMPLEX MISMATCH Sources checked: Reactome:R-HSA-983157 · Interaction of E3 with substrate and E2-Ub complex SOURCE WEAK OR INFERRED Actual route: 983157 -> input -> 976075 -> hasMember -> 8850899. Target membership and its explicit cytosol field are verified; the set-based compartment model lacks resolved target-specific localization support. SCF node976110, when present, is a Complex, not a F-box set. Supporting Evidence: PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. PMID:37161784 localises to the mitochondrial outer membrane in unstressed cells and mediates the constitutive ubiquitylation and degradation of the mitophagy receptors NIX and BNIP3 to suppress basal levels of mitophagy |
| GO:0006511 ubiquitin-dependent protein catabolic process | TAS PMID:10531035 Identification of a family of human F-box proteins. | TOO GENERAL | Summary: FBXL4 promotes SCF-dependent proteasomal substrate turnover. Reason: The 1999 F-box-family paper supports a broad ubiquitin-dependent catabolic role. Target-specific later studies establish receptor ubiquitination and proteasomal turnover by SCF-FBXL4, already represented by retained GO:0031146. The genuine ontology ancestry places that process under GO:0006511. The broader process adds no independently demonstrated alternative degradation route; it remains biologically true and is not rejected on evidence-code grounds. Supporting Evidence: PMID:10531035 In fact, F-box proteins are one of the four subunits of ubiquitin protein ligases called SCFs. PMID:36896912 SCF-FBXL4 ubiquitinates BNIP3 and NIX to target them for degradation. |
| GO:0016607 nuclear speck | IDA GO_REF:0000052 | KEEP AS NON CORE | Summary: HPA reports nuclear-speck staining in three human cell lines. Reason: The actual HPA subcellular page/XML classifies HPA029140 staining as supported nuclear speckles in A-431, U-251MG and U2OS. One blue/red/green image per line was inspected and shows green nuclear punctate staining, consistent with the curated compartment. The antigen sequence matches Q9UKA2 residues 26–110, and antigen-array validation reports a single peak against its own antigen; neither observation substitutes for endogenous knockout/knockdown localization validation. Only one antibody and no speck-specific marker channel were independently inspected. PMID:23993194 reported no nuclear signal under different tested conditions, whereas the mitochondrial studies establish another location without disproving this cell-line observation. Retain the curator-assigned location as contextual, not a core nuclear mechanism. The inherited outer-membrane quote remains context, not proof of nonspecific staining. Supporting Evidence: PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. |
| GO:0019005 SCF ubiquitin ligase complex | IBA GO_REF:0000033 | ACCEPT | Summary: FBXL4 is an SCF substrate-recognition subunit. Reason: Actual target experiments in PMID:36896912 show tagged human FBXL4 associated with endogenous SKP1 and CUL1; deleting or altering its F-box disrupts that association. This supports membership as the substrate-recognition component, not independent catalytic activity of FBXL4. The actual PANTHER v19 tree places human leaf PTN002547906 below asserted node PTN002547163, and the ungapped 621-residue alignment exactly matches official Q9UKA2. The selected positive IBD assertion and literal donor list were read; the selected file contains no NOT row. Target functional evidence agrees with the ancestral role, with no observed target-specific loss of this function. Donor count is not used as a confidence measure, and the complete experimental history of every descendant was not independently rereviewed. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN002547163 · PTN002547163 SUPPORTS TRANSFER The actual ancestral node is on the exact-sequence target path in PTHR13318, officially PARTNER OF PAIRED, ISOFORM B-RELATED; target subfamily PTHR13318:SF152 is F-BOX_LRR-REPEAT PROTEIN 4. Node placement and independent target assays support the specified conserved role, not every function in the broad family. Supporting Evidence: PMID:36896912 We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. PMID:36896912 Pathogenic FBXL4 mutations disrupt SCF-FBXL4 assembly and impair substrate degradation. |
| GO:0031146 SCF-dependent proteasomal ubiquitin-dependent protein catabolic process | IBA GO_REF:0000033 | ACCEPT | Summary: FBXL4 contributes to SCF-mediated proteasomal turnover of mitophagy receptors. Reason: Actual target experiments in PMID:36896912 link FBXL4, its F-box and CUL1 to BNIP3/NIX ubiquitination and proteasome-associated turnover. PMID:37161784 adds receptor stability/rescue and polyubiquitination evidence in human cells. FBXL4 contributes substrate recognition to the SCF machinery; the process does not require FBXL4 itself to hydrolyze peptide bonds. The actual PANTHER v19 tree places human leaf PTN002547906 below asserted node PTN002547163, and the ungapped 621-residue alignment exactly matches official Q9UKA2. The selected positive IBD assertion and literal donor list were read; the selected file contains no NOT row. Target functional evidence agrees with the ancestral role, with no observed target-specific loss of this function. Donor count is not used as a confidence measure, and the complete experimental history of every descendant was not independently rereviewed. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN002547163 · PTN002547163 SUPPORTS TRANSFER The actual ancestral node is on the exact-sequence target path in PTHR13318, officially PARTNER OF PAIRED, ISOFORM B-RELATED; target subfamily PTHR13318:SF152 is F-BOX_LRR-REPEAT PROTEIN 4. Node placement and independent target assays support the specified conserved role, not every function in the broad family. Supporting Evidence: PMID:36896912 SCF-FBXL4 ubiquitinates BNIP3 and NIX to target them for degradation. PMID:37161784 localises to the mitochondrial outer membrane in unstressed cells and mediates the constitutive ubiquitylation and degradation of the mitophagy receptors NIX and BNIP3 to suppress basal levels of mitophagy |
| GO:0031146 SCF-dependent proteasomal ubiquitin-dependent protein catabolic process | NAS PMID:33234069 The FBXL family of F-box proteins: variations on a theme. | ACCEPT | Summary: FBXL4 recruits substrates to an SCF ligase for proteasomal degradation. Reason: The family review provides NAS context, and target-specific primary studies establish the role through SCF association, F-box perturbations, in-cell ubiquitination, receptor half-life/proteasome experiments and rescue. FBXL4 participates by substrate recognition and recruitment, while the assembled SCF/E2 machinery performs ubiquitin transfer. BNIP3 and BNIP3L/NIX turnover are distinct from the downstream lysosomal removal of mitochondria. Contextual 2011 JMJD2A/KDM4A turnover evidence is separately assessed without implying a purified binary interaction or an independently imaged nuclear FBXL4 pool. Supporting Evidence: PMID:33234069 F-box proteins (FBPs) are substrate-recruiting subunits of Skp1-cullin1-FBP (SCF)-type E3 ubiquitin ligases that determine which proteins are ubiquitinated. PMID:36896912 SCF-FBXL4 ubiquitinates BNIP3 and NIX to target them for degradation. |
| GO:1901524 regulation of mitophagy | NAS PMID:36135912 Mitochondrial Fission and Fusion: Molecular Mechanisms, Biol... | TOO GENERAL | Summary: The experimentally resolved mitophagy-regulatory direction is negative. Reason: The mitochondrial-dynamics review discusses FBXL4 in the disease and turnover context; its fusion model is not treated as a direct assay of fusion chemistry. Independent primary experiments establish restraint of mitophagy through mitophagy-receptor turnover. Retained GO:1901525 is an actual ontology descendant of GO:1901524 and preserves this signed role. The unsigned parent adds no demonstrated opposite regulatory action in the evidence assessed here. Supporting Evidence: PMID:36135912 Then, mitochondrial disorders caused by defects in fission and fusion are summarized, including disorders related to MFN2, MSTO1, OPA1, YME1L1, FBXL4, DNM1L, and MFF genes. PMID:36896912 we found that knockout (KO) of FBXL4, a mitochondrial disease gene, hyperactivates mitophagy at basal conditions. |
| GO:1901525 negative regulation of mitophagy | IDA PMID:38992176 PPTC7 antagonizes mitophagy by promoting BNIP3 and NIX degra... | ACCEPT | Summary: SCF-FBXL4 limits mitophagy by promoting BNIP3 and BNIP3L/NIX turnover. Reason: The actual PMID38992176 target perturbation, turnover and mitophagy assays support an inhibitory role assisted by PPTC7-dependent receptor recruitment. FBXL4 still associates with SKP1/CUL1 after PPTC7 knockout, separating substrate recruitment from assembly of the SCF core. The 2025 PMID40025034 study independently supports cooperation in human-cell ubiquitination/rescue assays, but reports a different PPTC7 D78A outcome and different effects of component overexpression. Cell lines, expression levels and treatment contexts differ; neither paper establishes a universal monotonic dose response or resolves those differences. The shared inhibitory pathway remains supported. Supporting Evidence: PMID:38992176 The SCFFBXL4 ubiquitin ligase complex suppresses mitophagy by controlling the degradation of BNIP3 and NIX mitophagy receptors PMID:38992176 a pool of PPTC7 on the mitochondrial outer membrane acts as an adaptor linking BNIP3 and NIX to FBXL4, facilitating the turnover of these mitophagy receptors |
| GO:1990756 ubiquitin-like ligase-substrate adaptor activity | IDA PMID:38992176 PPTC7 antagonizes mitophagy by promoting BNIP3 and NIX degra... | ACCEPT | Summary: FBXL4 is the substrate-recognition/adaptor component of SCF-mediated ubiquitination. Reason: FBXL4 associates with SKP1/CUL1 through its F-box and connects receptor substrates to the SCF machinery; PPTC7 facilitates receptor recruitment. The source supports this adaptor role through co-complex, mutagenesis, in-cell ubiquitination and turnover experiments. PMID38992176 explicitly distinguishes its modeled ternary arrangement from experimentally demonstrated simultaneous ternary occupancy. PMID40025034 adds cofactor-dependent in-cell assays, not a purified reconstitution or independent PPTC7 E3 catalytic activity. The evidence supports the FBXL4 substrate-adaptor molecular function without asserting that every co-IP proves a purified binary contact, that PPTC7 assembles SKP1/CUL1, or that FBXL4 itself supplies ubiquitin-transfer chemistry. Supporting Evidence: PMID:38992176 a pool of PPTC7 on the mitochondrial outer membrane acts as an adaptor linking BNIP3 and NIX to FBXL4, facilitating the turnover of these mitophagy receptors PMID:37102372 FBXL4 restricts NIX and BNIP3 levels via direct interaction and protein destabilization PMID:36896912 SCF-FBXL4 ubiquitinates BNIP3 and NIX to target them for degradation. |
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Download this section (compressed HTML)Q: Are there SCF-FBXL4 substrates beyond BNIP3, NIX and possibly PPTC7, and do they contribute to the mtDNA depletion phenotype?
Q: Is the reported effect of FBXL4 on DRP1-mediated fission direct, or secondary to altered mitophagy and mitochondrial mass?
Experiment: Compare endogenous knock-in D78A and D290N in matched human-cell backgrounds, controlling protein abundance/localization and measuring receptor association, SCF assembly, receptor ubiquitination/turnover and mitophagy.
Type: Matched endogenous mutant, recruitment and turnover assays
Experiment: Use endogenous small epitope tags with controls for mitochondrial integrity, graded permeabilization and matched protease conditions, then combine protease accessibility with membrane extraction and orthogonal imaging.
Type: Endogenous topology, fractionation and imaging comparison
What is not known — curated, literature-grounded statements of the open unknowns (the inverse of core functions).
Gap: The assessed studies do not resolve why PPTC7 D78A and excess-expression outcomes differ between cellular contexts, or how endogenous recruitment states account for those differences.
OPEN BIOLOGY RESIDUAL_SUBGAP
What is known: FBXL4 and PPTC7 cooperate in receptor turnover. FBXL4 association with SKP1/CUL1 persists after PPTC7 loss. The 2024 and 2025 studies differ in cell line, expression and treatment context, and D290N retains a residual catalytic-activity caveat.
Significance: Distinguishing recruitment, catalytic and dosage effects would define when PPTC7 supports SCF-FBXL4 receptor turnover.
What would resolve it: Compare endogenous knock-in D78A and D290N in matched human-cell backgrounds, controlling protein abundance/localization and measuring receptor association, SCF assembly, receptor ubiquitination/turnover and mitophagy.
Provenance (the field's own admissions):
Gap: The assessed 2013 protection/fractionation results and later outer-membrane results do not resolve the endogenous FBXL4 membrane orientation and compartment differences across their experimental conditions.
OPEN BIOLOGY RESIDUAL_SUBGAP
What is known: Actual experiments support both the historical intermembrane-space interpretation and a functional outer-membrane pool. The preparations, expression and protease conditions differ, and the older paper prints different trypsin-concentration units. Neither result establishes exclusive localization in all cells.
Significance: The orientation of the substrate-recognition region determines how the mitochondrial pool accesses cytosolic SCF components and receptor domains.
What would resolve it: Use endogenous small epitope tags with controls for mitochondrial integrity, graded permeabilization and matched protease conditions, then combine protease accessibility with membrane extraction and orthogonal imaging.
Provenance (the field's own admissions):
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