{"gene":"FBXL5","run_date":"2026-06-09T23:54:43","timeline":{"discoveries":[{"year":2020,"finding":"The C-terminal substrate-binding domain of FBXL5 harbors a [2Fe2S] iron-sulfur cluster in the oxidized state. A cryo-EM structure of the IRP2-FBXL5-SKP1 complex revealed that this cluster organizes the FBXL5 C-terminal loop responsible for recruiting IRP2. IRP2 binding to FBXL5 depends on the oxidized state of the [2Fe2S] cluster maintained by ambient oxygen, explaining hypoxia-induced IRP2 stabilization. FBXL5 also sterically dislodges IRP2 from iron-responsive element RNA to facilitate its turnover.","method":"Cryo-EM structure determination, in vitro ubiquitination assay, mutagenesis, EPR spectroscopy","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structure with functional validation, multiple orthogonal methods (structural, biochemical, spectroscopic) in a single rigorous study","pmids":["32126207"],"is_preprint":false},{"year":2012,"finding":"The N-terminal domain of FBXL5 adopts a hemerythrin-like α-helical bundle fold containing an unusual diiron center. This domain senses iron and oxygen availability by distinct mechanisms: iron limitation causes substantive structural changes in the domain, whereas oxygen depletion does not produce the same conformational changes. The domain controls accessibility of a degradation sequence required for proteasomal degradation of FBXL5 itself.","method":"X-ray crystallography, biochemical assays, mutagenesis, spectroscopy","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — atomic crystal structure combined with biochemical and spectroscopic characterization across two independent papers (PMID 22253436, 22648410)","pmids":["22253436","22648410"],"is_preprint":false},{"year":2011,"finding":"FBXL5 mediates iron-dependent proteasomal degradation of IRP2 in vivo. Fbxl5-null mice die in utero with excessive iron accumulation; embryonic lethality is rescued by simultaneous deletion of IRP2 but not IRP1, establishing that unrestrained IRP2 activity is the primary cause of death and placing FBXL5 upstream of IRP2 in iron homeostasis.","method":"Genetic epistasis (Fbxl5−/− × Irp2−/− double knockout mice), immunoblotting, tissue iron measurements","journal":"Cell metabolism","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean genetic epistasis in vivo with defined molecular phenotype, replicated across multiple studies","pmids":["21907140"],"is_preprint":false},{"year":2012,"finding":"FBXL5 is required for maintenance of cellular and systemic iron homeostasis in vivo. FBXL5-null mice fail to sense increased cellular iron, show constitutive IRP2 accumulation and misexpression of IRP2 target genes, and die during embryogenesis; viability is restored by IRP2 but not IRP1 deletion. Heterozygous mice show increased intestinal iron absorption via enhanced duodenal IRP2 responsiveness and upregulation of DMT-1.","method":"Conditional and constitutive knockout mice, genetic epistasis, gene expression analysis, hematological measurements","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple genetic models with defined molecular and physiological phenotypes, replicates PMID 21907140 with additional mechanistic detail","pmids":["23135277"],"is_preprint":false},{"year":2014,"finding":"HERC2, a large HECT-type E3 ubiquitin ligase, interacts with FBXL5 and targets it for ubiquitin-dependent proteasomal degradation, controlling FBXL5 basal turnover. Inhibition of HERC2-FBXL5 interaction or HERC2 depletion stabilizes FBXL5, leading to decreased intracellular ferrous iron.","method":"Proteomics/mass spectrometry interactome, Co-IP, RNA interference, ferrous iron measurements","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal interaction identified by proteomics and Co-IP, functional consequence demonstrated, single lab","pmids":["24778179"],"is_preprint":false},{"year":2019,"finding":"FBXL5 interacts with the CIA-targeting complex (composed of MMS19, FAM96B, and CIAO1) in an oxygen-dependent manner. This interaction, robust at 21% O2 but severely diminished at 1% O2, promotes FBXL5-mediated degradation of IRPs and links IRP-dependent iron homeostasis with Fe-S cluster assembly machinery.","method":"Co-IP, mass spectrometry, cell-based IRP degradation assays under varying O2 conditions","journal":"Molecular cell","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP plus functional assays, single lab, two orthogonal methods","pmids":["31229404"],"is_preprint":false},{"year":2017,"finding":"FBXL5 promotes IRP1 polyubiquitination and degradation when cytosolic iron-sulfur (Fe-S) cluster assembly (CIA) is impaired. FBXL5 and CIA act synergistically through both IRP1 and IRP2 to control iron metabolism. IRP1 Ser-138 phosphorylation is required for iron rescue when CIA is inhibited. A negative feedback loop exists whereby elevated IRP expression induces FBXL5 protein levels.","method":"siRNA knockdown, IRP1 phosphorylation-site mutants, polyubiquitination assays, cell viability assays","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple genetic perturbations with biochemical readouts, single lab","pmids":["28768766"],"is_preprint":false},{"year":2013,"finding":"FBXL5 is localized in the nucleus where it interacts with the transcription factor Snail1, promotes its polyubiquitination, impairs Snail1 DNA binding, and reduces Snail1 protein stability. Although polyubiquitination occurs in the nucleus, Snail1 is degraded in the cytosol. Lats2 phosphorylation of Snail1 prevents nuclear export but not FBXL5-mediated polyubiquitination. FBXL5 is downregulated by iron depletion and γ-irradiation, explaining Snail1 stabilization under these stresses.","method":"shRNA screening, co-immunoprecipitation, ubiquitination assay, subcellular fractionation, nuclear localization imaging","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal methods (Co-IP, ubiquitination assay, localization), single lab","pmids":["24157836"],"is_preprint":false},{"year":2014,"finding":"FBXL5 interacts with cortactin and targets it for ERK-dependent ubiquitylation and proteasomal degradation. ERK-mediated serine phosphorylation of cortactin at S405/S418 is required for FBXL5-induced degradation; the cortactinS405A/S418A phospho-null mutant resists FBXL5-induced degradation and shows enhanced gastric cancer cell migration.","method":"Co-immunoprecipitation, ubiquitination assay, site-directed mutagenesis, cell migration assays","journal":"Tumour biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, ubiquitination assay, phosphorylation-site mutagenesis, single lab","pmids":["24867096"],"is_preprint":false},{"year":2014,"finding":"FBXL5 directly interacts with hSSB1 (single-stranded DNA-binding protein 1) and targets it for SCF-mediated ubiquitination and degradation. ATM-mediated phosphorylation of hSSB1 at T117 prevents FBXL5-induced degradation. FBXL5 overexpression abrogates ATM signaling, DNA damage checkpoint activation, and increases radio- and chemo-sensitivity.","method":"Co-immunoprecipitation, ubiquitination assay, phosphorylation-site mutagenesis, DNA damage assays","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, ubiquitination assay, phosphosite mutants, single lab","pmids":["25249620"],"is_preprint":false},{"year":2015,"finding":"FBXL5 directly interacts with CITED2 and promotes its ubiquitination-dependent proteasomal degradation. FBXL5 depletion increases CITED2 levels; FBXL5 overexpression decreases CITED2 levels, impairs CITED2 interaction with the CH1 domain of p300, and enables transcriptional activity of the HIF-1α N-terminal transactivation domain.","method":"Co-immunoprecipitation, RNA interference, overexpression, FRET/BRET assay in living cells, reporter assay","journal":"Archives of biochemistry and biophysics","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP and functional assays, multiple methods, single lab","pmids":["25956243"],"is_preprint":false},{"year":2007,"finding":"FBXL5 interacts with p150Glued (dynactin subunit) both in vitro and in vivo, co-localizes with it in the cytoplasm with peri-nuclear enrichment, and promotes its poly-ubiquitination and protein turnover.","method":"Co-immunoprecipitation, in vitro binding assay, immunofluorescence colocalization, overexpression ubiquitination assay","journal":"Biochemical and biophysical research communications","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single Co-IP and single lab, limited follow-up on mechanism","pmids":["17532294"],"is_preprint":false},{"year":2017,"finding":"FBXL5-mediated regulation of cellular iron homeostasis is required for hematopoietic stem cell (HSC) self-renewal. Conditional deletion of Fbxl5 in mouse HSCs causes cellular iron overload, reduced HSC number, and stem cell exhaustion upon bone marrow transplantation. Suppression of IRP2 accumulation in FBXL5-deficient HSCs restores stem cell function.","method":"Conditional knockout mice, bone marrow transplantation, transcriptomic analysis, genetic epistasis (IRP2 suppression)","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — conditional knockout with transplantation assays and genetic rescue via IRP2 suppression, multiple orthogonal readouts","pmids":["28714470"],"is_preprint":false},{"year":2017,"finding":"Brain-specific deletion of FBXL5 in nestin-expressing neural stem progenitor cells (NSPCs) leads to IRP2 stabilization, iron accumulation, ROS generation, and aberrant NSPC and astroglia proliferation in the cerebral cortex. Pharmacological manipulation implicated mTOR signaling as the downstream effector of FBXL5 deficiency.","method":"Conditional knockout mice (nestin-Cre), iron measurements, ROS assays, mTOR pathway inhibition","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — conditional knockout with defined molecular and cellular phenotypes, single lab","pmids":["28069738"],"is_preprint":false},{"year":2017,"finding":"Redox state of the FBXL5 diiron center governs its conformation and stability. EPR, direct electrochemistry, SRCD, and fluorescence spectroscopy showed that redox reactions at the diiron center are accompanied by conformational changes and iron release, which are mechanistically linked to FBXL5 stability and its function as an iron/oxygen sensor.","method":"EPR spectroscopy, direct electrochemistry, SRCD, fluorescence emission spectroscopy, redox kinetics","journal":"Archives of biochemistry and biophysics","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — multiple biophysical methods but single lab, no mutagenesis confirmation","pmids":["28131773"],"is_preprint":false},{"year":2021,"finding":"At tissue-level O2 concentrations, ISC deficiency can activate IRP2 and promote ferroptosis sensitivity independently of IRP1, FBXL5, and changes in IRP2 protein level. IRP2 responds to Fe-S cluster synthesis suppression via a previously unidentified mechanism that does not involve FBXL5-mediated degradation.","method":"ISC synthesis inhibition, IRP RNA-binding assays, IRP1/IRP2 double knockout, FBXL5 knockdown, ferroptosis assays","journal":"Science advances","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic knockouts with RNA-binding assays demonstrating FBXL5-independent mechanism; negative result for FBXL5 involvement is mechanistically informative","pmids":["34039609"],"is_preprint":false},{"year":2023,"finding":"FBXL5 promotes ubiquitination and proteasomal degradation of TFEB (transcription factor EB). In alcoholic fatty liver disease, FBXL5 is upregulated and its interaction with TFEB promotes TFEB degradation, contributing to lipid accumulation; TFEB knockdown reverses the effect of FBXL5 inhibition.","method":"Co-immunoprecipitation, ubiquitination assay, siRNA knockdown, overexpression in HepG2 cells","journal":"Cellular signalling","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single Co-IP and ubiquitination assay, single lab, limited mechanistic detail in abstract","pmids":["37743009"],"is_preprint":false},{"year":2023,"finding":"FBXL5 mRNA translation is suppressed by the RNA-binding protein G3BP1, which stabilizes IRP2 by binding to and suppressing FBXL5 mRNA translation. Sodium arsenite intoxication activates this G3BP1-FBXL5-IRP2 axis, elevating labile iron and triggering ferroptosis.","method":"35S-methionine labeling, RIP assay, siRNA knockdown, ferroptosis assays, mouse kidney injury model","journal":"Journal of hazardous materials","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — translational regulation established by metabolic labeling and RIP, functional consequence confirmed in vivo, single lab","pmids":["38118197"],"is_preprint":false},{"year":2023,"finding":"FBXL5 is required for redox homeostasis and spindle assembly during mouse oocyte meiotic maturation. Fbxl5 silencing caused meiotic failure, overproduction of ROS, and abnormal accumulation of CITED2. An in vitro ubiquitination assay confirmed that FBXL5 directly interacts with CITED2 and mediates its proteasomal degradation in oocytes.","method":"siRNA knockdown in mouse oocytes, ROS assay, immunofluorescence spindle imaging, in vitro ubiquitination assay, Co-IP","journal":"FASEB journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vitro ubiquitination assay plus loss-of-function phenotype, single lab","pmids":["37462473"],"is_preprint":false},{"year":2026,"finding":"FBXL5 promotes YAP1 protein degradation through the ubiquitin-proteasome pathway without altering YAP1 transcript levels. Galectin-3 (Gal-3) binds FBXL5 and enhances its expression, increasing YAP1 degradation and restraining colorectal cancer growth in vivo.","method":"Co-immunoprecipitation, ubiquitination assay, xenograft mouse model, cell proliferation assay","journal":"Frontiers in immunology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — Co-IP and ubiquitination assay, single lab, limited mechanistic detail; in vivo data support functional consequence","pmids":["42093989"],"is_preprint":false}],"current_model":"FBXL5 is the substrate-recognition subunit of an SCF (SKP1-CUL1-RBX1-FBXL5) E3 ubiquitin ligase complex that senses intracellular iron and oxygen via an N-terminal hemerythrin-like diiron center and a C-terminal [2Fe2S] cluster; when iron and oxygen are sufficient, these redox-sensitive clusters stabilize FBXL5 and enable it to recruit, polyubiquitinate, and direct IRP2 (and IRP1) to proteasomal degradation—a cryo-EM structure of the IRP2-FBXL5-SKP1 complex revealed how the oxidized [2Fe2S] cluster organizes the IRP2-binding loop and sterically dislodges IRP2 from IRE-RNA—while FBXL5 stability is itself regulated upstream by HERC2-mediated ubiquitination and by the CIA-targeting complex in an oxygen-dependent manner; additional substrates include Snail1 (nuclear ubiquitination controlling EMT), CITED2 (modulating HIF-1α transcription), hSSB1 (DNA damage response), cortactin (cell migration), p150Glued, TFEB, and YAP1, establishing FBXL5 as a multi-substrate iron/oxygen sensor whose primary in vivo role is maintenance of iron homeostasis in diverse tissues including liver, hematopoietic stem cells, neural progenitors, and cardiomyocytes."},"narrative":{"mechanistic_narrative":"FBXL5 is the substrate-recognition subunit of an SCF (SKP1-CUL1-RBX1) E3 ubiquitin ligase that functions as the master iron/oxygen sensor controlling cellular and systemic iron homeostasis [PMID:21907140, PMID:23135277]. It detects iron and oxygen through two redox-sensitive metal centers: an N-terminal hemerythrin-like α-helical bundle harboring a diiron center whose iron occupancy and redox state govern accessibility of a degradation sequence and thereby FBXL5's own stability, and a C-terminal [2Fe2S] cluster whose oxygen-maintained oxidized state organizes the substrate-binding loop [PMID:32126207, PMID:22253436, PMID:22648410, PMID:28131773]. When iron and oxygen are replete, stabilized FBXL5 recruits, polyubiquitinates, and targets the iron-regulatory proteins IRP2 and IRP1 for proteasomal degradation, with the oxidized [2Fe2S] cluster sterically dislodging IRP2 from iron-responsive-element RNA—explaining hypoxia- and iron-limitation-induced IRP2 stabilization [PMID:32126207, PMID:22253436, PMID:22648410]. Genetically, unrestrained IRP2 activity is the decisive output of FBXL5 loss: Fbxl5-null embryonic lethality and iron overload are rescued by deletion of IRP2 but not IRP1, and FBXL5-dependent iron control is required for hematopoietic stem cell self-renewal and neural progenitor homeostasis [PMID:21907140, PMID:23135277, PMID:28714470, PMID:28069738]. FBXL5 abundance is itself set upstream by HERC2-mediated ubiquitination and by oxygen-dependent association with the CIA-targeting complex (MMS19/FAM96B/CIAO1) that links it to Fe-S cluster assembly [PMID:24778179, PMID:31229404]. Beyond iron regulation, FBXL5 directs phosphorylation-gated degradation of additional substrates, including nuclear Snail1 in EMT control, hSSB1 in the DNA damage response, cortactin in cell migration, and CITED2 to modulate HIF-1α transactivation [PMID:24157836, PMID:24867096, PMID:25249620, PMID:25956243].","teleology":[{"year":2007,"claim":"First evidence that FBXL5 acts as a substrate-targeting E3 component identified a physical partner and ubiquitination target, establishing it as a functional ubiquitin ligase subunit.","evidence":"Co-IP, in vitro binding, colocalization, and ubiquitination assays with p150Glued in cells","pmids":["17532294"],"confidence":"Low","gaps":["Single Co-IP from a single lab with limited mechanistic follow-up","No demonstration of SCF complex assembly or physiological consequence"]},{"year":2011,"claim":"Genetic epistasis defined FBXL5's primary in vivo role by showing it degrades IRP2 to restrain iron accumulation, placing it upstream of IRP2 in iron homeostasis.","evidence":"Fbxl5−/− × Irp2−/− double-knockout mice, immunoblotting, tissue iron measurements","pmids":["21907140"],"confidence":"High","gaps":["Did not resolve how FBXL5 senses iron molecularly","Tissue- and cell-type-specific roles not yet dissected"]},{"year":2012,"claim":"Structural and physiological work established the iron/oxygen-sensing mechanism: an N-terminal hemerythrin-like diiron domain controls FBXL5 stability, and FBXL5 is required for cellular and systemic iron homeostasis.","evidence":"X-ray crystallography, spectroscopy, mutagenesis, plus conditional/constitutive knockout mice with iron and gene-expression readouts","pmids":["22253436","22648410","23135277"],"confidence":"High","gaps":["Did not explain how oxygen sensing distinct from iron sensing is transmitted to substrate recruitment","Mechanism of the C-terminal substrate domain unresolved"]},{"year":2013,"claim":"Identification of nuclear Snail1 as a substrate extended FBXL5 beyond iron biology to EMT regulation and linked its activity to iron- and irradiation-responsive stress signaling.","evidence":"shRNA, Co-IP, ubiquitination assay, subcellular fractionation, and localization imaging","pmids":["24157836"],"confidence":"Medium","gaps":["Single lab","Spatial paradox of nuclear ubiquitination with cytosolic degradation not fully resolved"]},{"year":2014,"claim":"FBXL5's own turnover was shown to be governed by HERC2, defining an upstream layer that sets sensor abundance, while phosphorylation-gated degradation of cortactin and hSSB1 revealed substrate selection coupled to ERK and ATM signaling.","evidence":"Interactome MS, Co-IP, RNAi, iron measurements (HERC2); Co-IP, ubiquitination, and phosphosite mutagenesis (cortactin, hSSB1)","pmids":["24778179","24867096","25249620"],"confidence":"Medium","gaps":["Each substrate characterized in a single lab","How phosphorylation switches gate FBXL5 recruitment mechanistically unknown"]},{"year":2015,"claim":"FBXL5-mediated degradation of CITED2 connected the ligase to HIF-1α transcriptional output, broadening its role in oxygen-responsive gene regulation.","evidence":"Co-IP, RNAi, overexpression, FRET/BRET, and reporter assays","pmids":["25956243"],"confidence":"Medium","gaps":["Single lab","In vivo relevance of the FBXL5-CITED2-HIF axis not established here"]},{"year":2017,"claim":"Tissue-level studies and CIA crosstalk established FBXL5 as essential for stem cell and progenitor iron control and linked it functionally to Fe-S cluster assembly through both IRP1 and IRP2.","evidence":"Conditional knockouts in HSCs and neural progenitors with IRP2-suppression rescue; siRNA, IRP1 phosphosite mutants, and polyubiquitination assays for CIA crosstalk","pmids":["28714470","28069738","28768766"],"confidence":"High","gaps":["mTOR as downstream effector in NSPCs inferred pharmacologically","Molecular coupling between CIA status and FBXL5 activity not fully defined"]},{"year":2019,"claim":"Oxygen-dependent association of FBXL5 with the CIA-targeting complex (MMS19/FAM96B/CIAO1) provided a mechanism linking ambient oxygen to FBXL5-mediated IRP degradation.","evidence":"Co-IP, mass spectrometry, and IRP degradation assays under varied O2","pmids":["31229404"],"confidence":"Medium","gaps":["Single lab","Structural basis of the oxygen-dependent interaction not determined"]},{"year":2020,"claim":"A cryo-EM structure of the IRP2-FBXL5-SKP1 complex resolved the C-terminal [2Fe2S] cluster as the oxygen-sensing module that organizes the substrate-binding loop and dislodges IRP2 from IRE-RNA, completing the dual-sensor mechanism.","evidence":"Cryo-EM, in vitro ubiquitination, mutagenesis, EPR spectroscopy","pmids":["32126207"],"confidence":"High","gaps":["Structure of the full SCF holocomplex on substrate not solved","Integration of N-terminal diiron and C-terminal [2Fe2S] signals into one stability/activity output not structurally unified"]},{"year":2021,"claim":"A negative-control study showed IRP2 can be activated by Fe-S cluster deficiency at tissue O2 levels independently of FBXL5, delimiting the boundary of the FBXL5-dependent regulatory axis.","evidence":"ISC inhibition, IRP RNA-binding assays, IRP1/IRP2 double knockout, FBXL5 knockdown, ferroptosis assays","pmids":["34039609"],"confidence":"Medium","gaps":["FBXL5-independent IRP2 activation mechanism not identified","Physiological contexts where each pathway dominates unclear"]},{"year":2023,"claim":"Translational and disease-context studies expanded FBXL5 regulation and substrate scope, showing G3BP1-mediated suppression of FBXL5 translation drives ferroptosis, and identifying TFEB and oocyte CITED2 as functionally relevant targets.","evidence":"35S labeling/RIP and in vivo kidney injury (G3BP1); Co-IP/ubiquitination/siRNA in HepG2 (TFEB); oocyte siRNA, ROS, spindle imaging, in vitro ubiquitination (CITED2)","pmids":["38118197","37743009","37462473"],"confidence":"Medium","gaps":["TFEB and YAP1 substrate findings rest on single Co-IP/ubiquitination studies","Whether these substrates depend on FBXL5's iron-sensing state is untested"]},{"year":2026,"claim":"FBXL5 was implicated in tumor suppression via Galectin-3-enhanced degradation of YAP1, extending its substrate repertoire to Hippo-pathway control.","evidence":"Co-IP, ubiquitination assay, xenograft model, proliferation assays","pmids":["42093989"],"confidence":"Low","gaps":["Single lab with limited mechanistic detail","Degron and recruitment determinants for YAP1 unknown"]},{"year":null,"claim":"How the N-terminal diiron and C-terminal [2Fe2S] sensors are jointly integrated to set both FBXL5 stability and substrate selectivity across its many non-IRP targets remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified structural model coupling both metal centers to substrate choice","Whether non-IRP substrates are regulated by iron/oxygen state is untested","Tissue-specific substrate prioritization unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,2,7,8,9,10]},{"term_id":"GO:0016740","term_label":"transferase activity","supporting_discovery_ids":[0,2]},{"term_id":"GO:0140299","term_label":"molecular sensor activity","supporting_discovery_ids":[0,1,14]},{"term_id":"GO:0140313","term_label":"molecular sequestering activity","supporting_discovery_ids":[0]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[7]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[11]}],"pathway":[{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[2,3,12]},{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[0,2,4]},{"term_id":"R-HSA-8953897","term_label":"Cellular responses to stimuli","supporting_discovery_ids":[0,5]}],"complexes":["SCF (SKP1-CUL1-RBX1-FBXL5) E3 ubiquitin ligase"],"partners":["IRP2","IRP1","SKP1","HERC2","SNAIL1","CITED2","HSSB1","CORTACTIN"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9UKA1","full_name":"F-box/LRR-repeat protein 5","aliases":["F-box and leucine-rich repeat protein 5","F-box protein FBL4/FBL5","p45SKP2-like protein"],"length_aa":691,"mass_kda":78.6,"function":"Component of some SCF (SKP1-cullin-F-box) protein ligase complex that plays a central role in iron homeostasis by promoting the ubiquitination and subsequent degradation of IREB2/IRP2 (PubMed:19762596, PubMed:19762597). The C-terminal domain of FBXL5 contains a redox-sensitive [2Fe-2S] cluster that, upon oxidation, promotes binding to IRP2 to effect its oxygen-dependent degradation (PubMed:32126207). Under iron deficiency conditions, the N-terminal hemerythrin-like (Hr) region, which contains a diiron metal center, cannot bind iron and undergoes conformational changes that destabilize the FBXL5 protein and cause its ubiquitination and degradation (PubMed:19762596, PubMed:19762597). When intracellular iron levels start rising, the Hr region is stabilized (PubMed:19762596, PubMed:19762597). Additional increases in iron levels facilitate the assembly and incorporation of a redox active [2Fe-2S] cluster in the C-terminal domain (PubMed:32126207). Only when oxygen level is high enough to maintain the cluster in its oxidized state can FBXL5 recruit IRP2 as a substrate for polyubiquitination and degradation (PubMed:32126207). Promotes ubiquitination and subsequent degradation of the dynactin complex component DCTN1 (PubMed:17532294). Within the nucleus, promotes the ubiquitination of SNAI1; preventing its interaction with DNA and promoting its degradation (PubMed:24157836). Negatively regulates DNA damage response by mediating the ubiquitin-proteasome degradation of the DNA repair protein NABP2 (PubMed:25249620)","subcellular_location":"Cytoplasm, perinuclear region; Nucleus","url":"https://www.uniprot.org/uniprotkb/Q9UKA1/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/FBXL5","classification":"Not Classified","n_dependent_lines":25,"n_total_lines":1208,"dependency_fraction":0.020695364238410598},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/FBXL5","total_profiled":1310},"omim":[{"mim_id":"605655","title":"F-BOX AND LEUCINE-RICH REPEAT PROTEIN 5; FBXL5","url":"https://www.omim.org/entry/605655"},{"mim_id":"147582","title":"IRON-RESPONSIVE ELEMENT-BINDING PROTEIN 2; IREB2","url":"https://www.omim.org/entry/147582"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/FBXL5"},"hgnc":{"alias_symbol":["FBL4","FBL5","FLR1"],"prev_symbol":["CC2D2A-AS1"]},"alphafold":{"accession":"Q9UKA1","domains":[{"cath_id":"1.20.120.520","chopping":"7-183","consensus_level":"high","plddt":85.1282,"start":7,"end":183}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9UKA1","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9UKA1-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9UKA1-F1-predicted_aligned_error_v6.png","plddt_mean":69.62},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=FBXL5","jax_strain_url":"https://www.jax.org/strain/search?query=FBXL5"},"sequence":{"accession":"Q9UKA1","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9UKA1.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9UKA1/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9UKA1"}},"corpus_meta":[{"pmid":"9235926","id":"PMC_9235926","title":"AP1-mediated multidrug resistance in Saccharomyces cerevisiae requires FLR1 encoding a transporter of the major facilitator superfamily.","date":"1997","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/9235926","citation_count":162,"is_preprint":false},{"pmid":"34039609","id":"PMC_34039609","title":"Iron-sulfur cluster deficiency can be sensed by IRP2 and regulates iron homeostasis and sensitivity to ferroptosis independent of IRP1 and FBXL5.","date":"2021","source":"Science advances","url":"https://pubmed.ncbi.nlm.nih.gov/34039609","citation_count":158,"is_preprint":false},{"pmid":"32126207","id":"PMC_32126207","title":"FBXL5 Regulates IRP2 Stability in Iron Homeostasis via an Oxygen-Responsive [2Fe2S] Cluster.","date":"2020","source":"Molecular cell","url":"https://pubmed.ncbi.nlm.nih.gov/32126207","citation_count":145,"is_preprint":false},{"pmid":"21907140","id":"PMC_21907140","title":"The FBXL5-IRP2 axis is integral to control of iron metabolism in 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A cryo-EM structure of the IRP2-FBXL5-SKP1 complex revealed that this cluster organizes the FBXL5 C-terminal loop responsible for recruiting IRP2. IRP2 binding to FBXL5 depends on the oxidized state of the [2Fe2S] cluster maintained by ambient oxygen, explaining hypoxia-induced IRP2 stabilization. FBXL5 also sterically dislodges IRP2 from iron-responsive element RNA to facilitate its turnover.\",\n      \"method\": \"Cryo-EM structure determination, in vitro ubiquitination assay, mutagenesis, EPR spectroscopy\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structure with functional validation, multiple orthogonal methods (structural, biochemical, spectroscopic) in a single rigorous study\",\n      \"pmids\": [\"32126207\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"The N-terminal domain of FBXL5 adopts a hemerythrin-like α-helical bundle fold containing an unusual diiron center. This domain senses iron and oxygen availability by distinct mechanisms: iron limitation causes substantive structural changes in the domain, whereas oxygen depletion does not produce the same conformational changes. The domain controls accessibility of a degradation sequence required for proteasomal degradation of FBXL5 itself.\",\n      \"method\": \"X-ray crystallography, biochemical assays, mutagenesis, spectroscopy\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — atomic crystal structure combined with biochemical and spectroscopic characterization across two independent papers (PMID 22253436, 22648410)\",\n      \"pmids\": [\"22253436\", \"22648410\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"FBXL5 mediates iron-dependent proteasomal degradation of IRP2 in vivo. Fbxl5-null mice die in utero with excessive iron accumulation; embryonic lethality is rescued by simultaneous deletion of IRP2 but not IRP1, establishing that unrestrained IRP2 activity is the primary cause of death and placing FBXL5 upstream of IRP2 in iron homeostasis.\",\n      \"method\": \"Genetic epistasis (Fbxl5−/− × Irp2−/− double knockout mice), immunoblotting, tissue iron measurements\",\n      \"journal\": \"Cell metabolism\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean genetic epistasis in vivo with defined molecular phenotype, replicated across multiple studies\",\n      \"pmids\": [\"21907140\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"FBXL5 is required for maintenance of cellular and systemic iron homeostasis in vivo. FBXL5-null mice fail to sense increased cellular iron, show constitutive IRP2 accumulation and misexpression of IRP2 target genes, and die during embryogenesis; viability is restored by IRP2 but not IRP1 deletion. Heterozygous mice show increased intestinal iron absorption via enhanced duodenal IRP2 responsiveness and upregulation of DMT-1.\",\n      \"method\": \"Conditional and constitutive knockout mice, genetic epistasis, gene expression analysis, hematological measurements\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple genetic models with defined molecular and physiological phenotypes, replicates PMID 21907140 with additional mechanistic detail\",\n      \"pmids\": [\"23135277\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"HERC2, a large HECT-type E3 ubiquitin ligase, interacts with FBXL5 and targets it for ubiquitin-dependent proteasomal degradation, controlling FBXL5 basal turnover. Inhibition of HERC2-FBXL5 interaction or HERC2 depletion stabilizes FBXL5, leading to decreased intracellular ferrous iron.\",\n      \"method\": \"Proteomics/mass spectrometry interactome, Co-IP, RNA interference, ferrous iron measurements\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal interaction identified by proteomics and Co-IP, functional consequence demonstrated, single lab\",\n      \"pmids\": [\"24778179\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"FBXL5 interacts with the CIA-targeting complex (composed of MMS19, FAM96B, and CIAO1) in an oxygen-dependent manner. This interaction, robust at 21% O2 but severely diminished at 1% O2, promotes FBXL5-mediated degradation of IRPs and links IRP-dependent iron homeostasis with Fe-S cluster assembly machinery.\",\n      \"method\": \"Co-IP, mass spectrometry, cell-based IRP degradation assays under varying O2 conditions\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP plus functional assays, single lab, two orthogonal methods\",\n      \"pmids\": [\"31229404\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"FBXL5 promotes IRP1 polyubiquitination and degradation when cytosolic iron-sulfur (Fe-S) cluster assembly (CIA) is impaired. FBXL5 and CIA act synergistically through both IRP1 and IRP2 to control iron metabolism. IRP1 Ser-138 phosphorylation is required for iron rescue when CIA is inhibited. A negative feedback loop exists whereby elevated IRP expression induces FBXL5 protein levels.\",\n      \"method\": \"siRNA knockdown, IRP1 phosphorylation-site mutants, polyubiquitination assays, cell viability assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple genetic perturbations with biochemical readouts, single lab\",\n      \"pmids\": [\"28768766\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"FBXL5 is localized in the nucleus where it interacts with the transcription factor Snail1, promotes its polyubiquitination, impairs Snail1 DNA binding, and reduces Snail1 protein stability. Although polyubiquitination occurs in the nucleus, Snail1 is degraded in the cytosol. Lats2 phosphorylation of Snail1 prevents nuclear export but not FBXL5-mediated polyubiquitination. FBXL5 is downregulated by iron depletion and γ-irradiation, explaining Snail1 stabilization under these stresses.\",\n      \"method\": \"shRNA screening, co-immunoprecipitation, ubiquitination assay, subcellular fractionation, nuclear localization imaging\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal methods (Co-IP, ubiquitination assay, localization), single lab\",\n      \"pmids\": [\"24157836\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"FBXL5 interacts with cortactin and targets it for ERK-dependent ubiquitylation and proteasomal degradation. ERK-mediated serine phosphorylation of cortactin at S405/S418 is required for FBXL5-induced degradation; the cortactinS405A/S418A phospho-null mutant resists FBXL5-induced degradation and shows enhanced gastric cancer cell migration.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, site-directed mutagenesis, cell migration assays\",\n      \"journal\": \"Tumour biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, ubiquitination assay, phosphorylation-site mutagenesis, single lab\",\n      \"pmids\": [\"24867096\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"FBXL5 directly interacts with hSSB1 (single-stranded DNA-binding protein 1) and targets it for SCF-mediated ubiquitination and degradation. ATM-mediated phosphorylation of hSSB1 at T117 prevents FBXL5-induced degradation. FBXL5 overexpression abrogates ATM signaling, DNA damage checkpoint activation, and increases radio- and chemo-sensitivity.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, phosphorylation-site mutagenesis, DNA damage assays\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, ubiquitination assay, phosphosite mutants, single lab\",\n      \"pmids\": [\"25249620\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"FBXL5 directly interacts with CITED2 and promotes its ubiquitination-dependent proteasomal degradation. FBXL5 depletion increases CITED2 levels; FBXL5 overexpression decreases CITED2 levels, impairs CITED2 interaction with the CH1 domain of p300, and enables transcriptional activity of the HIF-1α N-terminal transactivation domain.\",\n      \"method\": \"Co-immunoprecipitation, RNA interference, overexpression, FRET/BRET assay in living cells, reporter assay\",\n      \"journal\": \"Archives of biochemistry and biophysics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP and functional assays, multiple methods, single lab\",\n      \"pmids\": [\"25956243\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"FBXL5 interacts with p150Glued (dynactin subunit) both in vitro and in vivo, co-localizes with it in the cytoplasm with peri-nuclear enrichment, and promotes its poly-ubiquitination and protein turnover.\",\n      \"method\": \"Co-immunoprecipitation, in vitro binding assay, immunofluorescence colocalization, overexpression ubiquitination assay\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single Co-IP and single lab, limited follow-up on mechanism\",\n      \"pmids\": [\"17532294\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"FBXL5-mediated regulation of cellular iron homeostasis is required for hematopoietic stem cell (HSC) self-renewal. Conditional deletion of Fbxl5 in mouse HSCs causes cellular iron overload, reduced HSC number, and stem cell exhaustion upon bone marrow transplantation. Suppression of IRP2 accumulation in FBXL5-deficient HSCs restores stem cell function.\",\n      \"method\": \"Conditional knockout mice, bone marrow transplantation, transcriptomic analysis, genetic epistasis (IRP2 suppression)\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — conditional knockout with transplantation assays and genetic rescue via IRP2 suppression, multiple orthogonal readouts\",\n      \"pmids\": [\"28714470\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Brain-specific deletion of FBXL5 in nestin-expressing neural stem progenitor cells (NSPCs) leads to IRP2 stabilization, iron accumulation, ROS generation, and aberrant NSPC and astroglia proliferation in the cerebral cortex. Pharmacological manipulation implicated mTOR signaling as the downstream effector of FBXL5 deficiency.\",\n      \"method\": \"Conditional knockout mice (nestin-Cre), iron measurements, ROS assays, mTOR pathway inhibition\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — conditional knockout with defined molecular and cellular phenotypes, single lab\",\n      \"pmids\": [\"28069738\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Redox state of the FBXL5 diiron center governs its conformation and stability. EPR, direct electrochemistry, SRCD, and fluorescence spectroscopy showed that redox reactions at the diiron center are accompanied by conformational changes and iron release, which are mechanistically linked to FBXL5 stability and its function as an iron/oxygen sensor.\",\n      \"method\": \"EPR spectroscopy, direct electrochemistry, SRCD, fluorescence emission spectroscopy, redox kinetics\",\n      \"journal\": \"Archives of biochemistry and biophysics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — multiple biophysical methods but single lab, no mutagenesis confirmation\",\n      \"pmids\": [\"28131773\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"At tissue-level O2 concentrations, ISC deficiency can activate IRP2 and promote ferroptosis sensitivity independently of IRP1, FBXL5, and changes in IRP2 protein level. IRP2 responds to Fe-S cluster synthesis suppression via a previously unidentified mechanism that does not involve FBXL5-mediated degradation.\",\n      \"method\": \"ISC synthesis inhibition, IRP RNA-binding assays, IRP1/IRP2 double knockout, FBXL5 knockdown, ferroptosis assays\",\n      \"journal\": \"Science advances\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic knockouts with RNA-binding assays demonstrating FBXL5-independent mechanism; negative result for FBXL5 involvement is mechanistically informative\",\n      \"pmids\": [\"34039609\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"FBXL5 promotes ubiquitination and proteasomal degradation of TFEB (transcription factor EB). In alcoholic fatty liver disease, FBXL5 is upregulated and its interaction with TFEB promotes TFEB degradation, contributing to lipid accumulation; TFEB knockdown reverses the effect of FBXL5 inhibition.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, siRNA knockdown, overexpression in HepG2 cells\",\n      \"journal\": \"Cellular signalling\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single Co-IP and ubiquitination assay, single lab, limited mechanistic detail in abstract\",\n      \"pmids\": [\"37743009\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"FBXL5 mRNA translation is suppressed by the RNA-binding protein G3BP1, which stabilizes IRP2 by binding to and suppressing FBXL5 mRNA translation. Sodium arsenite intoxication activates this G3BP1-FBXL5-IRP2 axis, elevating labile iron and triggering ferroptosis.\",\n      \"method\": \"35S-methionine labeling, RIP assay, siRNA knockdown, ferroptosis assays, mouse kidney injury model\",\n      \"journal\": \"Journal of hazardous materials\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — translational regulation established by metabolic labeling and RIP, functional consequence confirmed in vivo, single lab\",\n      \"pmids\": [\"38118197\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"FBXL5 is required for redox homeostasis and spindle assembly during mouse oocyte meiotic maturation. Fbxl5 silencing caused meiotic failure, overproduction of ROS, and abnormal accumulation of CITED2. An in vitro ubiquitination assay confirmed that FBXL5 directly interacts with CITED2 and mediates its proteasomal degradation in oocytes.\",\n      \"method\": \"siRNA knockdown in mouse oocytes, ROS assay, immunofluorescence spindle imaging, in vitro ubiquitination assay, Co-IP\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro ubiquitination assay plus loss-of-function phenotype, single lab\",\n      \"pmids\": [\"37462473\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"FBXL5 promotes YAP1 protein degradation through the ubiquitin-proteasome pathway without altering YAP1 transcript levels. Galectin-3 (Gal-3) binds FBXL5 and enhances its expression, increasing YAP1 degradation and restraining colorectal cancer growth in vivo.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, xenograft mouse model, cell proliferation assay\",\n      \"journal\": \"Frontiers in immunology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — Co-IP and ubiquitination assay, single lab, limited mechanistic detail; in vivo data support functional consequence\",\n      \"pmids\": [\"42093989\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"FBXL5 is the substrate-recognition subunit of an SCF (SKP1-CUL1-RBX1-FBXL5) E3 ubiquitin ligase complex that senses intracellular iron and oxygen via an N-terminal hemerythrin-like diiron center and a C-terminal [2Fe2S] cluster; when iron and oxygen are sufficient, these redox-sensitive clusters stabilize FBXL5 and enable it to recruit, polyubiquitinate, and direct IRP2 (and IRP1) to proteasomal degradation—a cryo-EM structure of the IRP2-FBXL5-SKP1 complex revealed how the oxidized [2Fe2S] cluster organizes the IRP2-binding loop and sterically dislodges IRP2 from IRE-RNA—while FBXL5 stability is itself regulated upstream by HERC2-mediated ubiquitination and by the CIA-targeting complex in an oxygen-dependent manner; additional substrates include Snail1 (nuclear ubiquitination controlling EMT), CITED2 (modulating HIF-1α transcription), hSSB1 (DNA damage response), cortactin (cell migration), p150Glued, TFEB, and YAP1, establishing FBXL5 as a multi-substrate iron/oxygen sensor whose primary in vivo role is maintenance of iron homeostasis in diverse tissues including liver, hematopoietic stem cells, neural progenitors, and cardiomyocytes.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"FBXL5 is the substrate-recognition subunit of an SCF (SKP1-CUL1-RBX1) E3 ubiquitin ligase that functions as the master iron/oxygen sensor controlling cellular and systemic iron homeostasis [#2, #3]. It detects iron and oxygen through two redox-sensitive metal centers: an N-terminal hemerythrin-like α-helical bundle harboring a diiron center whose iron occupancy and redox state govern accessibility of a degradation sequence and thereby FBXL5's own stability, and a C-terminal [2Fe2S] cluster whose oxygen-maintained oxidized state organizes the substrate-binding loop [#0, #1, #14]. When iron and oxygen are replete, stabilized FBXL5 recruits, polyubiquitinates, and targets the iron-regulatory proteins IRP2 and IRP1 for proteasomal degradation, with the oxidized [2Fe2S] cluster sterically dislodging IRP2 from iron-responsive-element RNA—explaining hypoxia- and iron-limitation-induced IRP2 stabilization [#0, #1]. Genetically, unrestrained IRP2 activity is the decisive output of FBXL5 loss: Fbxl5-null embryonic lethality and iron overload are rescued by deletion of IRP2 but not IRP1, and FBXL5-dependent iron control is required for hematopoietic stem cell self-renewal and neural progenitor homeostasis [#2, #3, #12, #13]. FBXL5 abundance is itself set upstream by HERC2-mediated ubiquitination and by oxygen-dependent association with the CIA-targeting complex (MMS19/FAM96B/CIAO1) that links it to Fe-S cluster assembly [#4, #5]. Beyond iron regulation, FBXL5 directs phosphorylation-gated degradation of additional substrates, including nuclear Snail1 in EMT control, hSSB1 in the DNA damage response, cortactin in cell migration, and CITED2 to modulate HIF-1α transactivation [#7, #8, #9, #10].\",\n  \"teleology\": [\n    {\n      \"year\": 2007,\n      \"claim\": \"First evidence that FBXL5 acts as a substrate-targeting E3 component identified a physical partner and ubiquitination target, establishing it as a functional ubiquitin ligase subunit.\",\n      \"evidence\": \"Co-IP, in vitro binding, colocalization, and ubiquitination assays with p150Glued in cells\",\n      \"pmids\": [\"17532294\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single Co-IP from a single lab with limited mechanistic follow-up\", \"No demonstration of SCF complex assembly or physiological consequence\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Genetic epistasis defined FBXL5's primary in vivo role by showing it degrades IRP2 to restrain iron accumulation, placing it upstream of IRP2 in iron homeostasis.\",\n      \"evidence\": \"Fbxl5−/− × Irp2−/− double-knockout mice, immunoblotting, tissue iron measurements\",\n      \"pmids\": [\"21907140\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not resolve how FBXL5 senses iron molecularly\", \"Tissue- and cell-type-specific roles not yet dissected\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Structural and physiological work established the iron/oxygen-sensing mechanism: an N-terminal hemerythrin-like diiron domain controls FBXL5 stability, and FBXL5 is required for cellular and systemic iron homeostasis.\",\n      \"evidence\": \"X-ray crystallography, spectroscopy, mutagenesis, plus conditional/constitutive knockout mice with iron and gene-expression readouts\",\n      \"pmids\": [\"22253436\", \"22648410\", \"23135277\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not explain how oxygen sensing distinct from iron sensing is transmitted to substrate recruitment\", \"Mechanism of the C-terminal substrate domain unresolved\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Identification of nuclear Snail1 as a substrate extended FBXL5 beyond iron biology to EMT regulation and linked its activity to iron- and irradiation-responsive stress signaling.\",\n      \"evidence\": \"shRNA, Co-IP, ubiquitination assay, subcellular fractionation, and localization imaging\",\n      \"pmids\": [\"24157836\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab\", \"Spatial paradox of nuclear ubiquitination with cytosolic degradation not fully resolved\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"FBXL5's own turnover was shown to be governed by HERC2, defining an upstream layer that sets sensor abundance, while phosphorylation-gated degradation of cortactin and hSSB1 revealed substrate selection coupled to ERK and ATM signaling.\",\n      \"evidence\": \"Interactome MS, Co-IP, RNAi, iron measurements (HERC2); Co-IP, ubiquitination, and phosphosite mutagenesis (cortactin, hSSB1)\",\n      \"pmids\": [\"24778179\", \"24867096\", \"25249620\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Each substrate characterized in a single lab\", \"How phosphorylation switches gate FBXL5 recruitment mechanistically unknown\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"FBXL5-mediated degradation of CITED2 connected the ligase to HIF-1α transcriptional output, broadening its role in oxygen-responsive gene regulation.\",\n      \"evidence\": \"Co-IP, RNAi, overexpression, FRET/BRET, and reporter assays\",\n      \"pmids\": [\"25956243\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab\", \"In vivo relevance of the FBXL5-CITED2-HIF axis not established here\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Tissue-level studies and CIA crosstalk established FBXL5 as essential for stem cell and progenitor iron control and linked it functionally to Fe-S cluster assembly through both IRP1 and IRP2.\",\n      \"evidence\": \"Conditional knockouts in HSCs and neural progenitors with IRP2-suppression rescue; siRNA, IRP1 phosphosite mutants, and polyubiquitination assays for CIA crosstalk\",\n      \"pmids\": [\"28714470\", \"28069738\", \"28768766\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"mTOR as downstream effector in NSPCs inferred pharmacologically\", \"Molecular coupling between CIA status and FBXL5 activity not fully defined\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Oxygen-dependent association of FBXL5 with the CIA-targeting complex (MMS19/FAM96B/CIAO1) provided a mechanism linking ambient oxygen to FBXL5-mediated IRP degradation.\",\n      \"evidence\": \"Co-IP, mass spectrometry, and IRP degradation assays under varied O2\",\n      \"pmids\": [\"31229404\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab\", \"Structural basis of the oxygen-dependent interaction not determined\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"A cryo-EM structure of the IRP2-FBXL5-SKP1 complex resolved the C-terminal [2Fe2S] cluster as the oxygen-sensing module that organizes the substrate-binding loop and dislodges IRP2 from IRE-RNA, completing the dual-sensor mechanism.\",\n      \"evidence\": \"Cryo-EM, in vitro ubiquitination, mutagenesis, EPR spectroscopy\",\n      \"pmids\": [\"32126207\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structure of the full SCF holocomplex on substrate not solved\", \"Integration of N-terminal diiron and C-terminal [2Fe2S] signals into one stability/activity output not structurally unified\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"A negative-control study showed IRP2 can be activated by Fe-S cluster deficiency at tissue O2 levels independently of FBXL5, delimiting the boundary of the FBXL5-dependent regulatory axis.\",\n      \"evidence\": \"ISC inhibition, IRP RNA-binding assays, IRP1/IRP2 double knockout, FBXL5 knockdown, ferroptosis assays\",\n      \"pmids\": [\"34039609\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"FBXL5-independent IRP2 activation mechanism not identified\", \"Physiological contexts where each pathway dominates unclear\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Translational and disease-context studies expanded FBXL5 regulation and substrate scope, showing G3BP1-mediated suppression of FBXL5 translation drives ferroptosis, and identifying TFEB and oocyte CITED2 as functionally relevant targets.\",\n      \"evidence\": \"35S labeling/RIP and in vivo kidney injury (G3BP1); Co-IP/ubiquitination/siRNA in HepG2 (TFEB); oocyte siRNA, ROS, spindle imaging, in vitro ubiquitination (CITED2)\",\n      \"pmids\": [\"38118197\", \"37743009\", \"37462473\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"TFEB and YAP1 substrate findings rest on single Co-IP/ubiquitination studies\", \"Whether these substrates depend on FBXL5's iron-sensing state is untested\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"FBXL5 was implicated in tumor suppression via Galectin-3-enhanced degradation of YAP1, extending its substrate repertoire to Hippo-pathway control.\",\n      \"evidence\": \"Co-IP, ubiquitination assay, xenograft model, proliferation assays\",\n      \"pmids\": [\"42093989\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single lab with limited mechanistic detail\", \"Degron and recruitment determinants for YAP1 unknown\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How the N-terminal diiron and C-terminal [2Fe2S] sensors are jointly integrated to set both FBXL5 stability and substrate selectivity across its many non-IRP targets remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified structural model coupling both metal centers to substrate choice\", \"Whether non-IRP substrates are regulated by iron/oxygen state is untested\", \"Tissue-specific substrate prioritization unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 2, 7, 8, 9, 10]},\n      {\"term_id\": \"GO:0016740\", \"supporting_discovery_ids\": [0, 2]},\n      {\"term_id\": \"GO:0140299\", \"supporting_discovery_ids\": [0, 1, 14]},\n      {\"term_id\": \"GO:0140313\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [7]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [11]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [2, 3, 12]},\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [0, 2, 4]},\n      {\"term_id\": \"R-HSA-8953897\", \"supporting_discovery_ids\": [0, 5]}\n    ],\n    \"complexes\": [\"SCF (SKP1-CUL1-RBX1-FBXL5) E3 ubiquitin ligase\"],\n    \"partners\": [\"IRP2\", \"IRP1\", \"SKP1\", \"HERC2\", \"Snail1\", \"CITED2\", \"hSSB1\", \"cortactin\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"tie","faith_supported":6,"faith_total":6,"faith_pct":100.0}}