{"gene":"MAD2L1BP","run_date":"2026-06-10T02:59:50","timeline":{"discoveries":[{"year":2004,"finding":"p31comet binds selectively to the Cdc20-bound (closed) conformation of Mad2 but does not prevent the Mad2-Cdc20 interaction. Recombinant p31comet antagonizes Mad2-mediated inhibition of APC/C(Cdc20) in vitro and in Xenopus egg extracts. During checkpoint inactivation, p31comet forms a transient complex with APC/C(Cdc20)-bound Mad2 and enhances APC/C activity without disrupting Mad2-Cdc20 interaction. RNAi depletion of p31comet in HeLa cells delays mitotic exit after nocodazole removal.","method":"In vitro APC/C activity assay with recombinant protein, Xenopus egg extract assay, co-immunoprecipitation, RNAi in HeLa cells","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — in vitro reconstitution with recombinant proteins, Xenopus extract assay, and cell-based RNAi, multiple orthogonal methods in one study","pmids":["15257285"],"is_preprint":false},{"year":2006,"finding":"p31comet competes with O-Mad2 for binding to C-Mad2, thereby blocking the conformational dimerization of Mad2 that is required for spindle assembly checkpoint activation. NMR chemical shift perturbations showed that O-Mad2 undergoes major conformational rearrangement upon C-Mad2 binding, and p31comet inhibits this by occupying the C-Mad2 binding surface.","method":"NMR chemical shift perturbation, mutational analysis of Mad2 dimerization interface, SAC assay in Saccharomyces cerevisiae","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1 / Strong — NMR structural analysis combined with mutagenesis and in vivo checkpoint assay, multiple orthogonal methods","pmids":["16525508"],"is_preprint":false},{"year":2011,"finding":"p31comet promotes checkpoint silencing downstream of kinetochores by extracting Mad2 from the Mitotic Checkpoint Complex (MCC). p31comet binds Mad2 when it is within the MCC (bound to BubR1 and Cdc20). Recombinant p31comet added to checkpoint-arrested extracts removes Mad2 from the MCC; a p31comet mutant unable to bind Mad2 has no effect. RNAi of p31comet increases Mad2 bound to BubR1-Cdc20; overexpression decreases it.","method":"Co-immunoprecipitation, recombinant protein addition to checkpoint-arrested extracts, RNAi, overexpression, live-cell imaging","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal co-IP, extract-based reconstitution, mutant controls, RNAi and overexpression with multiple orthogonal readouts","pmids":["22100920"],"is_preprint":false},{"year":2011,"finding":"p31comet promotes disassembly of the MCC in an ATP-dependent manner; specifically, it promotes dissociation of Cdc20 from BubR1 within MCC. This process requires beta,gamma-hydrolyzable ATP. p31comet binds Mad2 yet causes release of Cdc20 from BubR1.","method":"Cell-free extracts from checkpoint-arrested HeLa cells, isolated MCC, ATP hydrolysis requirement assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — biochemical reconstitution with isolated MCC, ATP analog experiments, replicated concept in same lab and by other labs","pmids":["21300909"],"is_preprint":false},{"year":2011,"finding":"p31comet localizes to kinetochores in mitosis and traffics on and off kinetochores like Mad2. Cells depleted of p31comet arrest in metaphase with mature bipolar kinetochore-microtubule attachments and high cyclin B levels, demonstrating p31comet is required for timely mitotic exit even when the checkpoint is satisfied.","method":"Live-cell imaging, fluorescence microscopy, RNAi depletion in human and marsupial cells","journal":"Molecular biology of the cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct live-imaging of localization plus RNAi loss-of-function with specific mitotic phenotype, replicated across two cell systems","pmids":["21965286"],"is_preprint":false},{"year":2014,"finding":"TRIP13 AAA-ATPase works jointly with p31comet to disassemble MCC. TRIP13 promotes ATP- and p31comet-dependent disassembly of the Cdc20-Mad2 subcomplex and release of Mad2 from MCC, abrogating checkpoint inhibition of APC/C. TRIP13 was identified by fractionation of HeLa cell extracts as the factor responsible for this activity.","method":"Biochemical fractionation of HeLa cell extracts, in vitro disassembly assay, identification of TRIP13 by mass spectrometry, immunodepletion","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — biochemical fractionation, protein identification, in vitro reconstitution of disassembly with purified components","pmids":["25092294"],"is_preprint":false},{"year":2015,"finding":"Oligomeric TRIP13 binds both p31comet and MCC. p31comet and checkpoint complexes mutually promote each other's binding to oligomeric TRIP13. The substrate-binding site of TRIP13 has subsites specific for p31comet and C-Mad2-containing complex, and simultaneous occupancy of both subsites is required for high-affinity binding.","method":"Binding assays with purified recombinant proteins, co-immunoprecipitation, mutant analysis","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — purified component binding assays with multiple mutants, mechanistic model supported by multiple orthogonal binding experiments","pmids":["26324890"],"is_preprint":false},{"year":2014,"finding":"Phosphorylation of p31comet on Ser-102 weakens its binding to Mad2 and thereby enhances SAC activity (reduces p31comet-mediated SAC bypass). Attenuating p31comet affinity for Mad2 by phosphorylation at S102 promotes SAC activity during mitosis.","method":"Phosphorylation site mutagenesis (S102A), co-immunoprecipitation binding assay, SAC activity assay in cells","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — site-directed mutagenesis combined with binding assay and functional SAC readout, single lab","pmids":["24596092"],"is_preprint":false},{"year":2019,"finding":"Polo-like kinase 1 (Plk1) binds p31comet, phosphorylates it on S102, and suppresses its ability (together with TRIP13) to disassemble MCC. An S102A mutant of p31comet is largely resistant to Plk1-mediated inhibition. Plk1 inhibition in nocodazole-arrested HeLa cell extracts enhanced Mad2 release from checkpoint complexes.","method":"In vitro kinase assay with purified Plk1 and p31comet, co-immunoprecipitation, selective Plk1 inhibitor BI-2536 in cell extracts, S102A mutagenesis, MCC disassembly assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — purified kinase assay, site-directed mutagenesis, chemical inhibitor corroboration, functional MCC disassembly readout, multiple orthogonal methods","pmids":["31118282"],"is_preprint":false},{"year":2015,"finding":"IKK-β phosphorylates p31comet in Xenopus egg extracts during mitosis, and this phosphorylation enhances p31comet's role in SAC silencing to promote M-phase exit.","method":"Xenopus egg extract M-phase exit assay, kinase identification, phosphorylation analysis","journal":"Cell cycle (Georgetown, Tex.)","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — Xenopus egg extract functional assay with kinase identification, single lab, limited mechanistic detail in abstract","pmids":["25892037"],"is_preprint":false},{"year":2020,"finding":"p31comet binds the REV7-Shieldin complex in cells, promotes REV7 inactivation via TRIP13, and thereby promotes homologous recombination (HR) by enabling DNA end resection. p31comet also counteracts REV7 function in translesion synthesis (TLS) by releasing REV7 from REV3 in the Pol ζ complex. p31comet promotes extraction of REV7 from chromatin and causes PARP inhibitor resistance.","method":"Co-immunoprecipitation in cells, HR assay, chromatin fractionation, PARP inhibitor resistance assay, TRIP13 dependence assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP, functional HR assay, chromatin extraction assay in single lab with multiple orthogonal readouts","pmids":["33051298"],"is_preprint":false},{"year":2020,"finding":"p31comet-mediated suppression of MAD2 activation contributes to the timing of mitotic slippage. Reduction of kinetochore MAD2 during prolonged mitotic arrest is dependent on APC/C(CDC20), revealing an APC/C-to-SAC feedback loop. p31comet controls the rate of SAC weakening and thus the timing of cyclin B1 degradation and mitotic exit.","method":"RNAi depletion of p31comet, live-cell imaging, quantitative kinetochore MAD2 measurement, APC/C inhibitor experiments","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — RNAi with quantitative imaging and APC/C inhibitor experiments, single lab","pmids":["32029899"],"is_preprint":false},{"year":2009,"finding":"p31comet-induced cellular senescence requires direct interaction with Mad2; a p31comet mutant lacking the Mad2-binding region does not induce senescence. Senescence progression is dependent on p21(Waf1/Cip1) accumulation and is partly dependent on p53. Depletion of Mad2 by siRNA also induces senescence, phenocopying p31comet overexpression.","method":"Retroviral overexpression, domain deletion mutagenesis, siRNA knockdown, senescence assays (SA-β-gal, BrdU), flow cytometry","journal":"Molecular cancer research : MCR","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — functional mutagenesis of binding domain combined with loss-of-function siRNA and gain-of-function overexpression, single lab","pmids":["19276188"],"is_preprint":false},{"year":2012,"finding":"Depletion of p31comet increases mitotic checkpoint complex levels and prolongs mitotic block in response to spindle poisons. Cells with lower p31comet levels relative to MAD2 are less prone to mitotic slippage. The ratio of p31comet to MAD2 in cell lines correlates with susceptibility to mitotic slippage.","method":"RNAi depletion in multiple cancer cell lines, mitotic checkpoint complex quantification, cell death/slippage assays with spindle poisons","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — RNAi with MCC quantification and multiple functional readouts across multiple cell lines, single lab","pmids":["22544748"],"is_preprint":false},{"year":2015,"finding":"p31comet-induced cell death requires the C-terminal 30 amino acids of p31comet and is mediated by binding to and inactivating Mad2. Point mutation or deletion of the Mad2-binding region abolishes cytotoxic activity. Wild-type Mad2 (but not a non-p31comet-binding mutant) inhibits p31comet-induced cell death, confirming the mechanism involves Mad2 inactivation.","method":"Retroviral transduction, C-terminal deletion and point mutation analysis, colony formation assay, co-expression rescue experiments","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — domain mutagenesis with functional readout and rescue experiment, single lab","pmids":["26544187"],"is_preprint":false},{"year":2023,"finding":"Biallelic loss-of-function variants in MAD2L1BP causing C-terminal truncation of p31comet abolish its binding to MAD2. These variants cause female infertility due to oocyte metaphase I arrest. Microinjection of full-length MAD2L1BP cRNA rescued polar body extrusion in patient oocytes and mouse oocytes, while truncated variants could not.","method":"Whole-exome sequencing, co-immunoprecipitation binding assay, cRNA microinjection rescue in mouse and patient oocytes","journal":"eLife","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — patient genetics corroborated by binding assay and direct rescue experiment in human and mouse oocytes, multiple orthogonal methods","pmids":["37334967"],"is_preprint":false},{"year":2023,"finding":"A homozygous nonsense variant R253* in MAD2L1BP causes truncated p31comet that is defective in binding TRIP13, Mad2, and Rev7, and unable to support SAC silencing or homology-directed repair. An interaction between p31comet and TP53 was identified, and C-terminal truncation abrogated this interaction. Patient-derived cells showed high-grade mosaic variegated aneuploidy.","method":"Patient genetics, recombinant protein binding assays, SAC silencing assay, HDR assay, aneuploidy analysis of patient cells","journal":"JCI insight","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — recombinant protein binding assays and functional assays in patient-derived cells, single study with multiple readouts","pmids":["37796616"],"is_preprint":false},{"year":2025,"finding":"p31comet (MAD2L1BP) is modified by linear (Met1-linked) ubiquitination on its C-terminal lysine residues by the E3 ligase HOIP after cytokine stimulation. Ubiquitinated p31comet shows reduced binding to PLK1, which normally phosphorylates and inactivates p31comet. HOIP-deficient cells exhibit prolonged mitotic duration similar to p31comet knockout, and cells expressing a ubiquitination-deficient p31comet mutant take longer to complete M phase.","method":"Co-immunoprecipitation, ubiquitination assay, site-directed mutagenesis of C-terminal lysines, HOIP knockout cells, live-cell imaging of mitotic duration","journal":"Cell & bioscience","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP, mutagenesis, KO cell functional readout, single lab with multiple orthogonal methods","pmids":["40462232"],"is_preprint":false},{"year":2025,"finding":"p31comet Variant 1 (containing 32 additional N-terminal residues compared to Variant 2) exhibits reduced ability to bind MAD2 and reduced ability to induce mitotic progression. Variant 1 also shows reduced protein stability compared to Variant 2. Variant 1 is uniquely expressed in the testes.","method":"Splice variant cloning and expression, co-immunoprecipitation binding assay, mitotic progression assay, tissue expression analysis","journal":"International journal of molecular sciences","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — binding and functional assays comparing splice variants, single lab","pmids":["40243841"],"is_preprint":false},{"year":2013,"finding":"The Rb-E2F pathway regulates p31comet expression. A narrow range of p31comet:Mad2 ratios is compatible with cellular viability, and coordinate regulation of this antagonist-target pair is important for cell proliferation and checkpoint function.","method":"E2F reporter assay, expression analysis, cell viability assay with altered p31comet:Mad2 ratios","journal":"Cell cycle (Georgetown, Tex.)","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, expression-level regulation, limited mechanistic detail in abstract","pmids":["24131926"],"is_preprint":false}],"current_model":"MAD2L1BP (p31comet) is a Mad2-binding protein that silences the spindle assembly checkpoint (SAC) by two mechanisms: (1) competing with open-conformation O-Mad2 for binding to kinetochore-associated C-Mad2 to prevent new MCC assembly, and (2) binding C-Mad2 within the assembled MCC and recruiting the AAA-ATPase TRIP13, which uses ATP hydrolysis to extract Mad2 from the MCC, thereby releasing APC/C(Cdc20) from inhibition and allowing mitotic exit; p31comet activity is regulated by phosphorylation—PLK1 phosphorylates p31comet on S102 to suppress its MCC-disassembly activity during active checkpoint, while HOIP-mediated linear ubiquitination of p31comet reduces PLK1 binding and thus promotes p31comet activity; beyond mitosis, p31comet/TRIP13 also remodel REV7 complexes to promote homologous recombination and counteract translesion synthesis."},"narrative":{"mechanistic_narrative":"MAD2L1BP (p31comet) is a Mad2-binding adaptor that silences the spindle assembly checkpoint (SAC) to permit timely mitotic exit [PMID:15257285, PMID:21965286]. It acts at two points: it competes with open-conformation O-Mad2 for the dimerization surface of closed C-Mad2, blocking the Mad2 conformational dimerization required for checkpoint activation [PMID:16525508], and it binds C-Mad2 within the assembled Mitotic Checkpoint Complex (MCC), where it promotes ATP-dependent extraction of Mad2 and dissociation of Cdc20 from BubR1, releasing APC/C(Cdc20) from inhibition [PMID:22100920, PMID:21300909]. This disassembly is executed jointly with the AAA-ATPase TRIP13: p31comet and C-Mad2-containing checkpoint complexes occupy distinct subsites of oligomeric TRIP13 and mutually promote each other's high-affinity binding, coupling substrate recognition to ATP hydrolysis [PMID:25092294, PMID:26324890]. p31comet activity is set by post-translational control—PLK1 binds p31comet and phosphorylates it on Ser-102 to weaken Mad2 binding and suppress MCC disassembly during an active checkpoint, while HOIP-mediated linear (Met1-linked) ubiquitination of its C-terminal lysines reduces PLK1 binding and thereby promotes p31comet activity [PMID:24596092, PMID:31118282, PMID:40462232]. Beyond mitosis, p31comet partners with TRIP13 to remodel REV7 complexes, inactivating REV7-Shieldin to promote DNA end resection and homologous recombination and releasing REV7 from REV3 to counteract translesion synthesis [PMID:33051298]. Biallelic C-terminal truncating variants in MAD2L1BP that abolish Mad2/TRIP13/REV7 binding cause female infertility via oocyte metaphase I arrest and a mosaic variegated aneuploidy phenotype, with full-length cRNA rescuing the oocyte defect [PMID:37334967, PMID:37796616].","teleology":[{"year":2004,"claim":"Established that p31comet is a positive regulator of mitotic exit that antagonizes Mad2 inhibition of APC/C without disrupting the Mad2-Cdc20 interaction, defining it as a checkpoint silencer rather than a Mad2-Cdc20 competitor.","evidence":"in vitro APC/C activity assay with recombinant protein, Xenopus egg extract, co-IP, and RNAi in HeLa cells","pmids":["15257285"],"confidence":"High","gaps":["Did not resolve the conformational basis of selective binding to closed Mad2","Mechanism of how APC/C activity is enhanced left undefined"]},{"year":2006,"claim":"Provided the structural logic for checkpoint silencing by showing p31comet occupies the C-Mad2 dimerization surface and blocks O-Mad2/C-Mad2 conformational dimerization needed to propagate the checkpoint signal.","evidence":"NMR chemical shift perturbation, dimerization-interface mutagenesis, and SAC assay in S. cerevisiae","pmids":["16525508"],"confidence":"High","gaps":["Did not address action on the fully assembled MCC","No energy source for downstream disassembly identified"]},{"year":2011,"claim":"Showed p31comet acts downstream of kinetochores by binding Mad2 within the MCC and extracting it in an ATP-dependent reaction that releases Cdc20 from BubR1, and that p31comet is required for timely mitotic exit even when attachments are correct.","evidence":"co-IP, recombinant protein addition to checkpoint-arrested extracts, isolated-MCC ATP-hydrolysis assays, RNAi/overexpression, and live-cell imaging of kinetochore localization","pmids":["22100920","21300909","21965286"],"confidence":"High","gaps":["The ATPase responsible for the ATP-dependent step was not yet identified","Coupling between Mad2 extraction and Cdc20-BubR1 release left mechanistically open"]},{"year":2014,"claim":"Identified TRIP13 as the AAA-ATPase that, with p31comet, executes MCC disassembly, supplying the enzymatic engine for checkpoint silencing.","evidence":"biochemical fractionation of HeLa extracts, mass-spec identification, immunodepletion, and in vitro disassembly with purified components","pmids":["25092294"],"confidence":"High","gaps":["Stoichiometry and binding geometry of the p31comet-TRIP13-MCC assembly not yet defined"]},{"year":2015,"claim":"Defined the molecular architecture of the disassembly machine: oligomeric TRIP13 has separate subsites for p31comet and for C-Mad2 complexes, and co-occupancy drives high-affinity, productive engagement.","evidence":"purified-protein binding assays, co-IP, and mutant analysis","pmids":["26324890"],"confidence":"High","gaps":["Conformational changes during ATP-driven extraction not directly visualized"]},{"year":2014,"claim":"Revealed regulation of p31comet by phosphorylation, showing Ser-102 phosphorylation weakens Mad2 binding to limit premature SAC bypass.","evidence":"S102A mutagenesis, co-IP binding assay, and cellular SAC activity readout","pmids":["24596092"],"confidence":"Medium","gaps":["The kinase responsible was not identified in this study","Single-lab functional readout"]},{"year":2015,"claim":"Identified IKK-beta as a mitotic kinase that phosphorylates p31comet to enhance SAC silencing and M-phase exit, expanding the regulatory input on p31comet.","evidence":"Xenopus egg extract M-phase exit assay with kinase identification and phosphorylation analysis","pmids":["25892037"],"confidence":"Medium","gaps":["Phosphosite and mechanistic relationship to S102 control not resolved","Single-lab, limited mechanistic detail"]},{"year":2019,"claim":"Established PLK1 as the kinase that phosphorylates p31comet on S102 to suppress TRIP13/p31comet-mediated MCC disassembly during active checkpoint, integrating the 2014 phospho-regulation finding into a defined signaling input.","evidence":"in vitro kinase assay with purified PLK1, co-IP, BI-2536 inhibition in arrested extracts, S102A mutagenesis, and MCC disassembly assay","pmids":["31118282"],"confidence":"High","gaps":["Spatial/temporal control of PLK1 access to p31comet at kinetochores not fully defined"]},{"year":2020,"claim":"Extended p31comet/TRIP13 function beyond mitosis, showing it remodels REV7 complexes to promote homologous recombination and counteract translesion synthesis, conferring PARP-inhibitor resistance.","evidence":"co-IP, HR assay, chromatin fractionation, TRIP13-dependence, and PARP-inhibitor resistance assays","pmids":["33051298"],"confidence":"Medium","gaps":["Single-lab study","How p31comet selects REV7-Shieldin versus Pol-zeta REV7 not resolved"]},{"year":2020,"claim":"Showed p31comet sets the rate of SAC weakening and timing of mitotic slippage, linking its activity to an APC/C-to-SAC feedback loop during prolonged arrest.","evidence":"RNAi, quantitative kinetochore MAD2 imaging, and APC/C inhibitor experiments","pmids":["32029899"],"confidence":"Medium","gaps":["Single-lab","Molecular nature of the feedback step not defined"]},{"year":2023,"claim":"Demonstrated a human disease consequence, showing biallelic C-terminal truncating MAD2L1BP variants abolish Mad2/TRIP13/REV7 binding, cause oocyte metaphase I arrest and mosaic variegated aneuploidy, with full-length cRNA rescuing the oocyte phenotype.","evidence":"whole-exome sequencing, recombinant binding assays, SAC silencing and HDR assays, aneuploidy analysis, and cRNA microinjection rescue in patient and mouse oocytes","pmids":["37334967","37796616"],"confidence":"High","gaps":["The TP53 interaction reported requires further mechanistic definition","Genotype-phenotype breadth across patients not established"]},{"year":2025,"claim":"Added linear ubiquitination as a second post-translational control: HOIP modifies p31comet C-terminal lysines after cytokine stimulation to reduce PLK1 binding and thereby promote p31comet activity, with HOIP loss prolonging mitosis.","evidence":"co-IP, ubiquitination assays, C-terminal lysine mutagenesis, HOIP knockout cells, and live-cell imaging of mitotic duration","pmids":["40462232"],"confidence":"Medium","gaps":["Single-lab","Interplay between ubiquitination and S102 phosphorylation timing not resolved"]},{"year":null,"claim":"How the distinct regulatory inputs (PLK1/IKK-beta phosphorylation, HOIP ubiquitination, Rb-E2F expression control) are integrated in space and time to tune p31comet activity across mitotic and DNA-repair contexts remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified model linking phospho- and ubiquitin-regulation","Crosstalk between SAC silencing and REV7-remodeling roles undefined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[2,5,6]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[0,1,2]}],"localization":[{"term_id":"GO:0005815","term_label":"microtubule organizing center","supporting_discovery_ids":[4]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[10]}],"pathway":[{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[0,2,3,5]},{"term_id":"R-HSA-73894","term_label":"DNA Repair","supporting_discovery_ids":[10]}],"complexes":["MCC (mitotic checkpoint complex)","TRIP13-p31comet disassembly complex","REV7-Shieldin complex"],"partners":["MAD2L1","TRIP13","CDC20","BUB1B","PLK1","REV7","HOIP","TP53"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q15013","full_name":"MAD2L1-binding protein","aliases":["Caught by MAD2 protein","p31(comet)"],"length_aa":274,"mass_kda":31.1,"function":"May function to silence the spindle checkpoint and allow mitosis to proceed through anaphase by binding MAD2L1 after it has become dissociated from the MAD2L1-CDC20 complex","subcellular_location":"Nucleus; Cytoplasm, cytoskeleton, spindle","url":"https://www.uniprot.org/uniprotkb/Q15013/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/MAD2L1BP","classification":"Common Essential","n_dependent_lines":588,"n_total_lines":1208,"dependency_fraction":0.4867549668874172},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/MAD2L1BP","total_profiled":1310},"omim":[{"mim_id":"618136","title":"MAD2L1-BINDING PROTEIN; MAD2L1BP","url":"https://www.omim.org/entry/618136"},{"mim_id":"612425","title":"SHUGOSHIN-LIKE 2; SGOL2","url":"https://www.omim.org/entry/612425"},{"mim_id":"604507","title":"THYROID HORMONE RECEPTOR INTERACTOR 13; TRIP13","url":"https://www.omim.org/entry/604507"},{"mim_id":"601467","title":"MITOTIC ARREST-DEFICIENT 2 LIKE 1; MAD2L1","url":"https://www.omim.org/entry/601467"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nucleoplasm","reliability":"Supported"},{"location":"Nucleoli","reliability":"Supported"},{"location":"Nuclear membrane","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/MAD2L1BP"},"hgnc":{"alias_symbol":["CMT2","KIAA0110","dJ261G23.1","p31comet"],"prev_symbol":[]},"alphafold":{"accession":"Q15013","domains":[{"cath_id":"3.30.900.20","chopping":"55-98_119-271","consensus_level":"high","plddt":92.9954,"start":55,"end":271}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q15013","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q15013-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q15013-F1-predicted_aligned_error_v6.png","plddt_mean":80.0},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=MAD2L1BP","jax_strain_url":"https://www.jax.org/strain/search?query=MAD2L1BP"},"sequence":{"accession":"Q15013","fasta_url":"https://rest.uniprot.org/uniprotkb/Q15013.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q15013/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q15013"}},"corpus_meta":[{"pmid":"15257285","id":"PMC_15257285","title":"Conformation-specific binding of p31(comet) antagonizes the function of Mad2 in the spindle checkpoint.","date":"2004","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/15257285","citation_count":170,"is_preprint":false},{"pmid":"16525508","id":"PMC_16525508","title":"Determinants of conformational dimerization of Mad2 and its inhibition by p31comet.","date":"2006","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/16525508","citation_count":117,"is_preprint":false},{"pmid":"22100920","id":"PMC_22100920","title":"p31comet-mediated extraction of Mad2 from the MCC promotes efficient mitotic exit.","date":"2011","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/22100920","citation_count":115,"is_preprint":false},{"pmid":"25092294","id":"PMC_25092294","title":"Disassembly of mitotic checkpoint complexes by the joint action of the AAA-ATPase TRIP13 and p31(comet).","date":"2014","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/25092294","citation_count":108,"is_preprint":false},{"pmid":"21300909","id":"PMC_21300909","title":"p31comet Promotes disassembly of the mitotic checkpoint complex in an ATP-dependent process.","date":"2011","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/21300909","citation_count":82,"is_preprint":false},{"pmid":"26324890","id":"PMC_26324890","title":"Mode of interaction of TRIP13 AAA-ATPase with the Mad2-binding protein p31comet and with mitotic checkpoint complexes.","date":"2015","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/26324890","citation_count":64,"is_preprint":false},{"pmid":"21965286","id":"PMC_21965286","title":"p31(comet) acts to ensure timely spindle checkpoint silencing subsequent to kinetochore attachment.","date":"2011","source":"Molecular biology of the cell","url":"https://pubmed.ncbi.nlm.nih.gov/21965286","citation_count":52,"is_preprint":false},{"pmid":"32029899","id":"PMC_32029899","title":"Mitotic slippage is determined by p31comet and the weakening of the spindle-assembly checkpoint.","date":"2020","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/32029899","citation_count":46,"is_preprint":false},{"pmid":"27601671","id":"PMC_27601671","title":"P31comet, a member of the synaptonemal complex, participates in meiotic DSB formation in rice.","date":"2016","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/27601671","citation_count":44,"is_preprint":false},{"pmid":"33051298","id":"PMC_33051298","title":"p31comet promotes homologous recombination by inactivating REV7 through the TRIP13 ATPase.","date":"2020","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/33051298","citation_count":30,"is_preprint":false},{"pmid":"22544748","id":"PMC_22544748","title":"Depletion of p31comet protein promotes sensitivity to antimitotic drugs.","date":"2012","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/22544748","citation_count":29,"is_preprint":false},{"pmid":"19276188","id":"PMC_19276188","title":"p31comet Induces cellular senescence through p21 accumulation and Mad2 disruption.","date":"2009","source":"Molecular cancer research : MCR","url":"https://pubmed.ncbi.nlm.nih.gov/19276188","citation_count":22,"is_preprint":false},{"pmid":"24596092","id":"PMC_24596092","title":"Phosphorylation regulates the p31Comet-mitotic arrest-deficient 2 (Mad2) interaction to promote spindle assembly checkpoint (SAC) activity.","date":"2014","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/24596092","citation_count":22,"is_preprint":false},{"pmid":"31118282","id":"PMC_31118282","title":"Role of Polo-like kinase 1 in the regulation of the action of p31comet in the disassembly of mitotic checkpoint complexes.","date":"2019","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/31118282","citation_count":16,"is_preprint":false},{"pmid":"24255856","id":"PMC_24255856","title":"p31(comet) inactivates the chemically induced Mad2-dependent spindle assembly checkpoint and leads to resistance to anti-mitotic drugs.","date":"2013","source":"SpringerPlus","url":"https://pubmed.ncbi.nlm.nih.gov/24255856","citation_count":13,"is_preprint":false},{"pmid":"37334967","id":"PMC_37334967","title":"Biallelic variants in MAD2L1BP (p31) cause female infertility characterized by oocyte maturation arrest.","date":"2023","source":"eLife","url":"https://pubmed.ncbi.nlm.nih.gov/37334967","citation_count":12,"is_preprint":false},{"pmid":"24131926","id":"PMC_24131926","title":"Coordinated regulation of p31(Comet) and Mad2 expression is required for cellular proliferation.","date":"2013","source":"Cell cycle (Georgetown, Tex.)","url":"https://pubmed.ncbi.nlm.nih.gov/24131926","citation_count":12,"is_preprint":false},{"pmid":"37796616","id":"PMC_37796616","title":"Biallelic MAD2L1BP (p31comet) mutation is associated with mosaic aneuploidy and juvenile granulosa cell tumors.","date":"2023","source":"JCI insight","url":"https://pubmed.ncbi.nlm.nih.gov/37796616","citation_count":8,"is_preprint":false},{"pmid":"25892037","id":"PMC_25892037","title":"Phosphorylation of Xenopus p31(comet) potentiates mitotic checkpoint exit.","date":"2015","source":"Cell cycle (Georgetown, Tex.)","url":"https://pubmed.ncbi.nlm.nih.gov/25892037","citation_count":7,"is_preprint":false},{"pmid":"17934339","id":"PMC_17934339","title":"Mutation analysis of p31comet gene, a negative regulator of Mad2, in human hepatocellular carcinoma.","date":"2007","source":"Experimental & molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/17934339","citation_count":6,"is_preprint":false},{"pmid":"26544187","id":"PMC_26544187","title":"p31comet-Induced Cell Death Is Mediated by Binding and Inactivation of Mad2.","date":"2015","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/26544187","citation_count":4,"is_preprint":false},{"pmid":"33511944","id":"PMC_33511944","title":"The Mad2-Binding Protein p31comet as a Potential Target for Human Cancer Therapy.","date":"2021","source":"Current cancer drug targets","url":"https://pubmed.ncbi.nlm.nih.gov/33511944","citation_count":2,"is_preprint":false},{"pmid":"40462232","id":"PMC_40462232","title":"Linear ubiquitination of p31comet by HOIP couples cytokine response with mitotic regulation.","date":"2025","source":"Cell & bioscience","url":"https://pubmed.ncbi.nlm.nih.gov/40462232","citation_count":0,"is_preprint":false},{"pmid":"40243841","id":"PMC_40243841","title":"p31Comet Splice Variants Induce Distinct Spindle Assembly Checkpoint Dynamics due to Their Unique N-Termini.","date":"2025","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/40243841","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":12685,"output_tokens":5159,"usd":0.05772,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":13466,"output_tokens":4099,"usd":0.084903,"stage2_stop_reason":"end_turn"},"total_usd":0.142623,"stage1_batch_id":"msgbatch_01BbTCMyrRPyhtEq6b1MdwCN","stage2_batch_id":"msgbatch_011dEed5bbxYvTAaQWKJevxu","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2004,\n      \"finding\": \"p31comet binds selectively to the Cdc20-bound (closed) conformation of Mad2 but does not prevent the Mad2-Cdc20 interaction. Recombinant p31comet antagonizes Mad2-mediated inhibition of APC/C(Cdc20) in vitro and in Xenopus egg extracts. During checkpoint inactivation, p31comet forms a transient complex with APC/C(Cdc20)-bound Mad2 and enhances APC/C activity without disrupting Mad2-Cdc20 interaction. RNAi depletion of p31comet in HeLa cells delays mitotic exit after nocodazole removal.\",\n      \"method\": \"In vitro APC/C activity assay with recombinant protein, Xenopus egg extract assay, co-immunoprecipitation, RNAi in HeLa cells\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — in vitro reconstitution with recombinant proteins, Xenopus extract assay, and cell-based RNAi, multiple orthogonal methods in one study\",\n      \"pmids\": [\"15257285\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"p31comet competes with O-Mad2 for binding to C-Mad2, thereby blocking the conformational dimerization of Mad2 that is required for spindle assembly checkpoint activation. NMR chemical shift perturbations showed that O-Mad2 undergoes major conformational rearrangement upon C-Mad2 binding, and p31comet inhibits this by occupying the C-Mad2 binding surface.\",\n      \"method\": \"NMR chemical shift perturbation, mutational analysis of Mad2 dimerization interface, SAC assay in Saccharomyces cerevisiae\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — NMR structural analysis combined with mutagenesis and in vivo checkpoint assay, multiple orthogonal methods\",\n      \"pmids\": [\"16525508\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"p31comet promotes checkpoint silencing downstream of kinetochores by extracting Mad2 from the Mitotic Checkpoint Complex (MCC). p31comet binds Mad2 when it is within the MCC (bound to BubR1 and Cdc20). Recombinant p31comet added to checkpoint-arrested extracts removes Mad2 from the MCC; a p31comet mutant unable to bind Mad2 has no effect. RNAi of p31comet increases Mad2 bound to BubR1-Cdc20; overexpression decreases it.\",\n      \"method\": \"Co-immunoprecipitation, recombinant protein addition to checkpoint-arrested extracts, RNAi, overexpression, live-cell imaging\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal co-IP, extract-based reconstitution, mutant controls, RNAi and overexpression with multiple orthogonal readouts\",\n      \"pmids\": [\"22100920\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"p31comet promotes disassembly of the MCC in an ATP-dependent manner; specifically, it promotes dissociation of Cdc20 from BubR1 within MCC. This process requires beta,gamma-hydrolyzable ATP. p31comet binds Mad2 yet causes release of Cdc20 from BubR1.\",\n      \"method\": \"Cell-free extracts from checkpoint-arrested HeLa cells, isolated MCC, ATP hydrolysis requirement assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — biochemical reconstitution with isolated MCC, ATP analog experiments, replicated concept in same lab and by other labs\",\n      \"pmids\": [\"21300909\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"p31comet localizes to kinetochores in mitosis and traffics on and off kinetochores like Mad2. Cells depleted of p31comet arrest in metaphase with mature bipolar kinetochore-microtubule attachments and high cyclin B levels, demonstrating p31comet is required for timely mitotic exit even when the checkpoint is satisfied.\",\n      \"method\": \"Live-cell imaging, fluorescence microscopy, RNAi depletion in human and marsupial cells\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct live-imaging of localization plus RNAi loss-of-function with specific mitotic phenotype, replicated across two cell systems\",\n      \"pmids\": [\"21965286\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"TRIP13 AAA-ATPase works jointly with p31comet to disassemble MCC. TRIP13 promotes ATP- and p31comet-dependent disassembly of the Cdc20-Mad2 subcomplex and release of Mad2 from MCC, abrogating checkpoint inhibition of APC/C. TRIP13 was identified by fractionation of HeLa cell extracts as the factor responsible for this activity.\",\n      \"method\": \"Biochemical fractionation of HeLa cell extracts, in vitro disassembly assay, identification of TRIP13 by mass spectrometry, immunodepletion\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — biochemical fractionation, protein identification, in vitro reconstitution of disassembly with purified components\",\n      \"pmids\": [\"25092294\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Oligomeric TRIP13 binds both p31comet and MCC. p31comet and checkpoint complexes mutually promote each other's binding to oligomeric TRIP13. The substrate-binding site of TRIP13 has subsites specific for p31comet and C-Mad2-containing complex, and simultaneous occupancy of both subsites is required for high-affinity binding.\",\n      \"method\": \"Binding assays with purified recombinant proteins, co-immunoprecipitation, mutant analysis\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — purified component binding assays with multiple mutants, mechanistic model supported by multiple orthogonal binding experiments\",\n      \"pmids\": [\"26324890\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Phosphorylation of p31comet on Ser-102 weakens its binding to Mad2 and thereby enhances SAC activity (reduces p31comet-mediated SAC bypass). Attenuating p31comet affinity for Mad2 by phosphorylation at S102 promotes SAC activity during mitosis.\",\n      \"method\": \"Phosphorylation site mutagenesis (S102A), co-immunoprecipitation binding assay, SAC activity assay in cells\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — site-directed mutagenesis combined with binding assay and functional SAC readout, single lab\",\n      \"pmids\": [\"24596092\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Polo-like kinase 1 (Plk1) binds p31comet, phosphorylates it on S102, and suppresses its ability (together with TRIP13) to disassemble MCC. An S102A mutant of p31comet is largely resistant to Plk1-mediated inhibition. Plk1 inhibition in nocodazole-arrested HeLa cell extracts enhanced Mad2 release from checkpoint complexes.\",\n      \"method\": \"In vitro kinase assay with purified Plk1 and p31comet, co-immunoprecipitation, selective Plk1 inhibitor BI-2536 in cell extracts, S102A mutagenesis, MCC disassembly assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — purified kinase assay, site-directed mutagenesis, chemical inhibitor corroboration, functional MCC disassembly readout, multiple orthogonal methods\",\n      \"pmids\": [\"31118282\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"IKK-β phosphorylates p31comet in Xenopus egg extracts during mitosis, and this phosphorylation enhances p31comet's role in SAC silencing to promote M-phase exit.\",\n      \"method\": \"Xenopus egg extract M-phase exit assay, kinase identification, phosphorylation analysis\",\n      \"journal\": \"Cell cycle (Georgetown, Tex.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — Xenopus egg extract functional assay with kinase identification, single lab, limited mechanistic detail in abstract\",\n      \"pmids\": [\"25892037\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"p31comet binds the REV7-Shieldin complex in cells, promotes REV7 inactivation via TRIP13, and thereby promotes homologous recombination (HR) by enabling DNA end resection. p31comet also counteracts REV7 function in translesion synthesis (TLS) by releasing REV7 from REV3 in the Pol ζ complex. p31comet promotes extraction of REV7 from chromatin and causes PARP inhibitor resistance.\",\n      \"method\": \"Co-immunoprecipitation in cells, HR assay, chromatin fractionation, PARP inhibitor resistance assay, TRIP13 dependence assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP, functional HR assay, chromatin extraction assay in single lab with multiple orthogonal readouts\",\n      \"pmids\": [\"33051298\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"p31comet-mediated suppression of MAD2 activation contributes to the timing of mitotic slippage. Reduction of kinetochore MAD2 during prolonged mitotic arrest is dependent on APC/C(CDC20), revealing an APC/C-to-SAC feedback loop. p31comet controls the rate of SAC weakening and thus the timing of cyclin B1 degradation and mitotic exit.\",\n      \"method\": \"RNAi depletion of p31comet, live-cell imaging, quantitative kinetochore MAD2 measurement, APC/C inhibitor experiments\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — RNAi with quantitative imaging and APC/C inhibitor experiments, single lab\",\n      \"pmids\": [\"32029899\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"p31comet-induced cellular senescence requires direct interaction with Mad2; a p31comet mutant lacking the Mad2-binding region does not induce senescence. Senescence progression is dependent on p21(Waf1/Cip1) accumulation and is partly dependent on p53. Depletion of Mad2 by siRNA also induces senescence, phenocopying p31comet overexpression.\",\n      \"method\": \"Retroviral overexpression, domain deletion mutagenesis, siRNA knockdown, senescence assays (SA-β-gal, BrdU), flow cytometry\",\n      \"journal\": \"Molecular cancer research : MCR\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — functional mutagenesis of binding domain combined with loss-of-function siRNA and gain-of-function overexpression, single lab\",\n      \"pmids\": [\"19276188\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Depletion of p31comet increases mitotic checkpoint complex levels and prolongs mitotic block in response to spindle poisons. Cells with lower p31comet levels relative to MAD2 are less prone to mitotic slippage. The ratio of p31comet to MAD2 in cell lines correlates with susceptibility to mitotic slippage.\",\n      \"method\": \"RNAi depletion in multiple cancer cell lines, mitotic checkpoint complex quantification, cell death/slippage assays with spindle poisons\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — RNAi with MCC quantification and multiple functional readouts across multiple cell lines, single lab\",\n      \"pmids\": [\"22544748\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"p31comet-induced cell death requires the C-terminal 30 amino acids of p31comet and is mediated by binding to and inactivating Mad2. Point mutation or deletion of the Mad2-binding region abolishes cytotoxic activity. Wild-type Mad2 (but not a non-p31comet-binding mutant) inhibits p31comet-induced cell death, confirming the mechanism involves Mad2 inactivation.\",\n      \"method\": \"Retroviral transduction, C-terminal deletion and point mutation analysis, colony formation assay, co-expression rescue experiments\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — domain mutagenesis with functional readout and rescue experiment, single lab\",\n      \"pmids\": [\"26544187\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Biallelic loss-of-function variants in MAD2L1BP causing C-terminal truncation of p31comet abolish its binding to MAD2. These variants cause female infertility due to oocyte metaphase I arrest. Microinjection of full-length MAD2L1BP cRNA rescued polar body extrusion in patient oocytes and mouse oocytes, while truncated variants could not.\",\n      \"method\": \"Whole-exome sequencing, co-immunoprecipitation binding assay, cRNA microinjection rescue in mouse and patient oocytes\",\n      \"journal\": \"eLife\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — patient genetics corroborated by binding assay and direct rescue experiment in human and mouse oocytes, multiple orthogonal methods\",\n      \"pmids\": [\"37334967\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"A homozygous nonsense variant R253* in MAD2L1BP causes truncated p31comet that is defective in binding TRIP13, Mad2, and Rev7, and unable to support SAC silencing or homology-directed repair. An interaction between p31comet and TP53 was identified, and C-terminal truncation abrogated this interaction. Patient-derived cells showed high-grade mosaic variegated aneuploidy.\",\n      \"method\": \"Patient genetics, recombinant protein binding assays, SAC silencing assay, HDR assay, aneuploidy analysis of patient cells\",\n      \"journal\": \"JCI insight\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — recombinant protein binding assays and functional assays in patient-derived cells, single study with multiple readouts\",\n      \"pmids\": [\"37796616\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"p31comet (MAD2L1BP) is modified by linear (Met1-linked) ubiquitination on its C-terminal lysine residues by the E3 ligase HOIP after cytokine stimulation. Ubiquitinated p31comet shows reduced binding to PLK1, which normally phosphorylates and inactivates p31comet. HOIP-deficient cells exhibit prolonged mitotic duration similar to p31comet knockout, and cells expressing a ubiquitination-deficient p31comet mutant take longer to complete M phase.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, site-directed mutagenesis of C-terminal lysines, HOIP knockout cells, live-cell imaging of mitotic duration\",\n      \"journal\": \"Cell & bioscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP, mutagenesis, KO cell functional readout, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"40462232\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"p31comet Variant 1 (containing 32 additional N-terminal residues compared to Variant 2) exhibits reduced ability to bind MAD2 and reduced ability to induce mitotic progression. Variant 1 also shows reduced protein stability compared to Variant 2. Variant 1 is uniquely expressed in the testes.\",\n      \"method\": \"Splice variant cloning and expression, co-immunoprecipitation binding assay, mitotic progression assay, tissue expression analysis\",\n      \"journal\": \"International journal of molecular sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — binding and functional assays comparing splice variants, single lab\",\n      \"pmids\": [\"40243841\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"The Rb-E2F pathway regulates p31comet expression. A narrow range of p31comet:Mad2 ratios is compatible with cellular viability, and coordinate regulation of this antagonist-target pair is important for cell proliferation and checkpoint function.\",\n      \"method\": \"E2F reporter assay, expression analysis, cell viability assay with altered p31comet:Mad2 ratios\",\n      \"journal\": \"Cell cycle (Georgetown, Tex.)\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, expression-level regulation, limited mechanistic detail in abstract\",\n      \"pmids\": [\"24131926\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"MAD2L1BP (p31comet) is a Mad2-binding protein that silences the spindle assembly checkpoint (SAC) by two mechanisms: (1) competing with open-conformation O-Mad2 for binding to kinetochore-associated C-Mad2 to prevent new MCC assembly, and (2) binding C-Mad2 within the assembled MCC and recruiting the AAA-ATPase TRIP13, which uses ATP hydrolysis to extract Mad2 from the MCC, thereby releasing APC/C(Cdc20) from inhibition and allowing mitotic exit; p31comet activity is regulated by phosphorylation—PLK1 phosphorylates p31comet on S102 to suppress its MCC-disassembly activity during active checkpoint, while HOIP-mediated linear ubiquitination of p31comet reduces PLK1 binding and thus promotes p31comet activity; beyond mitosis, p31comet/TRIP13 also remodel REV7 complexes to promote homologous recombination and counteract translesion synthesis.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"MAD2L1BP (p31comet) is a Mad2-binding adaptor that silences the spindle assembly checkpoint (SAC) to permit timely mitotic exit [#0, #4]. It acts at two points: it competes with open-conformation O-Mad2 for the dimerization surface of closed C-Mad2, blocking the Mad2 conformational dimerization required for checkpoint activation [#1], and it binds C-Mad2 within the assembled Mitotic Checkpoint Complex (MCC), where it promotes ATP-dependent extraction of Mad2 and dissociation of Cdc20 from BubR1, releasing APC/C(Cdc20) from inhibition [#2, #3]. This disassembly is executed jointly with the AAA-ATPase TRIP13: p31comet and C-Mad2-containing checkpoint complexes occupy distinct subsites of oligomeric TRIP13 and mutually promote each other's high-affinity binding, coupling substrate recognition to ATP hydrolysis [#5, #6]. p31comet activity is set by post-translational control—PLK1 binds p31comet and phosphorylates it on Ser-102 to weaken Mad2 binding and suppress MCC disassembly during an active checkpoint, while HOIP-mediated linear (Met1-linked) ubiquitination of its C-terminal lysines reduces PLK1 binding and thereby promotes p31comet activity [#7, #8, #17]. Beyond mitosis, p31comet partners with TRIP13 to remodel REV7 complexes, inactivating REV7-Shieldin to promote DNA end resection and homologous recombination and releasing REV7 from REV3 to counteract translesion synthesis [#10]. Biallelic C-terminal truncating variants in MAD2L1BP that abolish Mad2/TRIP13/REV7 binding cause female infertility via oocyte metaphase I arrest and a mosaic variegated aneuploidy phenotype, with full-length cRNA rescuing the oocyte defect [#15, #16].\",\n  \"teleology\": [\n    {\n      \"year\": 2004,\n      \"claim\": \"Established that p31comet is a positive regulator of mitotic exit that antagonizes Mad2 inhibition of APC/C without disrupting the Mad2-Cdc20 interaction, defining it as a checkpoint silencer rather than a Mad2-Cdc20 competitor.\",\n      \"evidence\": \"in vitro APC/C activity assay with recombinant protein, Xenopus egg extract, co-IP, and RNAi in HeLa cells\",\n      \"pmids\": [\"15257285\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not resolve the conformational basis of selective binding to closed Mad2\", \"Mechanism of how APC/C activity is enhanced left undefined\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Provided the structural logic for checkpoint silencing by showing p31comet occupies the C-Mad2 dimerization surface and blocks O-Mad2/C-Mad2 conformational dimerization needed to propagate the checkpoint signal.\",\n      \"evidence\": \"NMR chemical shift perturbation, dimerization-interface mutagenesis, and SAC assay in S. cerevisiae\",\n      \"pmids\": [\"16525508\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not address action on the fully assembled MCC\", \"No energy source for downstream disassembly identified\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Showed p31comet acts downstream of kinetochores by binding Mad2 within the MCC and extracting it in an ATP-dependent reaction that releases Cdc20 from BubR1, and that p31comet is required for timely mitotic exit even when attachments are correct.\",\n      \"evidence\": \"co-IP, recombinant protein addition to checkpoint-arrested extracts, isolated-MCC ATP-hydrolysis assays, RNAi/overexpression, and live-cell imaging of kinetochore localization\",\n      \"pmids\": [\"22100920\", \"21300909\", \"21965286\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"The ATPase responsible for the ATP-dependent step was not yet identified\", \"Coupling between Mad2 extraction and Cdc20-BubR1 release left mechanistically open\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Identified TRIP13 as the AAA-ATPase that, with p31comet, executes MCC disassembly, supplying the enzymatic engine for checkpoint silencing.\",\n      \"evidence\": \"biochemical fractionation of HeLa extracts, mass-spec identification, immunodepletion, and in vitro disassembly with purified components\",\n      \"pmids\": [\"25092294\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Stoichiometry and binding geometry of the p31comet-TRIP13-MCC assembly not yet defined\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Defined the molecular architecture of the disassembly machine: oligomeric TRIP13 has separate subsites for p31comet and for C-Mad2 complexes, and co-occupancy drives high-affinity, productive engagement.\",\n      \"evidence\": \"purified-protein binding assays, co-IP, and mutant analysis\",\n      \"pmids\": [\"26324890\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Conformational changes during ATP-driven extraction not directly visualized\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Revealed regulation of p31comet by phosphorylation, showing Ser-102 phosphorylation weakens Mad2 binding to limit premature SAC bypass.\",\n      \"evidence\": \"S102A mutagenesis, co-IP binding assay, and cellular SAC activity readout\",\n      \"pmids\": [\"24596092\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"The kinase responsible was not identified in this study\", \"Single-lab functional readout\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Identified IKK-beta as a mitotic kinase that phosphorylates p31comet to enhance SAC silencing and M-phase exit, expanding the regulatory input on p31comet.\",\n      \"evidence\": \"Xenopus egg extract M-phase exit assay with kinase identification and phosphorylation analysis\",\n      \"pmids\": [\"25892037\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Phosphosite and mechanistic relationship to S102 control not resolved\", \"Single-lab, limited mechanistic detail\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Established PLK1 as the kinase that phosphorylates p31comet on S102 to suppress TRIP13/p31comet-mediated MCC disassembly during active checkpoint, integrating the 2014 phospho-regulation finding into a defined signaling input.\",\n      \"evidence\": \"in vitro kinase assay with purified PLK1, co-IP, BI-2536 inhibition in arrested extracts, S102A mutagenesis, and MCC disassembly assay\",\n      \"pmids\": [\"31118282\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Spatial/temporal control of PLK1 access to p31comet at kinetochores not fully defined\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Extended p31comet/TRIP13 function beyond mitosis, showing it remodels REV7 complexes to promote homologous recombination and counteract translesion synthesis, conferring PARP-inhibitor resistance.\",\n      \"evidence\": \"co-IP, HR assay, chromatin fractionation, TRIP13-dependence, and PARP-inhibitor resistance assays\",\n      \"pmids\": [\"33051298\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab study\", \"How p31comet selects REV7-Shieldin versus Pol-zeta REV7 not resolved\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Showed p31comet sets the rate of SAC weakening and timing of mitotic slippage, linking its activity to an APC/C-to-SAC feedback loop during prolonged arrest.\",\n      \"evidence\": \"RNAi, quantitative kinetochore MAD2 imaging, and APC/C inhibitor experiments\",\n      \"pmids\": [\"32029899\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab\", \"Molecular nature of the feedback step not defined\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Demonstrated a human disease consequence, showing biallelic C-terminal truncating MAD2L1BP variants abolish Mad2/TRIP13/REV7 binding, cause oocyte metaphase I arrest and mosaic variegated aneuploidy, with full-length cRNA rescuing the oocyte phenotype.\",\n      \"evidence\": \"whole-exome sequencing, recombinant binding assays, SAC silencing and HDR assays, aneuploidy analysis, and cRNA microinjection rescue in patient and mouse oocytes\",\n      \"pmids\": [\"37334967\", \"37796616\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"The TP53 interaction reported requires further mechanistic definition\", \"Genotype-phenotype breadth across patients not established\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Added linear ubiquitination as a second post-translational control: HOIP modifies p31comet C-terminal lysines after cytokine stimulation to reduce PLK1 binding and thereby promote p31comet activity, with HOIP loss prolonging mitosis.\",\n      \"evidence\": \"co-IP, ubiquitination assays, C-terminal lysine mutagenesis, HOIP knockout cells, and live-cell imaging of mitotic duration\",\n      \"pmids\": [\"40462232\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab\", \"Interplay between ubiquitination and S102 phosphorylation timing not resolved\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How the distinct regulatory inputs (PLK1/IKK-beta phosphorylation, HOIP ubiquitination, Rb-E2F expression control) are integrated in space and time to tune p31comet activity across mitotic and DNA-repair contexts remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified model linking phospho- and ubiquitin-regulation\", \"Crosstalk between SAC silencing and REV7-remodeling roles undefined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [2, 5, 6]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [0, 1, 2]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005815\", \"supporting_discovery_ids\": [4]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [10]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [0, 2, 3, 5]},\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [10]}\n    ],\n    \"complexes\": [\n      \"MCC (mitotic checkpoint complex)\",\n      \"TRIP13-p31comet disassembly complex\",\n      \"REV7-Shieldin complex\"\n    ],\n    \"partners\": [\n      \"MAD2L1\",\n      \"TRIP13\",\n      \"CDC20\",\n      \"BUB1B\",\n      \"PLK1\",\n      \"REV7\",\n      \"HOIP\",\n      \"TP53\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}