Affinage

MAD2L1BP

MAD2L1-binding protein · UniProt Q15013

Length
274 aa
Mass
31.1 kDa
Annotated
2026-06-10
24 papers in source corpus 20 papers cited in narrative 20 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 5/5 claims corpus-supported (100%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

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).

Mechanistic history

Synthesis pass · year-by-year structured walk · 12 steps
  1. 2004 High

    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

    PMID:15257285

    Open questions at the time
    • Did not resolve the conformational basis of selective binding to closed Mad2
    • Mechanism of how APC/C activity is enhanced left undefined
  2. 2006 High

    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

    PMID:16525508

    Open questions at the time
    • Did not address action on the fully assembled MCC
    • No energy source for downstream disassembly identified
  3. 2011 High

    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

    PMID:21300909 PMID:21965286 PMID:22100920

    Open questions at the time
    • The ATPase responsible for the ATP-dependent step was not yet identified
    • Coupling between Mad2 extraction and Cdc20-BubR1 release left mechanistically open
  4. 2014 High

    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

    PMID:25092294

    Open questions at the time
    • Stoichiometry and binding geometry of the p31comet-TRIP13-MCC assembly not yet defined
  5. 2015 High

    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

    PMID:26324890

    Open questions at the time
    • Conformational changes during ATP-driven extraction not directly visualized
  6. 2014 Medium

    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

    PMID:24596092

    Open questions at the time
    • The kinase responsible was not identified in this study
    • Single-lab functional readout
  7. 2015 Medium

    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

    PMID:25892037

    Open questions at the time
    • Phosphosite and mechanistic relationship to S102 control not resolved
    • Single-lab, limited mechanistic detail
  8. 2019 High

    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

    PMID:31118282

    Open questions at the time
    • Spatial/temporal control of PLK1 access to p31comet at kinetochores not fully defined
  9. 2020 Medium

    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

    PMID:33051298

    Open questions at the time
    • Single-lab study
    • How p31comet selects REV7-Shieldin versus Pol-zeta REV7 not resolved
  10. 2020 Medium

    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

    PMID:32029899

    Open questions at the time
    • Single-lab
    • Molecular nature of the feedback step not defined
  11. 2023 High

    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

    PMID:37334967 PMID:37796616

    Open questions at the time
    • The TP53 interaction reported requires further mechanistic definition
    • Genotype-phenotype breadth across patients not established
  12. 2025 Medium

    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

    PMID:40462232

    Open questions at the time
    • Single-lab
    • Interplay between ubiquitination and S102 phosphorylation timing not resolved

Open questions

Synthesis pass · forward-looking unresolved questions
  • 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.
  • No unified model linking phospho- and ubiquitin-regulation
  • Crosstalk between SAC silencing and REV7-remodeling roles undefined

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0060090 molecular adaptor activity 3 GO:0098772 molecular function regulator activity 3
Localization
GO:0005634 nucleus 1 GO:0005815 microtubule organizing center 1
Pathway
R-HSA-1640170 Cell Cycle 4 R-HSA-73894 DNA Repair 1
Partners
BUB1BCDC20HOIPMAD2L1PLK1REV7TP53TRIP13
Complex memberships
MCC (mitotic checkpoint complex)REV7-Shieldin complexTRIP13-p31comet disassembly complex

Evidence

Reading pass · 20 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2004 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. In vitro APC/C activity assay with recombinant protein, Xenopus egg extract assay, co-immunoprecipitation, RNAi in HeLa cells The EMBO journal High 15257285
2006 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. NMR chemical shift perturbation, mutational analysis of Mad2 dimerization interface, SAC assay in Saccharomyces cerevisiae The EMBO journal High 16525508
2011 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. Co-immunoprecipitation, recombinant protein addition to checkpoint-arrested extracts, RNAi, overexpression, live-cell imaging Journal of cell science High 22100920
2011 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. Cell-free extracts from checkpoint-arrested HeLa cells, isolated MCC, ATP hydrolysis requirement assay Proceedings of the National Academy of Sciences of the United States of America High 21300909
2011 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. Live-cell imaging, fluorescence microscopy, RNAi depletion in human and marsupial cells Molecular biology of the cell High 21965286
2014 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. Biochemical fractionation of HeLa cell extracts, in vitro disassembly assay, identification of TRIP13 by mass spectrometry, immunodepletion Proceedings of the National Academy of Sciences of the United States of America High 25092294
2015 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. Binding assays with purified recombinant proteins, co-immunoprecipitation, mutant analysis Proceedings of the National Academy of Sciences of the United States of America High 26324890
2014 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. Phosphorylation site mutagenesis (S102A), co-immunoprecipitation binding assay, SAC activity assay in cells The Journal of biological chemistry Medium 24596092
2019 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. In vitro kinase assay with purified Plk1 and p31comet, co-immunoprecipitation, selective Plk1 inhibitor BI-2536 in cell extracts, S102A mutagenesis, MCC disassembly assay Proceedings of the National Academy of Sciences of the United States of America High 31118282
2015 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. Xenopus egg extract M-phase exit assay, kinase identification, phosphorylation analysis Cell cycle (Georgetown, Tex.) Medium 25892037
2020 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. Co-immunoprecipitation in cells, HR assay, chromatin fractionation, PARP inhibitor resistance assay, TRIP13 dependence assay Proceedings of the National Academy of Sciences of the United States of America Medium 33051298
2020 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. RNAi depletion of p31comet, live-cell imaging, quantitative kinetochore MAD2 measurement, APC/C inhibitor experiments Oncogene Medium 32029899
2009 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. Retroviral overexpression, domain deletion mutagenesis, siRNA knockdown, senescence assays (SA-β-gal, BrdU), flow cytometry Molecular cancer research : MCR Medium 19276188
2012 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. RNAi depletion in multiple cancer cell lines, mitotic checkpoint complex quantification, cell death/slippage assays with spindle poisons The Journal of biological chemistry Medium 22544748
2015 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. Retroviral transduction, C-terminal deletion and point mutation analysis, colony formation assay, co-expression rescue experiments PloS one Medium 26544187
2023 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. Whole-exome sequencing, co-immunoprecipitation binding assay, cRNA microinjection rescue in mouse and patient oocytes eLife High 37334967
2023 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. Patient genetics, recombinant protein binding assays, SAC silencing assay, HDR assay, aneuploidy analysis of patient cells JCI insight Medium 37796616
2025 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. Co-immunoprecipitation, ubiquitination assay, site-directed mutagenesis of C-terminal lysines, HOIP knockout cells, live-cell imaging of mitotic duration Cell & bioscience Medium 40462232
2025 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. Splice variant cloning and expression, co-immunoprecipitation binding assay, mitotic progression assay, tissue expression analysis International journal of molecular sciences Medium 40243841
2013 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. E2F reporter assay, expression analysis, cell viability assay with altered p31comet:Mad2 ratios Cell cycle (Georgetown, Tex.) Low 24131926

Source papers

Stage 0 corpus · 24 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2004 Conformation-specific binding of p31(comet) antagonizes the function of Mad2 in the spindle checkpoint. The EMBO journal 170 15257285
2006 Determinants of conformational dimerization of Mad2 and its inhibition by p31comet. The EMBO journal 117 16525508
2011 p31comet-mediated extraction of Mad2 from the MCC promotes efficient mitotic exit. Journal of cell science 115 22100920
2014 Disassembly of mitotic checkpoint complexes by the joint action of the AAA-ATPase TRIP13 and p31(comet). Proceedings of the National Academy of Sciences of the United States of America 108 25092294
2011 p31comet Promotes disassembly of the mitotic checkpoint complex in an ATP-dependent process. Proceedings of the National Academy of Sciences of the United States of America 82 21300909
2015 Mode of interaction of TRIP13 AAA-ATPase with the Mad2-binding protein p31comet and with mitotic checkpoint complexes. Proceedings of the National Academy of Sciences of the United States of America 64 26324890
2011 p31(comet) acts to ensure timely spindle checkpoint silencing subsequent to kinetochore attachment. Molecular biology of the cell 52 21965286
2020 Mitotic slippage is determined by p31comet and the weakening of the spindle-assembly checkpoint. Oncogene 46 32029899
2016 P31comet, a member of the synaptonemal complex, participates in meiotic DSB formation in rice. Proceedings of the National Academy of Sciences of the United States of America 44 27601671
2020 p31comet promotes homologous recombination by inactivating REV7 through the TRIP13 ATPase. Proceedings of the National Academy of Sciences of the United States of America 30 33051298
2012 Depletion of p31comet protein promotes sensitivity to antimitotic drugs. The Journal of biological chemistry 29 22544748
2014 Phosphorylation regulates the p31Comet-mitotic arrest-deficient 2 (Mad2) interaction to promote spindle assembly checkpoint (SAC) activity. The Journal of biological chemistry 22 24596092
2009 p31comet Induces cellular senescence through p21 accumulation and Mad2 disruption. Molecular cancer research : MCR 22 19276188
2019 Role of Polo-like kinase 1 in the regulation of the action of p31comet in the disassembly of mitotic checkpoint complexes. Proceedings of the National Academy of Sciences of the United States of America 16 31118282
2013 p31(comet) inactivates the chemically induced Mad2-dependent spindle assembly checkpoint and leads to resistance to anti-mitotic drugs. SpringerPlus 13 24255856
2023 Biallelic variants in MAD2L1BP (p31) cause female infertility characterized by oocyte maturation arrest. eLife 12 37334967
2013 Coordinated regulation of p31(Comet) and Mad2 expression is required for cellular proliferation. Cell cycle (Georgetown, Tex.) 12 24131926
2023 Biallelic MAD2L1BP (p31comet) mutation is associated with mosaic aneuploidy and juvenile granulosa cell tumors. JCI insight 8 37796616
2015 Phosphorylation of Xenopus p31(comet) potentiates mitotic checkpoint exit. Cell cycle (Georgetown, Tex.) 7 25892037
2007 Mutation analysis of p31comet gene, a negative regulator of Mad2, in human hepatocellular carcinoma. Experimental & molecular medicine 6 17934339
2015 p31comet-Induced Cell Death Is Mediated by Binding and Inactivation of Mad2. PloS one 4 26544187
2021 The Mad2-Binding Protein p31comet as a Potential Target for Human Cancer Therapy. Current cancer drug targets 2 33511944
2025 p31Comet Splice Variants Induce Distinct Spindle Assembly Checkpoint Dynamics due to Their Unique N-Termini. International journal of molecular sciences 0 40243841
2025 Linear ubiquitination of p31comet by HOIP couples cytokine response with mitotic regulation. Cell & bioscience 0 40462232

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