{"gene":"FEM1B","run_date":"2026-06-09T23:54:43","timeline":{"discoveries":[{"year":2024,"finding":"CRL2FEM1B recognizes C-degrons containing a C-terminal proline via a bipartite mechanism that engages both the C-terminal proline and an upstream aromatic residue. Cryo-EM structures revealed the dimeric assembly of the complex and distinct dimerization states of unmodified vs. neddylated CRL2FEM1B, uncovering NEDD8-mediated activation. In vitro ubiquitination and cell-based assays confirmed that polyubiquitination and protein turnover depend on both FEM1B-degron interactions and the dimerization state.","method":"Cryo-EM structure determination, in vitro ubiquitination assay, cell-based degradation assays","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple cryo-EM structures combined with in vitro ubiquitination reconstitution and cell-based assays in one rigorous study","pmids":["38670995"],"is_preprint":false},{"year":2022,"finding":"A covalent ligand EN106 targets C186 of FEM1B and disrupts recognition of the reductive-stress substrate FNIP1. EN106 can be used as a FEM1B recruiter in PROTAC degraders (e.g., EN106-JQ1 degrades BRD4; EN106-dasatinib degrades BCR-ABL), establishing C186 as part of the natural E3 ligase–substrate binding site.","method":"Covalent ligand screening, PROTAC synthesis and cellular degradation assays, target engagement assays","journal":"Journal of the American Chemical Society","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — covalent active-site engagement with mutagenesis-level specificity (C186), reconstituted PROTAC degradation with multiple substrates, replicated across degrader pairs","pmids":["34994556"],"is_preprint":false},{"year":2021,"finding":"CRL2FEM1B recognizes the C-degron of the SMCR8 isoform (an autophagy regulator). Crystal/structural data of FEM1B bound to the SMCR8 C-degron provided mechanistic insight into substrate recognition and regulation of SMCR8 lifetime.","method":"Structural determination of FEM1B–SMCR8 C-degron complex, co-immunoprecipitation","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — structure presented from a single lab; limited functional validation reported in the abstract","pmids":["33892462"],"is_preprint":false},{"year":2026,"finding":"CRL2FEM1B recruits the transcription factor BACH1 for ubiquitin-mediated degradation by recognizing a degron directly formed by the redox-sensing molecule heme. By degrading BACH1 in response to heme, CRL2FEM1B modulates transcription of ferroptosis-protective genes (e.g., SLC7A11). Loss of CRL2FEM1B stabilizes BACH1 and sensitizes lung tumor cells to ferroptosis inducers.","method":"Co-immunoprecipitation, ubiquitination assays, in vitro and in vivo (preclinical) loss-of-function experiments","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal biochemical identification of heme-dependent substrate recruitment, multiple orthogonal assays, in vitro and preclinical validation","pmids":["42086045"],"is_preprint":false},{"year":2009,"finding":"FEM1B interacts with CHK1 (identified by yeast two-hybrid) and is required for chromatin loading of Rad9 and activation of CHK1 (Ser345 phosphorylation) in response to replication stress. FEM1B associates with chromatin (shown by fractionation), and FEM1B depletion by siRNA impairs ATR activity and sensitizes cells to replication block, phenocopying CHK1 loss.","method":"Yeast two-hybrid, siRNA knockdown, chromatin fractionation, CHK1 kinase activity assay, phosphorylation (immunoblot)","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — yeast two-hybrid plus functional siRNA experiments with multiple readouts; single lab","pmids":["19330022"],"is_preprint":false},{"year":2009,"finding":"RACK1 co-immunoprecipitates with endogenous FEM1B in metastatic colon cancer cells, stimulates FEM1B ubiquitination, and downregulates FEM1B protein levels. The N-terminal region of FEM1B is sufficient for RACK1 association. RACK1 knockdown elevates FEM1B and induces apoptosis, which is suppressed by blocking FEM1B upregulation.","method":"Co-immunoprecipitation, co-expression ubiquitination assay in HEK293T, siRNA knockdown, apoptosis assay","journal":"Cancer biology & therapy","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal co-IP and functional epistasis with rescue experiment; single lab","pmids":["19855191"],"is_preprint":false},{"year":2013,"finding":"Fem1b directly binds Gli1 (mammalian homolog of nematode TRA-1) via its VHL-box, promotes Gli1 ubiquitylation, suppresses Gli1 transcriptional activity, and attenuates an oncogenic Gli1 autoregulatory loop in cancer cells. These effects are dependent on the VHL-box motif of Fem1b.","method":"Co-immunoprecipitation, in-cell ubiquitylation assay, transcriptional reporter assay, domain mutagenesis (VHL-box)","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct binding confirmed with co-IP and GST pulldown-equivalent, plus functional VHL-box mutagenesis and ubiquitylation assay; single lab","pmids":["24076122"],"is_preprint":false},{"year":2011,"finding":"Mouse Fem1b binds Ankrd37 (identified by yeast two-hybrid and co-IP), facilitates nuclear translocation of Fem1b in co-transfected cells, and targets Ankrd37 for ubiquitin-mediated degradation in a Fem1b dose-dependent manner.","method":"Yeast two-hybrid, co-immunoprecipitation, transfection-based degradation assay, ubiquitination assay","journal":"Gene","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — yeast two-hybrid plus co-IP and dose-dependent degradation; single lab; note: an earlier companion paper (PMID:19171189) was withdrawn, but PMID:21723927 is distinct and peer-reviewed","pmids":["21723927"],"is_preprint":false},{"year":2010,"finding":"Fem1b protein is downregulated by the proteasome in malignant colon cancer cells; proteasome inhibitor treatment upregulates Fem1b protein, and antisense morpholino blockade of Fem1b upregulation suppresses the resulting apoptosis, demonstrating that Fem1b mediates proteasome inhibitor-induced apoptosis.","method":"Proteasome inhibitor treatment, morpholino antisense oligonucleotide, apoptosis assay, transfection over-expression","journal":"Molecular carcinogenesis","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — antisense rescue epistasis experiment with functional apoptosis readout; single lab, multiple cell lines","pmids":["19908242"],"is_preprint":false},{"year":2004,"finding":"FEM1B interacts with PHTF1 via the ANK domain of FEM1B (N-terminus of PHTF1 is the binding region), shown by yeast two-hybrid, co-immunoprecipitation, and co-expression. PHTF1 recruits FEM1B to the endoplasmic reticulum membrane.","method":"Yeast two-hybrid, co-immunoprecipitation, domain mapping, subcellular co-localization","journal":"Biology of reproduction","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — yeast two-hybrid plus co-IP with domain mapping; single lab","pmids":["15601915"],"is_preprint":false},{"year":2008,"finding":"Mouse Fem1b interacts with the homeodomain protein Nkx3.1 in GST pull-down and co-immunoprecipitation assays; both proteins are co-expressed in neonatal prostate and testis. Fem1b null mice (gene targeting) display defects in prostate ductal morphogenesis and secretory protein expression, similar to Nkx3.1 mutant phenotypes.","method":"GST pull-down, co-immunoprecipitation, gene-targeted knockout mouse, histology/developmental analysis","journal":"Developmental dynamics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — GST pull-down and co-IP plus KO mouse phenotype; single lab","pmids":["18816836"],"is_preprint":false},{"year":2023,"finding":"miR-29a-3p downregulates FEM1B expression, leading to stabilization of Gli1 (a FEM1B ubiquitination target); reduced FEM1B levels are sufficient to confer oxaliplatin resistance in colorectal cancer cells, and GLI1 knockdown reverses this resistance, placing FEM1B upstream of Gli1 in a chemoresistance pathway.","method":"miRNA overexpression, FEM1B knockdown, GLI1 knockdown, oxaliplatin resistance assay, in vivo mouse tumor model","journal":"American journal of cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis (FEM1B–Gli1 axis) with functional drug-resistance readout in vitro and in vivo; single lab","pmids":["38187042"],"is_preprint":false},{"year":2025,"finding":"FEM1B enhances TRAIL-induced apoptosis through the extrinsic caspase-dependent pathway (activating caspase-3 and caspase-8, but not caspase-9). In T lymphocyte lines, FEM1B co-immunoprecipitates with TRAF2 and downregulates TRAF2, diminishing TRAF2's inhibitory effect on caspase-8. In monocyte lines, FEM1B upregulates TRAIL-R2 to promote TRAIL-induced apoptosis.","method":"Co-immunoprecipitation, FEM1B knockdown/knockout, caspase activity assays, flow cytometry apoptosis assay, murine KO model","journal":"FEBS open bio","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP plus KO epistasis with specific caspase readouts and in vivo confirmation; single lab","pmids":["40392678"],"is_preprint":false},{"year":2025,"finding":"METTL3-mediated m6A modification reduces Fem1b mRNA stability in skeletal stem cells, causing lower FEM1B protein levels; reduced FEM1B in turn stabilizes Gli1, maintaining SSC quiescence. Genetic deletion of Mettl3 in murine SSCs elevates Fem1b and destabilizes Gli1, causing quiescence exit.","method":"Conditional Mettl3 knockout mouse, m6A sequencing, RNA stability assay, Gli1 protein level measurement","journal":"The EMBO journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis in vivo with mechanistic m6A profiling; single lab; Fem1b-Gli1 axis is consistent with prior studies","pmids":["40016417"],"is_preprint":false},{"year":2024,"finding":"Stop codon readthrough (SCR) of FEM1B generates a C-terminally extended, highly unstable isoform. The 81-nt proximal 3'UTR serves as the cis-signal for SCR and encodes the degron responsible for instability of the SCR product. CRISPR deletion of this region increases FEM1B expression, reduces SLBP (a FEM1B-mediated degradation target), reduces replication-dependent histones, and causes cell cycle delay.","method":"Stop codon readthrough reporter assays, CRISPR editing, protein stability assays, cell cycle analysis, SLBP expression measurement","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR functional epistasis plus SCR reporter and downstream substrate (SLBP) validation; single lab","pmids":["39140134"],"is_preprint":false},{"year":2005,"finding":"Mice with targeted inactivation of Fem1b (Fem1b-KO) display abnormal glucose tolerance due predominantly to defective glucose-stimulated and arginine-stimulated insulin secretion. Fem1b is expressed in pancreatic islet beta cells and non-beta cells and in INS-1E cells, implicating it in islet function.","method":"Gene-targeted knockout mouse, glucose tolerance test, insulin secretion assay (glucose- and arginine-stimulated)","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — clean KO mouse with specific physiological and cellular phenotype; single lab","pmids":["16024793"],"is_preprint":false},{"year":2024,"finding":"A de novo gain-of-function missense variant FEM1B p.Arg126Gln causes aberrant FEM1B activation. Overexpression of the Arg126Gln variant (but not wild-type) during mouse brain development (in utero electroporation) results in delayed neuronal migration. Patient cells exhibit oxidative stress and induction of type I interferon signaling.","method":"In utero electroporation in mouse brain, patient cell oxidative stress assay, interferon signaling measurement","journal":"Genetics in medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo functional variant assay (in utero electroporation) plus patient cell biochemistry; single cohort/lab","pmids":["38465576"],"is_preprint":false}],"current_model":"FEM1B functions as a substrate-recognition subunit of the CRL2 (Cullin 2-RING) E3 ubiquitin ligase complex, using a VHL-box motif and a bipartite degron-binding mechanism to recognize C-terminal degrons (including C-terminal proline motifs and heme-formed degrons) on substrates such as FNIP1, Gli1, BACH1, SMCR8, CDK5R1, SLBP, and Ankrd37, targeting them for polyubiquitination and proteasomal degradation; additionally, FEM1B participates in the DNA replication stress checkpoint by facilitating chromatin loading of Rad9 and CHK1 activation, promotes TRAIL-induced extrinsic apoptosis via TRAF2 downregulation and TRAIL-R2 upregulation, is itself regulated by RACK1-mediated ubiquitination and by m6A-dependent mRNA destabilization, and a gain-of-function variant (Arg126Gln) causes aberrant substrate degradation leading to a syndromic neurodevelopmental disorder."},"narrative":{"mechanistic_narrative":"FEM1B is the substrate-recognition subunit of a CRL2 (Cullin 2-RING) E3 ubiquitin ligase that selects substrates by binding C-terminal degrons and targets them for polyubiquitination and proteasomal degradation [PMID:38670995, PMID:34994556]. It recognizes C-degrons terminating in proline through a bipartite mechanism that engages both the C-terminal proline and an upstream aromatic residue, and cryo-EM structures show that the ligase assembles as a dimer whose neddylation state controls activation and substrate turnover [PMID:38670995]. Substrate recognition can be conditional: FEM1B reads a heme-formed degron on the transcription factor BACH1, coupling redox state to BACH1 degradation and thereby tuning expression of ferroptosis-protective genes such as SLC7A11 [PMID:42086045], and it engages the reductive-stress factor FNIP1 through a binding surface that includes Cys186 [PMID:34994556]. Through its VHL-box, FEM1B binds and ubiquitylates the transcription factor Gli1 to suppress an oncogenic Gli1 autoregulatory loop [PMID:24076122], an axis exploited in colorectal chemoresistance and in skeletal stem cell quiescence where reduced FEM1B stabilizes Gli1 [PMID:38187042, PMID:40016417]. Additional characterized substrates include the autophagy regulator SMCR8 [PMID:33892462], the histone mRNA factor SLBP [PMID:39140134], and Ankrd37 [PMID:21723927]. Beyond its ligase activity, FEM1B associates with chromatin and is required for Rad9 loading and CHK1 (Ser345) activation during replication stress [PMID:19330022], and it promotes TRAIL-induced extrinsic apoptosis by downregulating TRAF2 and upregulating TRAIL-R2 [PMID:40392678]. FEM1B abundance is itself controlled by RACK1-stimulated ubiquitination [PMID:19855191], by METTL3/m6A-dependent mRNA destabilization [PMID:40016417], and by a programmed stop-codon-readthrough event that produces an unstable extended isoform [PMID:39140134]. A de novo gain-of-function variant, p.Arg126Gln, causes aberrant FEM1B activation and a syndromic neurodevelopmental disorder, with delayed neuronal migration and patient-cell oxidative stress and type I interferon induction [PMID:38465576].","teleology":[{"year":2004,"claim":"Established the first physical partner and a subcellular context for FEM1B, showing its ANK domain binds PHTF1, which tethers FEM1B to the ER membrane.","evidence":"Yeast two-hybrid, co-IP, domain mapping and co-localization","pmids":["15601915"],"confidence":"Medium","gaps":["Did not define a catalytic/E3 function","No demonstration that PHTF1 is a degradation substrate"]},{"year":2005,"claim":"A Fem1b knockout linked the gene to a physiological process, revealing a requirement for glucose- and arginine-stimulated insulin secretion in islet beta cells.","evidence":"Gene-targeted KO mouse with glucose tolerance and insulin secretion assays","pmids":["16024793"],"confidence":"Medium","gaps":["Molecular mechanism connecting FEM1B to insulin secretion unresolved","No substrate implicated in the islet phenotype"]},{"year":2008,"claim":"Connected FEM1B to a developmental program by showing it binds Nkx3.1 and is required for prostate ductal morphogenesis.","evidence":"GST pull-down, co-IP, and KO mouse developmental analysis","pmids":["18816836"],"confidence":"Medium","gaps":["Whether Nkx3.1 is a ubiquitination substrate not tested","Mechanism of developmental defect unknown"]},{"year":2009,"claim":"Identified a checkpoint role distinct from ligase activity, showing FEM1B associates with chromatin and is needed for Rad9 loading and CHK1 activation under replication stress.","evidence":"Yeast two-hybrid, siRNA, chromatin fractionation, CHK1 kinase and phospho-immunoblot assays","pmids":["19330022"],"confidence":"Medium","gaps":["Direct biochemical role in Rad9 loading not reconstituted","Single lab","Relationship to E3 ligase function unclear"]},{"year":2009,"claim":"Revealed that FEM1B is itself a regulated target, with RACK1 stimulating FEM1B ubiquitination and controlling its pro-apoptotic levels in colon cancer.","evidence":"Reciprocal co-IP, co-expression ubiquitination assay, siRNA and apoptosis rescue","pmids":["19855191"],"confidence":"Medium","gaps":["E3 ligase mediating FEM1B ubiquitination not identified","Single lab"]},{"year":2010,"claim":"Demonstrated functionally that proteasome-controlled FEM1B levels drive proteasome-inhibitor-induced apoptosis in malignant colon cells.","evidence":"Proteasome inhibitor treatment, morpholino antisense rescue, apoptosis assay","pmids":["19908242"],"confidence":"Medium","gaps":["Downstream apoptotic effectors not defined","Mechanism of FEM1B-driven death unresolved"]},{"year":2011,"claim":"Provided early evidence of FEM1B acting as a degradation factor by identifying Ankrd37 as a dose-dependent ubiquitination target.","evidence":"Yeast two-hybrid, co-IP, transfection-based degradation and ubiquitination assays","pmids":["21723927"],"confidence":"Medium","gaps":["Degron on Ankrd37 not mapped","CRL2 context not yet established","Single lab"]},{"year":2013,"claim":"Defined a VHL-box-dependent substrate, showing FEM1B binds and ubiquitylates Gli1 to suppress an oncogenic Gli1 autoregulatory loop.","evidence":"Co-IP, in-cell ubiquitylation, reporter assay, VHL-box mutagenesis","pmids":["24076122"],"confidence":"Medium","gaps":["Full CRL2 reconstitution not shown","Degron recognition mechanism unresolved at this stage"]},{"year":2021,"claim":"Extended the substrate repertoire with structural insight into FEM1B reading the C-degron of the autophagy regulator SMCR8.","evidence":"Structural determination of FEM1B–SMCR8 C-degron complex and co-IP","pmids":["33892462"],"confidence":"Medium","gaps":["Limited functional validation","Cellular consequences of SMCR8 turnover not fully resolved","Single lab"]},{"year":2022,"claim":"Pinpointed Cys186 as part of the natural substrate-binding site and converted FEM1B into a ligand-addressable E3 for targeted protein degradation.","evidence":"Covalent ligand (EN106) screening, PROTAC synthesis (EN106-JQ1, EN106-dasatinib), target engagement and degradation assays","pmids":["34994556"],"confidence":"High","gaps":["Full structural basis of FNIP1 recognition not solved here","In vivo applicability of degraders untested"]},{"year":2023,"claim":"Placed FEM1B upstream of Gli1 in a clinically relevant axis, showing miR-29a-3p-mediated FEM1B loss stabilizes Gli1 and confers oxaliplatin resistance.","evidence":"miRNA overexpression, FEM1B and GLI1 knockdown, oxaliplatin resistance assay, mouse tumor model","pmids":["38187042"],"confidence":"Medium","gaps":["Direct miR-29a-3p–FEM1B binding not detailed","Single lab"]},{"year":2024,"claim":"Solved the mechanistic basis of FEM1B substrate selection, revealing bipartite C-terminal proline degron recognition and NEDD8-controlled dimeric activation of CRL2FEM1B.","evidence":"Cryo-EM structures, in vitro ubiquitination reconstitution, cell-based degradation assays","pmids":["38670995"],"confidence":"High","gaps":["Generality of bipartite recognition across all substrates not exhaustively mapped","Dynamics of dimerization in cells not directly observed"]},{"year":2024,"claim":"Showed FEM1B abundance is set by a programmed stop-codon-readthrough degron and identified SLBP as a substrate controlling histone supply and cell cycle progression.","evidence":"SCR reporter assays, CRISPR deletion, protein stability and cell cycle analysis, SLBP measurement","pmids":["39140134"],"confidence":"Medium","gaps":["Trans-factors driving FEM1B readthrough unknown","Single lab"]},{"year":2024,"claim":"Linked FEM1B to human disease, demonstrating that the gain-of-function p.Arg126Gln variant drives aberrant activation, delayed neuronal migration, and a neurodevelopmental phenotype.","evidence":"In utero electroporation in mouse brain, patient cell oxidative stress and interferon signaling assays","pmids":["38465576"],"confidence":"Medium","gaps":["Specific dysregulated substrate driving the neurodevelopmental phenotype not identified","Single cohort/lab"]},{"year":2025,"claim":"Defined a ligase-independent and a ligase-related role in extrinsic apoptosis, with FEM1B downregulating TRAF2 and upregulating TRAIL-R2 to enhance TRAIL-induced caspase-8/3 activation.","evidence":"Co-IP, knockdown/knockout, caspase activity and flow-cytometry apoptosis assays, murine KO","pmids":["40392678"],"confidence":"Medium","gaps":["Whether TRAF2 is a direct ubiquitination substrate not established","Single lab"]},{"year":2025,"claim":"Connected FEM1B regulation to stem cell biology, showing METTL3/m6A destabilizes Fem1b mRNA to stabilize Gli1 and maintain skeletal stem cell quiescence.","evidence":"Conditional Mettl3 KO mouse, m6A sequencing, RNA stability and Gli1 protein assays","pmids":["40016417"],"confidence":"Medium","gaps":["Direct m6A reader controlling Fem1b decay not specified","Single lab"]},{"year":2026,"claim":"Revealed conditional, cofactor-formed degron recognition, showing CRL2FEM1B reads a heme-formed degron on BACH1 to couple redox state to ferroptosis-gene regulation.","evidence":"Co-IP, ubiquitination assays, in vitro and preclinical loss-of-function experiments","pmids":["42086045"],"confidence":"High","gaps":["Structural basis of heme-degron recognition not resolved","Breadth of heme-formed substrates unknown"]},{"year":null,"claim":"How FEM1B's ligase-independent functions (replication checkpoint, apoptosis) mechanistically relate to its CRL2 substrate-degradation role, and which substrate drives the neurodevelopmental disorder, remain open.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unifying mechanism linking E3 activity to chromatin/checkpoint function","Disease-driving substrate unidentified","In vivo substrate hierarchy not established"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,1,3,6]},{"term_id":"GO:0016874","term_label":"ligase activity","supporting_discovery_ids":[0,6]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,6,2]},{"term_id":"GO:0140299","term_label":"molecular sensor activity","supporting_discovery_ids":[3]}],"localization":[{"term_id":"GO:0005783","term_label":"endoplasmic reticulum","supporting_discovery_ids":[9]},{"term_id":"GO:0005694","term_label":"chromosome","supporting_discovery_ids":[4]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[7]}],"pathway":[{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[0,1,3,6]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[12,8]},{"term_id":"R-HSA-73894","term_label":"DNA Repair","supporting_discovery_ids":[4]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[3,6]},{"term_id":"R-HSA-8953897","term_label":"Cellular responses to stimuli","supporting_discovery_ids":[3,1]}],"complexes":["CRL2 (Cullin 2-RING) E3 ubiquitin ligase"],"partners":["FNIP1","BACH1","GLI1","SMCR8","SLBP","ANKRD37","RACK1","TRAF2"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9UK73","full_name":"Protein fem-1 homolog B","aliases":["FEM1-beta","Fem-1-like death receptor-binding protein alpha","Fem-1-like in apoptotic pathway protein alpha","F1A-alpha"],"length_aa":627,"mass_kda":70.3,"function":"Substrate-recognition component of a Cul2-RING (CRL2) E3 ubiquitin-protein ligase complex of the DesCEND (destruction via C-end degrons) pathway, which recognizes a C-degron located at the extreme C terminus of target proteins, leading to their ubiquitination and degradation (PubMed:29779948, PubMed:33398168, PubMed:33398170). The C-degron recognized by the DesCEND pathway is usually a motif of less than ten residues and can be present in full-length proteins, truncated proteins or proteolytically cleaved forms (PubMed:29779948, PubMed:33398168, PubMed:33398170). The CRL2(FEM1B) complex specifically recognizes proteins ending with -Gly-Leu-Asp-Arg, such as CDK5R1, leading to their ubiquitination and degradation (PubMed:33398168, PubMed:33398170). Also acts as a regulator of the reductive stress response by mediating ubiquitination of reduced FNIP1: in response to reductive stress, the CRL2(FEM1B) complex specifically recognizes a conserved Cys degron in FNIP1 when this degron is reduced, leading to FNIP1 degradation and subsequent activation of mitochondria to recalibrate reactive oxygen species (ROS) (By similarity). Mechanistically, recognizes and binds reduced FNIP1 through two interface zinc ions, which act as a molecular glue that recruit reduced FNIP1 to FEM1B (By similarity). Promotes ubiquitination of GLI1, suppressing GLI1 transcriptional activator activity (PubMed:24076122). Promotes ubiquitination and degradation of ANKRD37 (By similarity). Promotes ubiquitination and degradation of SLBP (PubMed:28118078). Involved in apoptosis by acting as a death receptor-associated protein that mediates apoptosis (PubMed:10542291). Also involved in glucose homeostasis in pancreatic islet (By similarity). May also act as an adapter/mediator in replication stress-induced signaling that leads to the activation of CHEK1 (PubMed:19330022)","subcellular_location":"Cytoplasm; Nucleus","url":"https://www.uniprot.org/uniprotkb/Q9UK73/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/FEM1B","classification":"Not Classified","n_dependent_lines":30,"n_total_lines":1208,"dependency_fraction":0.024834437086092714},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/FEM1B","total_profiled":1310},"omim":[{"mim_id":"621263","title":"NEURODEVELOPMENTAL DISORDER WITH BEHAVIORAL, EAR, AND SKELETAL ABNORMALITIES; NEDBES","url":"https://www.omim.org/entry/621263"},{"mim_id":"613539","title":"FEM1 HOMOLOG B; FEM1B","url":"https://www.omim.org/entry/613539"},{"mim_id":"608767","title":"FEM1 HOMOLOG C; FEM1C","url":"https://www.omim.org/entry/608767"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nucleoplasm","reliability":"Supported"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/FEM1B"},"hgnc":{"alias_symbol":[],"prev_symbol":[]},"alphafold":{"accession":"Q9UK73","domains":[{"cath_id":"1.25.40.20","chopping":"1-152","consensus_level":"medium","plddt":96.0805,"start":1,"end":152},{"cath_id":"-","chopping":"380-507","consensus_level":"medium","plddt":95.3089,"start":380,"end":507}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9UK73","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9UK73-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9UK73-F1-predicted_aligned_error_v6.png","plddt_mean":94.44},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=FEM1B","jax_strain_url":"https://www.jax.org/strain/search?query=FEM1B"},"sequence":{"accession":"Q9UK73","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9UK73.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9UK73/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9UK73"}},"corpus_meta":[{"pmid":"34994556","id":"PMC_34994556","title":"Discovery of a Covalent FEM1B Recruiter for Targeted Protein Degradation Applications.","date":"2022","source":"Journal of the American Chemical Society","url":"https://pubmed.ncbi.nlm.nih.gov/34994556","citation_count":166,"is_preprint":false},{"pmid":"37521275","id":"PMC_37521275","title":"Glucose-responsive, antioxidative HA-PBA-FA/EN106 hydrogel enhanced diabetic wound healing through modulation of FEM1b-FNIP1 axis and promoting angiogenesis.","date":"2023","source":"Bioactive materials","url":"https://pubmed.ncbi.nlm.nih.gov/37521275","citation_count":59,"is_preprint":false},{"pmid":"19855191","id":"PMC_19855191","title":"RACK1 downregulates levels of the pro-apoptotic protein Fem1b in apoptosis-resistant colon cancer cells.","date":"2009","source":"Cancer biology & therapy","url":"https://pubmed.ncbi.nlm.nih.gov/19855191","citation_count":28,"is_preprint":false},{"pmid":"16024793","id":"PMC_16024793","title":"Abnormal glucose homeostasis and pancreatic islet function in mice with inactivation of the Fem1b gene.","date":"2005","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/16024793","citation_count":25,"is_preprint":false},{"pmid":"10623617","id":"PMC_10623617","title":"Sequence, organization, and expression of the human FEM1B gene.","date":"2000","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/10623617","citation_count":22,"is_preprint":false},{"pmid":"18757445","id":"PMC_18757445","title":"FEM1A and FEM1B: novel candidate genes for polycystic ovary syndrome.","date":"2008","source":"Human reproduction (Oxford, England)","url":"https://pubmed.ncbi.nlm.nih.gov/18757445","citation_count":21,"is_preprint":false},{"pmid":"21723927","id":"PMC_21723927","title":"Mouse Fem1b interacts with and induces ubiquitin-mediated degradation of Ankrd37.","date":"2011","source":"Gene","url":"https://pubmed.ncbi.nlm.nih.gov/21723927","citation_count":20,"is_preprint":false},{"pmid":"19908242","id":"PMC_19908242","title":"Fem1b, a proapoptotic protein, mediates proteasome inhibitor-induced apoptosis of human colon cancer cells.","date":"2010","source":"Molecular carcinogenesis","url":"https://pubmed.ncbi.nlm.nih.gov/19908242","citation_count":20,"is_preprint":false},{"pmid":"38670995","id":"PMC_38670995","title":"Mechanism of Ψ-Pro/C-degron recognition by the CRL2FEM1B ubiquitin ligase.","date":"2024","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/38670995","citation_count":19,"is_preprint":false},{"pmid":"24076122","id":"PMC_24076122","title":"Fem1b promotes ubiquitylation and suppresses transcriptional activity of Gli1.","date":"2013","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/24076122","citation_count":18,"is_preprint":false},{"pmid":"33892462","id":"PMC_33892462","title":"Structural insights into SMCR8 C-degron recognition by FEM1B.","date":"2021","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/33892462","citation_count":14,"is_preprint":false},{"pmid":"39804678","id":"PMC_39804678","title":"Development of the First-in-Class FEM1B-Recruiting Histone Deacetylase Degraders.","date":"2025","source":"Journal of medicinal chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/39804678","citation_count":10,"is_preprint":false},{"pmid":"15601915","id":"PMC_15601915","title":"Putative homeodomain transcription factor 1 interacts with the feminization factor homolog fem1b in male germ cells.","date":"2004","source":"Biology of reproduction","url":"https://pubmed.ncbi.nlm.nih.gov/15601915","citation_count":10,"is_preprint":false},{"pmid":"18816836","id":"PMC_18816836","title":"Mouse Fem1b interacts with the Nkx3.1 homeoprotein and is required for proper male secondary sexual development.","date":"2008","source":"Developmental dynamics : an official publication of the American Association of Anatomists","url":"https://pubmed.ncbi.nlm.nih.gov/18816836","citation_count":9,"is_preprint":false},{"pmid":"38187042","id":"PMC_38187042","title":"Induction of resistance to oxaliplatin in cancer by a microRNA/Fem1B/Gli1 pathway.","date":"2023","source":"American journal of cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/38187042","citation_count":7,"is_preprint":false},{"pmid":"27323097","id":"PMC_27323097","title":"Molecular cloning and expression analysis of Fem1b from oriental river prawn Macrobrachium nipponense.","date":"2016","source":"Genetics and molecular research : GMR","url":"https://pubmed.ncbi.nlm.nih.gov/27323097","citation_count":6,"is_preprint":false},{"pmid":"19330022","id":"PMC_19330022","title":"Human FEM1B is required for Rad9 recruitment and CHK1 activation in response to replication stress.","date":"2009","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/19330022","citation_count":6,"is_preprint":false},{"pmid":"38465576","id":"PMC_38465576","title":"A recurrent missense variant in the E3 ubiquitin ligase substrate recognition subunit FEM1B causes a rare syndromic neurodevelopmental disorder.","date":"2024","source":"Genetics in medicine : official journal of the American College of Medical Genetics","url":"https://pubmed.ncbi.nlm.nih.gov/38465576","citation_count":5,"is_preprint":false},{"pmid":"10768616","id":"PMC_10768616","title":"Rapid communication: the human FEM1B gene maps to chromosome 15q22 and is excluded as the gene for Bardet-Biedl syndrome, type 4.","date":"2000","source":"The American journal of the medical sciences","url":"https://pubmed.ncbi.nlm.nih.gov/10768616","citation_count":3,"is_preprint":false},{"pmid":"36003920","id":"PMC_36003920","title":"miR-29b Regulates Lung Cancer Progression by Downregulating FEM1B and Inhibiting the FOX01/AKT Pathway.","date":"2022","source":"Computational and mathematical methods in medicine","url":"https://pubmed.ncbi.nlm.nih.gov/36003920","citation_count":2,"is_preprint":false},{"pmid":"40016417","id":"PMC_40016417","title":"An epitranscriptomic program maintains skeletal stem cell quiescence via a METTL3-FEM1B-GLI1 axis.","date":"2025","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/40016417","citation_count":1,"is_preprint":false},{"pmid":"20952268","id":"PMC_20952268","title":"Fem1b antigen in the stool of ApcMin mice as a biomarker of early Wnt signaling activation in intestinal neoplasia.","date":"2010","source":"Cancer epidemiology","url":"https://pubmed.ncbi.nlm.nih.gov/20952268","citation_count":1,"is_preprint":false},{"pmid":"39140134","id":"PMC_39140134","title":"Hominini-specific regulation of the cell cycle by stop codon readthrough of FEM1B.","date":"2024","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/39140134","citation_count":0,"is_preprint":false},{"pmid":"42086045","id":"PMC_42086045","title":"CRL2FEM1B uses heme to recruit BACH1 for degradation and regulate ferroptosis in lung cancer.","date":"2026","source":"Molecular cell","url":"https://pubmed.ncbi.nlm.nih.gov/42086045","citation_count":0,"is_preprint":false},{"pmid":"40392678","id":"PMC_40392678","title":"FEM1B enhances TRAIL-induced apoptosis in T lymphocytes and monocytes.","date":"2025","source":"FEBS open bio","url":"https://pubmed.ncbi.nlm.nih.gov/40392678","citation_count":0,"is_preprint":false},{"pmid":"37829534","id":"PMC_37829534","title":"Retracted: miR-29b Regulates Lung Cancer Progression by Downregulating FEM1B and Inhibiting the FOX01/AKT Pathway.","date":"2023","source":"Computational and mathematical methods in medicine","url":"https://pubmed.ncbi.nlm.nih.gov/37829534","citation_count":0,"is_preprint":false},{"pmid":"19171189","id":"PMC_19171189","title":"WITHDRAWN: Fem1b interacts with Ankrd37 in mouse testis and induces its degradation by ubiquitin-mediated proteolysis pathway.","date":"2009","source":"Cellular signalling","url":"https://pubmed.ncbi.nlm.nih.gov/19171189","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":13732,"output_tokens":4362,"usd":0.053313,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":12350,"output_tokens":5022,"usd":0.09365,"stage2_stop_reason":"end_turn"},"total_usd":0.146963,"stage1_batch_id":"msgbatch_01AsR7g8dGTEiEw9U83bJeWn","stage2_batch_id":"msgbatch_01YV5ZyxtFNUZEjxRcpd9jzb","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2024,\n      \"finding\": \"CRL2FEM1B recognizes C-degrons containing a C-terminal proline via a bipartite mechanism that engages both the C-terminal proline and an upstream aromatic residue. Cryo-EM structures revealed the dimeric assembly of the complex and distinct dimerization states of unmodified vs. neddylated CRL2FEM1B, uncovering NEDD8-mediated activation. In vitro ubiquitination and cell-based assays confirmed that polyubiquitination and protein turnover depend on both FEM1B-degron interactions and the dimerization state.\",\n      \"method\": \"Cryo-EM structure determination, in vitro ubiquitination assay, cell-based degradation assays\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple cryo-EM structures combined with in vitro ubiquitination reconstitution and cell-based assays in one rigorous study\",\n      \"pmids\": [\"38670995\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"A covalent ligand EN106 targets C186 of FEM1B and disrupts recognition of the reductive-stress substrate FNIP1. EN106 can be used as a FEM1B recruiter in PROTAC degraders (e.g., EN106-JQ1 degrades BRD4; EN106-dasatinib degrades BCR-ABL), establishing C186 as part of the natural E3 ligase–substrate binding site.\",\n      \"method\": \"Covalent ligand screening, PROTAC synthesis and cellular degradation assays, target engagement assays\",\n      \"journal\": \"Journal of the American Chemical Society\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — covalent active-site engagement with mutagenesis-level specificity (C186), reconstituted PROTAC degradation with multiple substrates, replicated across degrader pairs\",\n      \"pmids\": [\"34994556\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"CRL2FEM1B recognizes the C-degron of the SMCR8 isoform (an autophagy regulator). Crystal/structural data of FEM1B bound to the SMCR8 C-degron provided mechanistic insight into substrate recognition and regulation of SMCR8 lifetime.\",\n      \"method\": \"Structural determination of FEM1B–SMCR8 C-degron complex, co-immunoprecipitation\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — structure presented from a single lab; limited functional validation reported in the abstract\",\n      \"pmids\": [\"33892462\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"CRL2FEM1B recruits the transcription factor BACH1 for ubiquitin-mediated degradation by recognizing a degron directly formed by the redox-sensing molecule heme. By degrading BACH1 in response to heme, CRL2FEM1B modulates transcription of ferroptosis-protective genes (e.g., SLC7A11). Loss of CRL2FEM1B stabilizes BACH1 and sensitizes lung tumor cells to ferroptosis inducers.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assays, in vitro and in vivo (preclinical) loss-of-function experiments\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal biochemical identification of heme-dependent substrate recruitment, multiple orthogonal assays, in vitro and preclinical validation\",\n      \"pmids\": [\"42086045\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"FEM1B interacts with CHK1 (identified by yeast two-hybrid) and is required for chromatin loading of Rad9 and activation of CHK1 (Ser345 phosphorylation) in response to replication stress. FEM1B associates with chromatin (shown by fractionation), and FEM1B depletion by siRNA impairs ATR activity and sensitizes cells to replication block, phenocopying CHK1 loss.\",\n      \"method\": \"Yeast two-hybrid, siRNA knockdown, chromatin fractionation, CHK1 kinase activity assay, phosphorylation (immunoblot)\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — yeast two-hybrid plus functional siRNA experiments with multiple readouts; single lab\",\n      \"pmids\": [\"19330022\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"RACK1 co-immunoprecipitates with endogenous FEM1B in metastatic colon cancer cells, stimulates FEM1B ubiquitination, and downregulates FEM1B protein levels. The N-terminal region of FEM1B is sufficient for RACK1 association. RACK1 knockdown elevates FEM1B and induces apoptosis, which is suppressed by blocking FEM1B upregulation.\",\n      \"method\": \"Co-immunoprecipitation, co-expression ubiquitination assay in HEK293T, siRNA knockdown, apoptosis assay\",\n      \"journal\": \"Cancer biology & therapy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal co-IP and functional epistasis with rescue experiment; single lab\",\n      \"pmids\": [\"19855191\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Fem1b directly binds Gli1 (mammalian homolog of nematode TRA-1) via its VHL-box, promotes Gli1 ubiquitylation, suppresses Gli1 transcriptional activity, and attenuates an oncogenic Gli1 autoregulatory loop in cancer cells. These effects are dependent on the VHL-box motif of Fem1b.\",\n      \"method\": \"Co-immunoprecipitation, in-cell ubiquitylation assay, transcriptional reporter assay, domain mutagenesis (VHL-box)\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct binding confirmed with co-IP and GST pulldown-equivalent, plus functional VHL-box mutagenesis and ubiquitylation assay; single lab\",\n      \"pmids\": [\"24076122\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Mouse Fem1b binds Ankrd37 (identified by yeast two-hybrid and co-IP), facilitates nuclear translocation of Fem1b in co-transfected cells, and targets Ankrd37 for ubiquitin-mediated degradation in a Fem1b dose-dependent manner.\",\n      \"method\": \"Yeast two-hybrid, co-immunoprecipitation, transfection-based degradation assay, ubiquitination assay\",\n      \"journal\": \"Gene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — yeast two-hybrid plus co-IP and dose-dependent degradation; single lab; note: an earlier companion paper (PMID:19171189) was withdrawn, but PMID:21723927 is distinct and peer-reviewed\",\n      \"pmids\": [\"21723927\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Fem1b protein is downregulated by the proteasome in malignant colon cancer cells; proteasome inhibitor treatment upregulates Fem1b protein, and antisense morpholino blockade of Fem1b upregulation suppresses the resulting apoptosis, demonstrating that Fem1b mediates proteasome inhibitor-induced apoptosis.\",\n      \"method\": \"Proteasome inhibitor treatment, morpholino antisense oligonucleotide, apoptosis assay, transfection over-expression\",\n      \"journal\": \"Molecular carcinogenesis\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — antisense rescue epistasis experiment with functional apoptosis readout; single lab, multiple cell lines\",\n      \"pmids\": [\"19908242\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"FEM1B interacts with PHTF1 via the ANK domain of FEM1B (N-terminus of PHTF1 is the binding region), shown by yeast two-hybrid, co-immunoprecipitation, and co-expression. PHTF1 recruits FEM1B to the endoplasmic reticulum membrane.\",\n      \"method\": \"Yeast two-hybrid, co-immunoprecipitation, domain mapping, subcellular co-localization\",\n      \"journal\": \"Biology of reproduction\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — yeast two-hybrid plus co-IP with domain mapping; single lab\",\n      \"pmids\": [\"15601915\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Mouse Fem1b interacts with the homeodomain protein Nkx3.1 in GST pull-down and co-immunoprecipitation assays; both proteins are co-expressed in neonatal prostate and testis. Fem1b null mice (gene targeting) display defects in prostate ductal morphogenesis and secretory protein expression, similar to Nkx3.1 mutant phenotypes.\",\n      \"method\": \"GST pull-down, co-immunoprecipitation, gene-targeted knockout mouse, histology/developmental analysis\",\n      \"journal\": \"Developmental dynamics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — GST pull-down and co-IP plus KO mouse phenotype; single lab\",\n      \"pmids\": [\"18816836\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"miR-29a-3p downregulates FEM1B expression, leading to stabilization of Gli1 (a FEM1B ubiquitination target); reduced FEM1B levels are sufficient to confer oxaliplatin resistance in colorectal cancer cells, and GLI1 knockdown reverses this resistance, placing FEM1B upstream of Gli1 in a chemoresistance pathway.\",\n      \"method\": \"miRNA overexpression, FEM1B knockdown, GLI1 knockdown, oxaliplatin resistance assay, in vivo mouse tumor model\",\n      \"journal\": \"American journal of cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis (FEM1B–Gli1 axis) with functional drug-resistance readout in vitro and in vivo; single lab\",\n      \"pmids\": [\"38187042\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"FEM1B enhances TRAIL-induced apoptosis through the extrinsic caspase-dependent pathway (activating caspase-3 and caspase-8, but not caspase-9). In T lymphocyte lines, FEM1B co-immunoprecipitates with TRAF2 and downregulates TRAF2, diminishing TRAF2's inhibitory effect on caspase-8. In monocyte lines, FEM1B upregulates TRAIL-R2 to promote TRAIL-induced apoptosis.\",\n      \"method\": \"Co-immunoprecipitation, FEM1B knockdown/knockout, caspase activity assays, flow cytometry apoptosis assay, murine KO model\",\n      \"journal\": \"FEBS open bio\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP plus KO epistasis with specific caspase readouts and in vivo confirmation; single lab\",\n      \"pmids\": [\"40392678\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"METTL3-mediated m6A modification reduces Fem1b mRNA stability in skeletal stem cells, causing lower FEM1B protein levels; reduced FEM1B in turn stabilizes Gli1, maintaining SSC quiescence. Genetic deletion of Mettl3 in murine SSCs elevates Fem1b and destabilizes Gli1, causing quiescence exit.\",\n      \"method\": \"Conditional Mettl3 knockout mouse, m6A sequencing, RNA stability assay, Gli1 protein level measurement\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis in vivo with mechanistic m6A profiling; single lab; Fem1b-Gli1 axis is consistent with prior studies\",\n      \"pmids\": [\"40016417\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Stop codon readthrough (SCR) of FEM1B generates a C-terminally extended, highly unstable isoform. The 81-nt proximal 3'UTR serves as the cis-signal for SCR and encodes the degron responsible for instability of the SCR product. CRISPR deletion of this region increases FEM1B expression, reduces SLBP (a FEM1B-mediated degradation target), reduces replication-dependent histones, and causes cell cycle delay.\",\n      \"method\": \"Stop codon readthrough reporter assays, CRISPR editing, protein stability assays, cell cycle analysis, SLBP expression measurement\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR functional epistasis plus SCR reporter and downstream substrate (SLBP) validation; single lab\",\n      \"pmids\": [\"39140134\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Mice with targeted inactivation of Fem1b (Fem1b-KO) display abnormal glucose tolerance due predominantly to defective glucose-stimulated and arginine-stimulated insulin secretion. Fem1b is expressed in pancreatic islet beta cells and non-beta cells and in INS-1E cells, implicating it in islet function.\",\n      \"method\": \"Gene-targeted knockout mouse, glucose tolerance test, insulin secretion assay (glucose- and arginine-stimulated)\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean KO mouse with specific physiological and cellular phenotype; single lab\",\n      \"pmids\": [\"16024793\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"A de novo gain-of-function missense variant FEM1B p.Arg126Gln causes aberrant FEM1B activation. Overexpression of the Arg126Gln variant (but not wild-type) during mouse brain development (in utero electroporation) results in delayed neuronal migration. Patient cells exhibit oxidative stress and induction of type I interferon signaling.\",\n      \"method\": \"In utero electroporation in mouse brain, patient cell oxidative stress assay, interferon signaling measurement\",\n      \"journal\": \"Genetics in medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo functional variant assay (in utero electroporation) plus patient cell biochemistry; single cohort/lab\",\n      \"pmids\": [\"38465576\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"FEM1B functions as a substrate-recognition subunit of the CRL2 (Cullin 2-RING) E3 ubiquitin ligase complex, using a VHL-box motif and a bipartite degron-binding mechanism to recognize C-terminal degrons (including C-terminal proline motifs and heme-formed degrons) on substrates such as FNIP1, Gli1, BACH1, SMCR8, CDK5R1, SLBP, and Ankrd37, targeting them for polyubiquitination and proteasomal degradation; additionally, FEM1B participates in the DNA replication stress checkpoint by facilitating chromatin loading of Rad9 and CHK1 activation, promotes TRAIL-induced extrinsic apoptosis via TRAF2 downregulation and TRAIL-R2 upregulation, is itself regulated by RACK1-mediated ubiquitination and by m6A-dependent mRNA destabilization, and a gain-of-function variant (Arg126Gln) causes aberrant substrate degradation leading to a syndromic neurodevelopmental disorder.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"FEM1B is the substrate-recognition subunit of a CRL2 (Cullin 2-RING) E3 ubiquitin ligase that selects substrates by binding C-terminal degrons and targets them for polyubiquitination and proteasomal degradation [#0, #1]. It recognizes C-degrons terminating in proline through a bipartite mechanism that engages both the C-terminal proline and an upstream aromatic residue, and cryo-EM structures show that the ligase assembles as a dimer whose neddylation state controls activation and substrate turnover [#0]. Substrate recognition can be conditional: FEM1B reads a heme-formed degron on the transcription factor BACH1, coupling redox state to BACH1 degradation and thereby tuning expression of ferroptosis-protective genes such as SLC7A11 [#3], and it engages the reductive-stress factor FNIP1 through a binding surface that includes Cys186 [#1]. Through its VHL-box, FEM1B binds and ubiquitylates the transcription factor Gli1 to suppress an oncogenic Gli1 autoregulatory loop [#6], an axis exploited in colorectal chemoresistance and in skeletal stem cell quiescence where reduced FEM1B stabilizes Gli1 [#11, #13]. Additional characterized substrates include the autophagy regulator SMCR8 [#2], the histone mRNA factor SLBP [#14], and Ankrd37 [#7]. Beyond its ligase activity, FEM1B associates with chromatin and is required for Rad9 loading and CHK1 (Ser345) activation during replication stress [#4], and it promotes TRAIL-induced extrinsic apoptosis by downregulating TRAF2 and upregulating TRAIL-R2 [#12]. FEM1B abundance is itself controlled by RACK1-stimulated ubiquitination [#5], by METTL3/m6A-dependent mRNA destabilization [#13], and by a programmed stop-codon-readthrough event that produces an unstable extended isoform [#14]. A de novo gain-of-function variant, p.Arg126Gln, causes aberrant FEM1B activation and a syndromic neurodevelopmental disorder, with delayed neuronal migration and patient-cell oxidative stress and type I interferon induction [#16].\",\n  \"teleology\": [\n    {\n      \"year\": 2004,\n      \"claim\": \"Established the first physical partner and a subcellular context for FEM1B, showing its ANK domain binds PHTF1, which tethers FEM1B to the ER membrane.\",\n      \"evidence\": \"Yeast two-hybrid, co-IP, domain mapping and co-localization\",\n      \"pmids\": [\"15601915\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Did not define a catalytic/E3 function\", \"No demonstration that PHTF1 is a degradation substrate\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"A Fem1b knockout linked the gene to a physiological process, revealing a requirement for glucose- and arginine-stimulated insulin secretion in islet beta cells.\",\n      \"evidence\": \"Gene-targeted KO mouse with glucose tolerance and insulin secretion assays\",\n      \"pmids\": [\"16024793\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular mechanism connecting FEM1B to insulin secretion unresolved\", \"No substrate implicated in the islet phenotype\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Connected FEM1B to a developmental program by showing it binds Nkx3.1 and is required for prostate ductal morphogenesis.\",\n      \"evidence\": \"GST pull-down, co-IP, and KO mouse developmental analysis\",\n      \"pmids\": [\"18816836\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether Nkx3.1 is a ubiquitination substrate not tested\", \"Mechanism of developmental defect unknown\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Identified a checkpoint role distinct from ligase activity, showing FEM1B associates with chromatin and is needed for Rad9 loading and CHK1 activation under replication stress.\",\n      \"evidence\": \"Yeast two-hybrid, siRNA, chromatin fractionation, CHK1 kinase and phospho-immunoblot assays\",\n      \"pmids\": [\"19330022\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct biochemical role in Rad9 loading not reconstituted\", \"Single lab\", \"Relationship to E3 ligase function unclear\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Revealed that FEM1B is itself a regulated target, with RACK1 stimulating FEM1B ubiquitination and controlling its pro-apoptotic levels in colon cancer.\",\n      \"evidence\": \"Reciprocal co-IP, co-expression ubiquitination assay, siRNA and apoptosis rescue\",\n      \"pmids\": [\"19855191\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"E3 ligase mediating FEM1B ubiquitination not identified\", \"Single lab\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Demonstrated functionally that proteasome-controlled FEM1B levels drive proteasome-inhibitor-induced apoptosis in malignant colon cells.\",\n      \"evidence\": \"Proteasome inhibitor treatment, morpholino antisense rescue, apoptosis assay\",\n      \"pmids\": [\"19908242\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Downstream apoptotic effectors not defined\", \"Mechanism of FEM1B-driven death unresolved\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Provided early evidence of FEM1B acting as a degradation factor by identifying Ankrd37 as a dose-dependent ubiquitination target.\",\n      \"evidence\": \"Yeast two-hybrid, co-IP, transfection-based degradation and ubiquitination assays\",\n      \"pmids\": [\"21723927\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Degron on Ankrd37 not mapped\", \"CRL2 context not yet established\", \"Single lab\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Defined a VHL-box-dependent substrate, showing FEM1B binds and ubiquitylates Gli1 to suppress an oncogenic Gli1 autoregulatory loop.\",\n      \"evidence\": \"Co-IP, in-cell ubiquitylation, reporter assay, VHL-box mutagenesis\",\n      \"pmids\": [\"24076122\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Full CRL2 reconstitution not shown\", \"Degron recognition mechanism unresolved at this stage\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Extended the substrate repertoire with structural insight into FEM1B reading the C-degron of the autophagy regulator SMCR8.\",\n      \"evidence\": \"Structural determination of FEM1B–SMCR8 C-degron complex and co-IP\",\n      \"pmids\": [\"33892462\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Limited functional validation\", \"Cellular consequences of SMCR8 turnover not fully resolved\", \"Single lab\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Pinpointed Cys186 as part of the natural substrate-binding site and converted FEM1B into a ligand-addressable E3 for targeted protein degradation.\",\n      \"evidence\": \"Covalent ligand (EN106) screening, PROTAC synthesis (EN106-JQ1, EN106-dasatinib), target engagement and degradation assays\",\n      \"pmids\": [\"34994556\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Full structural basis of FNIP1 recognition not solved here\", \"In vivo applicability of degraders untested\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Placed FEM1B upstream of Gli1 in a clinically relevant axis, showing miR-29a-3p-mediated FEM1B loss stabilizes Gli1 and confers oxaliplatin resistance.\",\n      \"evidence\": \"miRNA overexpression, FEM1B and GLI1 knockdown, oxaliplatin resistance assay, mouse tumor model\",\n      \"pmids\": [\"38187042\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct miR-29a-3p–FEM1B binding not detailed\", \"Single lab\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Solved the mechanistic basis of FEM1B substrate selection, revealing bipartite C-terminal proline degron recognition and NEDD8-controlled dimeric activation of CRL2FEM1B.\",\n      \"evidence\": \"Cryo-EM structures, in vitro ubiquitination reconstitution, cell-based degradation assays\",\n      \"pmids\": [\"38670995\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Generality of bipartite recognition across all substrates not exhaustively mapped\", \"Dynamics of dimerization in cells not directly observed\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Showed FEM1B abundance is set by a programmed stop-codon-readthrough degron and identified SLBP as a substrate controlling histone supply and cell cycle progression.\",\n      \"evidence\": \"SCR reporter assays, CRISPR deletion, protein stability and cell cycle analysis, SLBP measurement\",\n      \"pmids\": [\"39140134\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Trans-factors driving FEM1B readthrough unknown\", \"Single lab\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Linked FEM1B to human disease, demonstrating that the gain-of-function p.Arg126Gln variant drives aberrant activation, delayed neuronal migration, and a neurodevelopmental phenotype.\",\n      \"evidence\": \"In utero electroporation in mouse brain, patient cell oxidative stress and interferon signaling assays\",\n      \"pmids\": [\"38465576\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Specific dysregulated substrate driving the neurodevelopmental phenotype not identified\", \"Single cohort/lab\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Defined a ligase-independent and a ligase-related role in extrinsic apoptosis, with FEM1B downregulating TRAF2 and upregulating TRAIL-R2 to enhance TRAIL-induced caspase-8/3 activation.\",\n      \"evidence\": \"Co-IP, knockdown/knockout, caspase activity and flow-cytometry apoptosis assays, murine KO\",\n      \"pmids\": [\"40392678\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether TRAF2 is a direct ubiquitination substrate not established\", \"Single lab\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Connected FEM1B regulation to stem cell biology, showing METTL3/m6A destabilizes Fem1b mRNA to stabilize Gli1 and maintain skeletal stem cell quiescence.\",\n      \"evidence\": \"Conditional Mettl3 KO mouse, m6A sequencing, RNA stability and Gli1 protein assays\",\n      \"pmids\": [\"40016417\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct m6A reader controlling Fem1b decay not specified\", \"Single lab\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Revealed conditional, cofactor-formed degron recognition, showing CRL2FEM1B reads a heme-formed degron on BACH1 to couple redox state to ferroptosis-gene regulation.\",\n      \"evidence\": \"Co-IP, ubiquitination assays, in vitro and preclinical loss-of-function experiments\",\n      \"pmids\": [\"42086045\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of heme-degron recognition not resolved\", \"Breadth of heme-formed substrates unknown\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How FEM1B's ligase-independent functions (replication checkpoint, apoptosis) mechanistically relate to its CRL2 substrate-degradation role, and which substrate drives the neurodevelopmental disorder, remain open.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unifying mechanism linking E3 activity to chromatin/checkpoint function\", \"Disease-driving substrate unidentified\", \"In vivo substrate hierarchy not established\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 1, 3, 6]},\n      {\"term_id\": \"GO:0016874\", \"supporting_discovery_ids\": [0, 6]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0, 6, 2]},\n      {\"term_id\": \"GO:0140299\", \"supporting_discovery_ids\": [3]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005783\", \"supporting_discovery_ids\": [9]},\n      {\"term_id\": \"GO:0005694\", \"supporting_discovery_ids\": [4]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [7]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [0, 1, 3, 6]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [12, 8]},\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [4]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [3, 6]},\n      {\"term_id\": \"R-HSA-8953897\", \"supporting_discovery_ids\": [3, 1]}\n    ],\n    \"complexes\": [\"CRL2 (Cullin 2-RING) E3 ubiquitin ligase\"],\n    \"partners\": [\"FNIP1\", \"BACH1\", \"Gli1\", \"SMCR8\", \"SLBP\", \"Ankrd37\", \"RACK1\", \"TRAF2\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":8,"faith_total":8,"faith_pct":100.0}}