Affinage

MAD2L1

Mitotic spindle assembly checkpoint protein MAD2A · UniProt Q13257

Length
205 aa
Mass
23.5 kDa
Annotated
2026-04-28
100 papers in source corpus 40 papers cited in narrative 40 extracted findings

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

MAD2L1 (MAD2) is a conformationally metamorphic spindle assembly checkpoint (SAC) protein that prevents premature anaphase by inhibiting the anaphase-promoting complex/cyclosome (APC/C). MAD2 exists in interconvertible open (O-Mad2) and closed (C-Mad2) conformations; at unattached kinetochores, Mad1-bound C-Mad2 acts as a template to catalyze conversion of cytosolic O-Mad2 into C-Mad2 complexed with CDC20, generating the mitotic checkpoint complex (MCC: MAD2–BUBR1–BUB3–CDC20) that directly inhibits APC/C-mediated ubiquitination of securin and cyclin B (PMID:15694304, PMID:22037211, PMID:9637688). Checkpoint silencing is driven by the AAA+ ATPase TRIP13, which, together with the adaptor p31comet, threads the MAD2 N-terminus through its axial pore and mechanically unfolds C-Mad2 back to inactive O-Mad2, disassembling the MCC (PMID:29973720, PMID:25918846). Beyond canonical SAC signaling, C-Mad2 forms a separase-inhibitory complex with shugoshin 2 (SGO2) that functionally substitutes for securin, and MAD2 overexpression hyperstabilizes kinetochore–microtubule attachments independently of the checkpoint through effects on Aurora B localization (PMID:32322060, PMID:22405866). MAD2 abundance is transcriptionally controlled by the β-TrCP–REST degradation axis and TRRAP-associated histone acetyltransferases, while phosphorylation at S195 inhibits the O-to-C conformational transition and Cdc20 binding, providing post-translational tuning of checkpoint strength (PMID:18354482, PMID:21041666, PMID:15549134).

Mechanistic history

Synthesis pass · year-by-year structured walk · 19 steps
  1. 1996 High

    Establishing MAD2 as an essential vertebrate SAC component resolved whether the yeast checkpoint had a functional human counterpart and showed MAD2 monitors kinetochore–spindle attachment.

    Evidence Antibody electroporation in HeLa cells caused premature mitotic exit; immunofluorescence showed kinetochore localization only on unattached chromosomes

    PMID:8824189

    Open questions at the time
    • Downstream target of MAD2 inhibition not identified
    • Mechanism of MAD2 removal from attached kinetochores unknown
  2. 1997 High

    Demonstrating that MAD2 directly associates with and inhibits APC/C via CDC20 identified the molecular target of checkpoint signaling.

    Evidence Co-IP of MAD2 with APC/C in mitotic cells; purified MAD2 arrested Xenopus extracts in metaphase by blocking cyclin B ubiquitination

    PMID:9356466

    Open questions at the time
    • Structural basis of MAD2–CDC20 interaction unknown
    • Whether MAD2 alone is sufficient for full APC/C inhibition unclear
  3. 1998 High

    Discovery that MAD2 forms a ternary MAD2–CDC20–APC complex and that its oligomeric state determines inhibitory potency revealed that MAD2's structural conformation is functionally critical.

    Evidence Xenopus embryo injection, in vitro APC assay, and gel filtration showed tetrameric but not monomeric MAD2 inhibited APC/C

    PMID:9637688

    Open questions at the time
    • Atomic structure not yet determined
    • Nature of the two folded states unresolved
  4. 2000 High

    The NMR structure of MAD2 revealed a novel α/β fold and showed that the flexible C-terminal region becomes ordered upon CDC20 binding, providing the first atomic framework for the checkpoint interaction.

    Evidence Solution NMR structure determination plus deletion mutagenesis defining the minimal CDC20-binding region

    PMID:10700282

    Open questions at the time
    • Whether MAD2 adopts multiple stable folds not yet recognized
    • Structure of the MAD2–CDC20 complex not determined
  5. 2001 High

    Multiple studies established that Mad1 recruits Mad2 to kinetochores via direct binding, that Mad2 forms mutually exclusive complexes with Mad1 and Cdc20, and that Mad2 haploinsufficiency causes chromosomal instability — linking checkpoint protein dosage to genome integrity.

    Evidence RNAi of Mad1 ablated Mad2 kinetochore localization; NMR showed ligand binding triggers extensive conformational change; heterozygous Mad2 knockout in mice and human cells produced premature chromatid separation; monomeric Mad2 mutant retained checkpoint activity

    PMID:11201745 PMID:11707408 PMID:11804586

    Open questions at the time
    • Identity of the two Mad2 conformational states not yet defined at atomic level
    • Mechanism of signal amplification from kinetochores unclear
  6. 2002 High

    Identification of BubR1 as a synergistic co-inhibitor with Mad2 of APC/C-Cdc20, and discovery of p31comet as a Mad2-binding negative regulator, defined both the composition of the inhibitory signal and its silencing mechanism.

    Evidence In vitro APC/C inhibition with purified proteins showed BubR1–Mad2 synergy; co-IP identified CMT2/p31comet whose overexpression caused premature securin destruction

    PMID:11907259 PMID:12456649

    Open questions at the time
    • Stoichiometry and order of MCC assembly unknown
    • Molecular mechanism of p31comet-mediated checkpoint silencing unresolved
  7. 2003 High

    Aurora B and the Ndc80 complex were shown to act upstream of Mad2 kinetochore recruitment, while cell-cycle-dependent phosphorylation of Mad2 was found to negatively regulate its binding to Mad1 and APC/C.

    Evidence Aurora B inhibition/RNAi abolished Mad2 kinetochore localization; Nuf2/Hec1 RNAi reduced Mad1/Mad2 at kinetochores; phospho-mimetic Mad2 mutants lost Mad1/APC interactions

    PMID:12574116 PMID:12719470 PMID:14654001

    Open questions at the time
    • Specific kinase(s) phosphorylating Mad2 in vivo not identified
    • Hierarchy between Aurora B and Ndc80 in Mad2 recruitment unclear
  8. 2004 High

    NMR determination of two natively folded Mad2 states (O-Mad2 and C-Mad2) and demonstration that C-Mad2 is the active APC/C inhibitor established the two-state model, while TRRAP-dependent histone acetylation was shown to control Mad2 transcription.

    Evidence NMR structures of both conformers; C-Mad2 was more potent in APC/C inhibition; ChIP showed TRRAP/HAT at Mad2 promoter; Mad2 ectopic expression rescued Trrap-deficient cells

    PMID:15024386 PMID:15549134

    Open questions at the time
    • How Mad1 catalyzes O-to-C conversion not structurally resolved
    • Whether transcriptional regulation is tissue-specific unknown
  9. 2005 High

    The 'Mad2 template model' was established: kinetochore-bound Mad1–C-Mad2 serves as a template/receptor for cytosolic O-Mad2, catalytically converting it to C-Mad2–Cdc20 and amplifying checkpoint signaling away from kinetochores.

    Evidence Mutagenesis, FRAP live imaging, and co-IP showed O-Mad2:C-Mad2 interaction is essential for checkpoint function

    PMID:15694304

    Open questions at the time
    • Structural basis of O-Mad2:C-Mad2 asymmetric dimer unknown
    • Rate-limiting step of catalytic conversion not defined
  10. 2006 High

    Characterization of the O-Mad2:C-Mad2 conformational dimer interface by NMR and yeast genetics, plus demonstration that p31comet competes with O-Mad2 for C-Mad2 binding, unified the template and silencing models.

    Evidence NMR chemical shift perturbation, interface mutations abolished SAC in S. cerevisiae, competition assays showed p31comet displaces O-Mad2

    PMID:16525508

    Open questions at the time
    • Crystal structure of O-Mad2:C-Mad2 dimer not yet obtained
    • Whether p31comet binding is regulated by post-translational modification unknown
  11. 2007 High

    The crystal structure of the asymmetric O-Mad2:C-Mad2 dimer provided atomic-level validation of the template model and revealed the hydrophobic rearrangements underlying the topological conformational switch.

    Evidence X-ray crystallography with functional mutagenesis validation

    PMID:18022367

    Open questions at the time
    • Structure of complete Mad1–C-Mad2 core complex at kinetochore not determined
    • Energetic landscape of O-to-C transition not quantified
  12. 2008 High

    Mps1 kinase was established as the key catalyst for O-Mad2 recruitment to kinetochore-bound Mad1–C-Mad2, the REST–β-TrCP degradation axis was shown to control MAD2 transcription, and the C-Mad2 symmetric dimer structure was determined.

    Evidence Catalytically inactive Mps1 restored Mad1 but not Mad2 kinetochore localization; stable REST mutant suppressed Mad2 expression producing checkpoint defects; crystal structure of C-C Mad2 dimer

    PMID:18318601 PMID:18354482 PMID:18541701

    Open questions at the time
    • Direct Mps1 substrates enabling O-Mad2 recruitment not identified
    • Whether REST regulation of Mad2 is relevant in non-transformed cells unclear
  13. 2010 High

    Sustained Mps1 activity was shown to be continuously required for O-Mad2 recruitment even after the Mad1–C-Mad2 core is kinetochore-bound, and phosphorylation at S195 was demonstrated to block the O-to-C conformational transition by inhibiting Cdc20 binding.

    Evidence Timed Mps1 inhibitor (AZ3146) treatment dissected sequential steps; intein-mediated semisynthesis of pS195-Mad2 plus NMR showed blocked conformational change

    PMID:20624899 PMID:21041666

    Open questions at the time
    • Kinase responsible for S195 phosphorylation in vivo not identified
    • Whether Mps1 directly phosphorylates Mad2 or acts indirectly unknown
  14. 2011 High

    The order of MCC assembly was defined — BUB3–BUBR1–CDC20 forms in G2, then C-Mad2 joins during mitosis — and the C-Mad2 residues (R133/Q134) mediating BUBR1 interaction were mapped, establishing that C-Mad2 incorporation is the rate-limiting, mitosis-specific step.

    Evidence Biochemical fractionation of endogenous MCC across cell cycle; mutagenesis showed R133A/Q134A abolished BUBR1 binding and APC/C inhibition

    PMID:21525009 PMID:22037211

    Open questions at the time
    • Whether MCC assembly occurs only at kinetochores or also in the cytoplasm not resolved
    • Structural model of complete MCC not yet available
  15. 2012 High

    A second mechanism of APC/C inhibition was uncovered — Mad2 directly competes with APC/C for a binding site on Cdc20 — and Mad2 overexpression was found to hyperstabilize kinetochore–microtubule attachments independently of the SAC via Aurora B relocalization.

    Evidence In vitro competition assay showed Mad2 and APC/C compete for Cdc20; Cdc20 mutant unable to bind Mad2 abrogated SAC; Mad2 overexpression with Mad1 depletion still altered k-MT stability and Aurora B localization

    PMID:22405866 PMID:23007648

    Open questions at the time
    • Relative contribution of two inhibitory mechanisms in vivo not quantified
    • Mechanism by which Mad2–Cdc20 alters Aurora B localization unknown
  16. 2015 High

    TRIP13 was structurally and biochemically established as the AAA+ ATPase that, with p31comet adaptor, catalyzes C-Mad2→O-Mad2 remodeling to silence the SAC, while the I-Mad2 intermediate structure revealed the allosteric pathway of the conformational transition.

    Evidence Cryo-EM/crystal structures of TRIP13; NMR showed local unfolding of Mad2 C-terminal region; crystal structure of I-Mad2:C-Mad2 defined intermediate state and allosteric network

    PMID:25918846 PMID:26305957 PMID:29208896

    Open questions at the time
    • Kinetics of TRIP13-mediated remodeling in vivo not measured
    • Whether TRIP13 acts on MCC-bound or free C-Mad2 in cells debated
  17. 2018 High

    Cryo-EM of the TRIP13–p31comet–C-MAD2–CDC20 complex revealed the complete mechanical mechanism: p31comet positions MAD2's N-terminus into TRIP13's axial pore, and ATP-driven translocation unwinds the αA helix, destabilizing the β-sheet that locks the closed conformation.

    Evidence Cryo-electron microscopy of the quaternary complex with molecular modeling

    PMID:29973720

    Open questions at the time
    • Whether TRIP13 can remodel MCC already bound to APC/C in vivo remains unclear
    • Regulation of TRIP13 activity during the cell cycle not defined
  18. 2020 High

    Discovery that C-Mad2 forms a complex with SGO2 that directly inhibits separase via a pseudo-substrate mechanism — functionally replacing securin — revealed a checkpoint-effector role for MAD2 beyond APC/C inhibition.

    Evidence Co-IP, in vitro separase inhibition reconstitution, genetic knockouts; TRIP13–p31comet disassembled SGO2–MAD2 in vitro

    PMID:32322060

    Open questions at the time
    • Whether SGO2–MAD2 complex has physiological importance outside securin-null contexts unknown
    • Structural basis of SGO2–MAD2 interaction not determined
  19. 2021 High

    Real-time biosensor imaging at kinetochores revealed that Mad2–Cdc20 assembly is driven by a tripartite mechanism requiring localized substrate delivery plus two phosphorylation-dependent interactions that geometrically constrain Cdc20 for Mad2 capture.

    Evidence Live-cell imaging with kinetochore-specific biosensor probe; phosphorylation-dependent interaction dissection

    PMID:33384372

    Open questions at the time
    • Identities of all kinases providing the two phosphorylation inputs at kinetochores not fully resolved
    • Whether geometric constraint model applies in all cell types unknown

Open questions

Synthesis pass · forward-looking unresolved questions
  • Key unresolved questions include: the in vivo kinase(s) responsible for MAD2 S195 phosphorylation and its cell-cycle timing; the structural basis of the complete kinetochore-bound Mad1–C-Mad2 catalytic platform; whether TRIP13 acts preferentially on free C-Mad2, MCC, or APC/C-bound MCC in living cells; and the physiological significance of the SGO2–MAD2 separase inhibitory complex in wild-type cells.
  • No in vivo identification of the S195 kinase
  • No structure of kinetochore-assembled Mad1–Mad2 catalytic platform
  • Relative TRIP13 substrates (free C-Mad2 vs MCC vs APC/C-bound MCC) not resolved in cells

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0098772 molecular function regulator activity 6 GO:0060090 molecular adaptor activity 3
Localization
GO:0005694 chromosome 7 GO:0005635 nuclear envelope 2 GO:0005829 cytosol 2
Pathway
R-HSA-1640170 Cell Cycle 8 R-HSA-392499 Metabolism of proteins 6
Partners
BUB3BUBR1CDC20MAD1L1MAD2L1BPSGO2TPRTRIP13
Complex memberships
MCC (MAD2-BUBR1-BUB3-CDC20)Mad1-C-Mad2 core complexSGO2-MAD2 separase inhibitory complex

Evidence

Reading pass · 40 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
1996 hsMAD2 (MAD2L1) is a necessary component of the mitotic checkpoint in HeLa cells; antibody electroporation caused premature mitotic exit. MAD2 localizes to kinetochores after chromosome condensation but is absent from kinetochores at metaphase, suggesting it monitors spindle-kinetochore attachment completion. Antibody electroporation (loss-of-function), immunofluorescence localization Science High 8824189
1997 MAD2 associates with the cyclosome/anaphase-promoting complex (APC/C) upon mitotic checkpoint activation, and purified MAD2 arrests cycling Xenopus egg extracts in metaphase by blocking cyclin B ubiquitination, establishing MAD2 as a direct inhibitor of APC/C. Co-immunoprecipitation, Xenopus egg extract cell-free assay, in vitro ubiquitination assay Proceedings of the National Academy of Sciences High 9356466
1998 MAD2 prevents APC/C activation by forming a ternary hMAD2-CDC20-APC complex; MAD2 injection into Xenopus embryos arrests cells in mitosis with inactive APC. MAD2 exists in two folded states (tetramer and monomer), both binding CDC20, but only the tetramer inhibits APC/C, indicating structural state is critical for checkpoint signaling. Xenopus embryo microinjection, in vitro APC activity assay, gel filtration/biochemical fractionation, co-immunoprecipitation Genes & Development High 9637688
2000 Solution NMR structure of human MAD2 determined; MAD2 has a novel three-layered α/β fold. The minimal MAD2-binding region of CDC20 is a 40-residue segment N-terminal to its WD40 repeats, and the C-terminal flexible region of MAD2 is required for CDC20 binding, becoming structured upon complex formation. NMR spectroscopy, deletion mutagenesis, NMR titration Nature Structural Biology High 10700282
2001 RNAi-mediated suppression of Mad1 in mammalian cells causes loss of Mad2 kinetochore localization and impairment of the spindle checkpoint. Binding of Mad2 to Mad1 or Cdc20 peptides triggers extensive rearrangement of Mad2 tertiary structure, suggesting a common conformational change mechanism upon ligand binding. RNAi knockdown, NMR spectroscopy, peptide-binding structural analysis Molecular Cell High 11804586
2001 MAD2 haplo-insufficiency (deletion of one MAD2 allele) results in a defective mitotic checkpoint, premature sister-chromatid separation in the presence of spindle inhibitors, and elevated chromosome mis-segregation rates, demonstrating partial loss of MAD2 is sufficient to cause chromosomal instability. Gene deletion (heterozygous knockout in mice and human cancer cells), flow cytometry, chromosome analysis Nature High 11201745
2001 Mad2 interaction with Mad1 is crucial for localization of Mad2 to kinetochores; at kinetochores, Mad2 interacts with Cdc20. Mad2 forms mutually exclusive, oligomerization-independent complexes with Mad1 and Cdc20. A monomeric Mad2 point mutant still causes cell cycle arrest of comparable strength, showing oligomerization is not required for checkpoint function. Co-immunoprecipitation, kinetochore localization by immunofluorescence, Xenopus embryo injection, mutational analysis The EMBO Journal High 11707408
2001 HsMad1 and HsMAD2 associate with nuclear pore complexes throughout interphase, as demonstrated by co-localization with nucleoporin antibodies and co-purification with enriched nuclear envelope fractions, suggesting a non-mitotic role for the Mad1/Mad2 complex. Immunofluorescence co-localization, subcellular fractionation/co-purification Journal of Cell Science Medium 11181178
2002 BubR1 and Mad2 each independently inhibit Cdc20-APC/C activation. At physiological concentrations, BubR1 and Mad2 mutually promote each other's binding to Cdc20 and act synergistically to quantitatively inhibit APC/C; BubR1 is ~12-fold more potent than Mad2 as an APC/C inhibitor. BubR1-Cdc20 inhibition does not require BubR1 kinase activity. In vitro APC/C activity assay with purified recombinant proteins, quantitative biochemistry Molecular Biology of the Cell High 11907259
2002 A novel MAD2-binding protein, CMT2 (later identified as p31comet), interacts with MAD2. Formation of the CMT2-MAD2 complex coincides with dissociation of the p55CDC-MAD2 complex upon spindle attachment completion. CMT2 overexpression causes premature securin destruction and mitotic exit, while CMT2 depletion delays anaphase onset. Co-immunoprecipitation, overexpression and depletion functional assays The EMBO Journal Medium 12456649
2003 Aurora B kinase activity is required for kinetochore localization of spindle checkpoint components BubR1, Mad2, and Cenp-E. Aurora B inhibition with ZM447439 or Aurora B RNAi prevents mitotic arrest after spindle damage and abolishes kinetochore recruitment of Mad2, linking chromosome alignment sensing to checkpoint protein localization. Small molecule Aurora kinase inhibition (ZM447439), RNAi, immunofluorescence localization Journal of Cell Biology High 12719470
2003 Nuf2 and Hec1 (Ndc80 complex components) are required for retention of Mad1 and Mad2 at kinetochores; RNAi depletion of either protein causes >5-fold reduction of Mad1 and Mad2 at kinetochores, which is microtubule-dependent and reversible upon spindle depolymerization. RNAi knockdown, immunofluorescence quantification of kinetochore localization Current Biology Medium 14654001
2003 MAD2 phosphorylation on multiple serine residues occurs in a cell cycle-dependent manner in vivo; only unphosphorylated MAD2 interacts with Mad1 or the APC/C. A phospho-mimicking MAD2 mutant (S→D) fails to interact with Mad1 or APC/C and acts as a dominant-negative antagonist of wild-type MAD2. In vivo phosphorylation analysis, co-immunoprecipitation, dominant-negative overexpression The EMBO Journal Medium 12574116
2004 Mad2 adopts two distinct natively folded conformations at equilibrium without cofactor binding (termed N1-Mad2/O-Mad2 and N2-Mad2/C-Mad2). NMR structure of N2-Mad2 determined. N2/C-Mad2 is more potent in APC/C inhibition. Interconversion is slow in vitro but accelerated by a Mad1 fragment. Overexpression of a Mad2 mutant that sequesters N2-Mad2 partially blocks checkpoint signaling in cells. NMR spectroscopy, in vitro APC/C inhibition assay, equilibrium conformational analysis, cell overexpression Nature Structural & Molecular Biology High 15024386
2005 The Mad1-bound closed conformer of Mad2 (C-Mad2) serves as a template/receptor for cytosolic open Mad2 (O-Mad2) at kinetochores; O-Mad2 and C-Mad2 interaction is essential for the spindle checkpoint. This interaction enables conversion of O-Mad2 into C-Mad2 bound to Cdc20, constituting the 'Mad2 template model' for amplification of the checkpoint signal away from kinetochores. Mutational analysis, fluorescence microscopy (live imaging, FRAP), co-immunoprecipitation, epistasis Current Biology High 15694304
2006 The molecular determinants of the O-Mad2:C-Mad2 conformational dimer were characterized. Mutation of individual interface residues abrogates the SAC in S. cerevisiae. NMR chemical shift perturbations show O-Mad2 undergoes major conformational rearrangement upon binding C-Mad2. p31comet competes with O-Mad2 for C-Mad2 binding, explaining its negative regulatory role on the SAC. NMR spectroscopy, yeast genetic assay (SAC abolition), mutational analysis, co-immunoprecipitation The EMBO Journal High 16525508
1999 Mad2 binds to phosphorylated kinetochores but not to unphosphorylated ones in lysed PtK1 cells, providing a molecular link between attachment-sensitive kinetochore phosphorylation and Mad2 recruitment to unattached kinetochores. Lysed cell in vitro kinetochore binding assay, phosphatase treatment, immunofluorescence Current Biology Medium 10375530
1999 FAT10, an MHC-encoded ubiquitin-like protein, noncovalently associates with MAD2 as identified by yeast two-hybrid screening and co-immunoprecipitation, suggesting FAT10 may modulate MAD2 activity during cell growth. Yeast two-hybrid, co-immunoprecipitation Proceedings of the National Academy of Sciences Low 10200259
2007 Crystal structure of the O-Mad2:C-Mad2 conformational dimer reveals an asymmetric interface explaining selective dimerization. Buried hydrophobic residues undergo rearrangement correlated with the topological change. The structure supports a catalytic model where C-Mad2 template facilitates O-Mad2 binding to Cdc20. X-ray crystallography, mutational functional validation Cell High 18022367
2008 Crystal structure of the symmetric C-Mad2:C-Mad2 (C-C) dimer determined, revealing the structural basis for unliganded C-Mad2 (but not O-Mad2 or ligand-bound C-Mad2) forming symmetric dimers. The Mad1-Mad2 core complex facilitates conversion of O-Mad2 to C-Mad2 in vitro. X-ray crystallography, in vitro conformational conversion assay, cell-based functional assays PLoS Biology High 18318601
2008 Mps1 catalytic activity is required for recruitment of Mad2 (but not Mad1) to kinetochores; catalytically inactive Mps1 restores Mad1 kinetochore localization but not Mad2. Mps1 kinase activity restrains anaphase during unperturbed mitosis. RNAi complementation with catalytically inactive mutant and analogue-sensitive allele, immunofluorescence Journal of Cell Biology High 18541701
2008 SCF(β-TrCP) degrades REST during G2, allowing transcriptional derepression of Mad2 (a REST target gene). Expression of stable REST (unable to bind β-TrCP) or oncogenic REST-FS inhibits Mad2 expression, producing checkpoint defects analogous to Mad2+/- cells, demonstrating transcriptional regulation of Mad2 by the β-TrCP-REST axis. Unbiased protein interaction screen, co-immunoprecipitation, stable mutant expression, flow cytometry, fluorescence microscopy Nature High 18354482
2010 Sustained Mps1 activity during mitosis is required for recruitment of open Mad2 (O-Mad2) to the Mad1-C-Mad2 core complex at kinetochores. Mps1 inhibition after mitotic entry leaves the Mad1-C-Mad2 core complex kinetochore-bound but abolishes O-Mad2 recruitment. Mps1 can dimerize and transphosphorylate in cells, promoting its own release from kinetochores to facilitate O-Mad2 recruitment. Novel Mps1 inhibitor (AZ3146), immunofluorescence, co-immunoprecipitation Journal of Cell Biology High 20624899
2010 Phosphorylation of MAD2 on S195 inhibits its conformational transition from O-Mad2 to C-Mad2. Phospho-mimicking Mad2(S195D) fails to bind Cdc20 but retains binding to high-affinity ligands Mad1 and MBP1. Overexpression of Mad2(S195D) causes checkpoint defects in human cells. Phospho-mimetic mutagenesis, intein-mediated semisynthesis of phosphorylated protein, NMR, co-immunoprecipitation, cell-based checkpoint assay Proceedings of the National Academy of Sciences High 21041666
2011 Endogenous human mitotic checkpoint complex (MCC) is assembled by first forming a BUBR1:BUB3:CDC20 complex in G2, followed by selective incorporation of closed MAD2 (C-MAD2) during mitosis. A recombinant MCC containing C-MAD2 effectively inhibits APC/C, whereas BUBR1:BUB3:CDC20 alone is ineffective at comparable concentrations. Biochemical fractionation, co-immunoprecipitation, in vitro APC/C inhibition assay, expression of conformation-locked MAD2 mutant Cell Cycle High 22037211
2011 BUBR1 directly interacts with closed MAD2 (C-MAD2) via Arg133 and Gln134 of C-MAD2; this interaction is essential for MCC-mediated inhibition of APC/C. The same C-MAD2 residues are required for MAD2 dimerization and p31comet binding. Co-immunoprecipitation with mutant proteins, in vitro APC/C inhibition assay Journal of Biological Chemistry Medium 21525009
2012 Mad2 inhibits Cdc20 by binding directly to a site on Cdc20 required for APC/C binding; Mad2 and APC/C compete for Cdc20 in vitro. A Cdc20 mutant that does not stably bind Mad2 abrogates the SAC in vivo, uncovering a second mechanism by which the SAC inhibits APC/C. In vitro competition assay, co-immunoprecipitation, cell-based SAC functional assay with Cdc20 mutant Journal of Cell Biology High 23007648
2012 Mad2 overexpression hyperstabilizes kinetochore-microtubule (k-MT) attachments independent of the mitotic checkpoint by altering centromeric localization and activity of Aurora B kinase. This checkpoint-independent function of Mad2 requires Cdc20 and explains why Mad2 overexpression increases chromosome missegregation. Mad2 overexpression, Mad1 depletion (to uncouple checkpoint), k-MT attachment stability assay, Aurora B localization/activity measurement Current Biology High 22405866
2013 Tpr (nuclear pore complex protein) stabilizes Mad1 and Mad2 protein levels before mitosis by forming a complex (TM2 complex) with them during interphase and mitosis. Tpr is required for Mad1-C-Mad2 recruitment to NPCs and for normal Mad2 levels at kinetochores; overexpression of GFP-Mad2 restores SAC response in Tpr-depleted cells. Tpr may regulate SAC proteostasis through SUMO-isopeptidases SENP1 and SENP2 at NPCs. Co-immunoprecipitation, protein half-life measurement, RNAi depletion, rescue by GFP-Mad2 overexpression, immunofluorescence Journal of Cell Biology Medium 24344181
2015 TRIP13 AAA+ ATPase, aided by adapter protein p31comet, converts MAD2 from the signaling-active closed conformer (C-MAD2) to the inactive open conformer (O-MAD2), thereby inactivating the spindle assembly checkpoint and promoting disassembly of mitotic checkpoint complexes. The PCH-2 (C. elegans TRIP13 ortholog) structure reveals it as a new AAA+ protein remodeler with substrate-recognition domain related to NSF and p97. Cryo-EM/structural analysis of C. elegans TRIP13, in vitro MAD2 conformational conversion assay, functional genetics eLife High 25918846
2015 TRIP13, together with p31comet, prevents APC/C inhibition by free MCC components but cannot reactivate APC/C already bound to MCC. Crystal structure of human TRIP13 determined. TRIP13 and p31comet catalyze conversion of C-Mad2 to O-Mad2 through local unfolding of the Mad2 C-terminal region without disrupting the stable folded core, as shown by NMR. NMR spectroscopy, X-ray crystallography of human TRIP13, in vitro APC/C inhibition assay, mutagenesis Nature Communications High 29208896
2015 Structure of an intermediate Mad2 conformer (I-Mad2) bound to C-Mad2 determined by X-ray crystallography and NMR; I-Mad2 retains O-Mad2 fold but core elements move toward C-Mad2 configuration. An allosteric network connects the C-Mad2-binding site to the conformationally malleable C-terminal region. Mutations at the I-Mad2:C-Mad2 interface hinder I-Mad2 formation and impede the structural transition. X-ray crystallography, NMR spectroscopy, mutagenesis Proceedings of the National Academy of Sciences High 26305957
2018 Cryo-EM structures of the TRIP13-p31comet-C-MAD2-CDC20 complex reveal that p31comet recruits C-MAD2 to TRIP13 hexameric ring, positioning MAD2's N-terminus (MAD2NT) into TRIP13's axial pore. ATP-driven translocation pushes on and rotates the p31comet-C-MAD2 complex, unwinding a region of the αA helix of C-MAD2 required to stabilize its β-sheet, destabilizing C-MAD2 in favor of O-MAD2. Cryo-electron microscopy, molecular modeling Nature High 29973720
2021 Kinetochore-catalyzed Mad2-Cdc20 assembly occurs through a tripartite mechanism: localized delivery of Mad2 and Cdc20 substrates, plus two phosphorylation-dependent interactions that geometrically constrain their positions and prime Cdc20 for interaction with Mad2. This was established with a probe specifically monitoring the assembly reaction at kinetochores in living cells. Live-cell imaging with specific biosensor probe for Mad2-Cdc20 assembly at kinetochores, phosphorylation-dependent interaction analysis Science High 33384372
2020 MAD2 associates with shugoshin 2 (SGO2) in a SAC-activated manner to create a separase inhibitor (SGO2-MAD2 complex) that can functionally replace securin. SGO2-MAD2 sequesters most separase in securin-knockout cells and uses a pseudo-substrate sequence to block the active site of separase. TRIP13-p31comet liberates separase from SGO2-MAD2 in vitro. Co-immunoprecipitation, in vitro reconstitution (separase inhibition assay), genetic knockouts, TRIP13-p31comet in vitro disassembly assay Nature High 32322060
1997 Human MAD2 (hMAD2) interacts with the C-terminal 30 amino acids of the insulin receptor (IR) cytoplasmic domain but not with IGF-IR; this interaction does not require IR kinase activity and is reduced upon IR autophosphorylation/activation, suggesting MAD2 is released from activated IR. Yeast two-hybrid, GST pulldown in vitro, co-immunoprecipitation from mammalian cells Journal of Biological Chemistry Low 9092546
2004 TRRAP (HAT cofactor) controls mitotic checkpoint integrity by regulating transcription of Mad1 and Mad2 genes through histone H4 and H3 acetylation at their promoters. Trrap associates with HATs Tip60 and PCAF at Mad1/Mad2 promoters in a cell cycle-dependent manner; ectopic expression of Mad1 and Mad2 fully restores the mitotic checkpoint in Trrap-deficient cells. Chromatin immunoprecipitation, RNAi/conditional knockout, ectopic expression rescue The EMBO Journal Medium 15549134
2009 Nek2 kinase physically associates with Mad2 and Cdc20 and can phosphorylate both proteins in vitro; overexpression of Nek2 enhances Mad2-induced mitotic delay, suggesting Nek2 regulates the Mad2-Cdc20 mitotic checkpoint complex. Co-immunoprecipitation, in vitro kinase assay, overexpression functional assay Experimental and Molecular Pathology Low 20034488
2013 Chk1 co-localizes and physically associates with Mad2 in cells under unstressed and DNA-damaged conditions. Chk1 phosphorylates Mad2 in vitro on multiple sites; a Mad2 mutant lacking all six Chk1 phosphorylatable sites cannot be phosphorylated by Chk1, suggesting a crosslink between DNA damage and mitotic spindle checkpoints. Co-immunoprecipitation, in vitro kinase assay, mutagenesis, immunofluorescence co-localization Cell Cycle Low 23454898
2014 The FAT10-MAD2 interaction interface was mapped to FAT10's first ubiquitin-like domain (NMR structure determined). Disruption of FAT10-MAD2 interaction through mutation of specific MAD2-binding residues dramatically limited FAT10's pro-malignant capacity (tumor growth in vivo, aneuploidy, proliferation, migration, invasion) without affecting FAT10's other interactions. NMR structure of FAT10 domain, mutagenesis of binding interface, in vivo tumor xenograft assay, in vitro cellular assays Proceedings of the National Academy of Sciences Medium 25422469

Source papers

Stage 0 corpus · 100 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2003 Aurora B couples chromosome alignment with anaphase by targeting BubR1, Mad2, and Cenp-E to kinetochores. The Journal of cell biology 1049 12719470
2001 MAD2 haplo-insufficiency causes premature anaphase and chromosome instability in mammalian cells. Nature 637 11201745
1996 Identification of a human mitotic checkpoint gene: hsMAD2. Science (New York, N.Y.) 526 8824189
1998 The checkpoint protein MAD2 and the mitotic regulator CDC20 form a ternary complex with the anaphase-promoting complex to control anaphase initiation. Genes & development 507 9637688
2006 Mad2 overexpression promotes aneuploidy and tumorigenesis in mice. Cancer cell 506 17189715
2005 The Mad1/Mad2 complex as a template for Mad2 activation in the spindle assembly checkpoint. Current biology : CB 354 15694304
2010 Sustained Mps1 activity is required in mitosis to recruit O-Mad2 to the Mad1-C-Mad2 core complex. The Journal of cell biology 278 20624899
2002 The Mad2 spindle checkpoint protein undergoes similar major conformational changes upon binding to either Mad1 or Cdc20. Molecular cell 268 11804586
2002 Checkpoint protein BubR1 acts synergistically with Mad2 to inhibit anaphase-promoting complex. Molecular biology of the cell 265 11907259
2004 The Mad2 spindle checkpoint protein has two distinct natively folded states. Nature structural & molecular biology 256 15024386
2001 Mammalian mad2 and bub1/bubR1 recognize distinct spindle-attachment and kinetochore-tension checkpoints. Proceedings of the National Academy of Sciences of the United States of America 241 11274370
2007 The Mad2 conformational dimer: structure and implications for the spindle assembly checkpoint. Cell 205 18022367
2005 Mad2 prevents aneuploidy and premature proteolysis of cyclin B and securin during meiosis I in mouse oocytes. Genes & development 189 15655110
1997 MAD2 associates with the cyclosome/anaphase-promoting complex and inhibits its activity. Proceedings of the National Academy of Sciences of the United States of America 188 9356466
2008 Control of chromosome stability by the beta-TrCP-REST-Mad2 axis. Nature 168 18354482
2000 Structure of the Mad2 spindle assembly checkpoint protein and its interaction with Cdc20. Nature structural biology 167 10700282
2001 Mitotic checkpoint proteins HsMAD1 and HsMAD2 are associated with nuclear pore complexes in interphase. Journal of cell science 162 11181178
2001 Bub3 interaction with Mad2, Mad3 and Cdc20 is mediated by WD40 repeats and does not require intact kinetochores. The EMBO journal 159 11726501
2008 Mps1 kinase activity restrains anaphase during an unperturbed mitosis and targets Mad2 to kinetochores. The Journal of cell biology 156 18541701
2015 TRIP13 is a protein-remodeling AAA+ ATPase that catalyzes MAD2 conformation switching. eLife 142 25918846
1999 A MHC-encoded ubiquitin-like protein (FAT10) binds noncovalently to the spindle assembly checkpoint protein MAD2. Proceedings of the National Academy of Sciences of the United States of America 142 10200259
2003 Nuf2 and Hec1 are required for retention of the checkpoint proteins Mad1 and Mad2 to kinetochores. Current biology : CB 139 14654001
2002 Identification of a MAD2-binding protein, CMT2, and its role in mitosis. The EMBO journal 138 12456649
2001 Mad2 binding to Mad1 and Cdc20, rather than oligomerization, is required for the spindle checkpoint. The EMBO journal 136 11707408
2002 Significance of MAD2 expression to mitotic checkpoint control in ovarian cancer cells. Cancer research 128 11912137
2008 Protein metamorphosis: the two-state behavior of Mad2. Structure (London, England : 1993) 126 19000814
2006 Determinants of conformational dimerization of Mad2 and its inhibition by p31comet. The EMBO journal 117 16525508
2018 Mechanism for remodelling of the cell cycle checkpoint protein MAD2 by the ATPase TRIP13. Nature 111 29973720
1999 Identification of frequent impairment of the mitotic checkpoint and molecular analysis of the mitotic checkpoint genes, hsMAD2 and p55CDC, in human lung cancers. Oncogene 105 10439037
2007 Changing Mad2 levels affects chromosome segregation and spindle assembly checkpoint control in female mouse meiosis I. PloS one 99 18043727
2005 Generating chromosome instability through the simultaneous deletion of Mad2 and p53. Proceedings of the National Academy of Sciences of the United States of America 95 16055552
2006 Structural activation of Mad2 in the mitotic spindle checkpoint: the two-state Mad2 model versus the Mad2 template model. The Journal of cell biology 93 16636141
2002 Mad2 and BubR1 function in a single checkpoint pathway that responds to a loss of tension. Molecular biology of the cell 88 12388768
2008 Insights into mad2 regulation in the spindle checkpoint revealed by the crystal structure of the symmetric mad2 dimer. PLoS biology 86 18318601
2001 Molecular analyses of the mitotic checkpoint components hsMAD2, hBUB1 and hBUB3 in human cancer. International journal of cancer 86 11400114
2003 Mad2 phosphorylation regulates its association with Mad1 and the APC/C. The EMBO journal 85 12574116
1999 Mutational inactivation of mitotic checkpoint genes, hsMAD2 and hBUB1, is rare in sporadic digestive tract cancers. Japanese journal of cancer research : Gann 84 10543255
2017 MiR-200c-5p suppresses proliferation and metastasis of human hepatocellular carcinoma (HCC) via suppressing MAD2L1. Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie 73 28609841
2016 Negative Selection and Chromosome Instability Induced by Mad2 Overexpression Delay Breast Cancer but Facilitate Oncogene-Independent Outgrowth. Cell reports 67 27292643
2012 Mad2 and the APC/C compete for the same site on Cdc20 to ensure proper chromosome segregation. The Journal of cell biology 67 23007648
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
2012 Checkpoint-independent stabilization of kinetochore-microtubule attachments by Mad2 in human cells. Current biology : CB 63 22405866
2005 The roles of MAD1, MAD2 and MAD3 in meiotic progression and the segregation of nonexchange chromosomes. Nature genetics 58 15951820
2013 Spindle assembly checkpoint robustness requires Tpr-mediated regulation of Mad1/Mad2 proteostasis. The Journal of cell biology 57 24344181
2011 Overexpression of MAD2 predicts clinical outcome in primary lung cancer patients. Lung cancer (Amsterdam, Netherlands) 57 21376419
2009 Re-examination of siRNA specificity questions role of PICH and Tao1 in the spindle checkpoint and identifies Mad2 as a sensitive target for small RNAs. Chromosoma 57 19904549
2004 Intra-oocyte localization of MAD2 and its relationship with kinetochores, microtubules, and chromosomes in rat oocytes during meiosis. Biology of reproduction 56 15115722
2014 Disruption of FAT10-MAD2 binding inhibits tumor progression. Proceedings of the National Academy of Sciences of the United States of America 54 25422469
2001 Mitotic checkpoint protein hsMAD2 as a marker predicting liver metastasis of human gastric cancers. Japanese journal of cancer research : Gann 52 11572763
2020 Securin-independent regulation of separase by checkpoint-induced shugoshin-MAD2. Nature 51 32322060
2019 miR-30a-3p Targets MAD2L1 and Regulates Proliferation of Gastric Cancer Cells. OncoTargets and therapy 51 31908496
2016 Biodistribution and pharmacokinetics of Mad2 siRNA-loaded EGFR-targeted chitosan nanoparticles in cisplatin sensitive and resistant lung cancer models. Nanomedicine (London, England) 51 26980454
2012 The Mad1-Mad2 balancing act--a damaged spindle checkpoint in chromosome instability and cancer. Journal of cell science 51 23093575
2012 Functional interaction between the Arabidopsis orthologs of spindle assembly checkpoint proteins MAD1 and MAD2 and the nucleoporin NUA. Plant molecular biology 50 22457071
2003 Spindle checkpoint proteins Mad1 and Mad2 are required for cytostatic factor-mediated metaphase arrest. The Journal of cell biology 50 14691134
2006 Accumulation of Mad2-Cdc20 complex during spindle checkpoint activation requires binding of open and closed conformers of Mad2 in Saccharomyces cerevisiae. The Journal of cell biology 49 16818718
1999 Mad2 binding by phosphorylated kinetochores links error detection and checkpoint action in mitosis. Current biology : CB 48 10375530
2021 A tripartite mechanism catalyzes Mad2-Cdc20 assembly at unattached kinetochores. Science (New York, N.Y.) 47 33384372
2019 The HuR CMLD-2 inhibitor exhibits antitumor effects via MAD2 downregulation in thyroid cancer cells. Scientific reports 47 31089242
2007 Genomic instability and increased expression of BUB1B and MAD2L1 genes in ductal breast carcinoma. Cancer letters 47 17498870
2014 Mad2 and BubR1 modulates tumourigenesis and paclitaxel response in MKN45 gastric cancer cells. Cell cycle (Georgetown, Tex.) 46 25483095
2011 BUBR1 and closed MAD2 (C-MAD2) interact directly to assemble a functional mitotic checkpoint complex. The Journal of biological chemistry 46 21525009
2009 Cellular senescence induced by aberrant MAD2 levels impacts on paclitaxel responsiveness in vitro. British journal of cancer 46 19935801
2008 The Mad2 partial unfolding model: regulating mitosis through Mad2 conformational switching. The Journal of cell biology 46 19029339
2003 Spindle checkpoint component Mad2 contributes to biorientation of homologous chromosomes. Current biology : CB 46 14614824
2004 HAT cofactor Trrap regulates the mitotic checkpoint by modulation of Mad1 and Mad2 expression. The EMBO journal 44 15549134
2017 Mad2 Overexpression Uncovers a Critical Role for TRIP13 in Mitotic Exit. Cell reports 42 28564602
2014 miR-28-5p promotes chromosomal instability in VHL-associated cancers by inhibiting Mad2 translation. Cancer research 42 24491803
2010 Functional evaluation of missense variations in the human MAD1L1 and MAD2L1 genes and their impact on susceptibility to lung cancer. Journal of medical genetics 42 20516147
1993 The mitotic feedback control gene MAD2 encodes the alpha-subunit of a prenyltransferase. Nature 42 8232541
1997 Interaction of MAD2 with the carboxyl terminus of the insulin receptor but not with the IGFIR. Evidence for release from the insulin receptor after activation. The Journal of biological chemistry 40 9092546
2002 Characterization of regions in hsMAD1 needed for binding hsMAD2. A polymorphic change in an hsMAD1 leucine zipper affects MAD1-MAD2 interaction and spindle checkpoint function. The Journal of biological chemistry 39 12042300
2009 Nek2 targets the mitotic checkpoint proteins Mad2 and Cdc20: a mechanism for aneuploidy in cancer. Experimental and molecular pathology 38 20034488
2004 Transcriptional abnormality of the hsMAD2 mitotic checkpoint gene is a potential link to hepatocellular carcinogenesis. Cancer research 38 15574775
2017 Mechanistic insight into TRIP13-catalyzed Mad2 structural transition and spindle checkpoint silencing. Nature communications 36 29208896
2020 Targeting MAD2 modulates stemness and tumorigenesis in human Gastric Cancer cell lines. Theranostics 35 32863948
2015 TRIP13PCH-2 promotes Mad2 localization to unattached kinetochores in the spindle checkpoint response. The Journal of cell biology 34 26527744
2010 Phosphorylation of the spindle checkpoint protein Mad2 regulates its conformational transition. Proceedings of the National Academy of Sciences of the United States of America 34 21041666
2016 Novel Mad2-targeting miR-493-3p controls mitotic fidelity and cancer cells' sensitivity to paclitaxel. Oncotarget 32 26943585
2011 The spindle checkpoint protein Mad2 regulates APC/C activity during prometaphase and metaphase of meiosis I in Saccharomyces cerevisiae. Molecular biology of the cell 32 21697504
2015 Structure of an intermediate conformer of the spindle checkpoint protein Mad2. Proceedings of the National Academy of Sciences of the United States of America 31 26305957
2012 Kinase activity of fission yeast Mph1 is required for Mad2 and Mad3 to stably bind the anaphase promoting complex. Current biology : CB 31 22281223
2016 Distinct responses to reduplicated chromosomes require distinct Mad2 responses. eLife 30 27159240
2006 Role of MEK/ERK pathway in the MAD2-mediated cisplatin sensitivity in testicular germ cell tumour cells. British journal of cancer 30 16880791
2005 Explaining the oligomerization properties of the spindle assembly checkpoint protein Mad2. Philosophical transactions of the Royal Society of London. Series B, Biological sciences 29 15897186
2008 Aberrant MAD2 expression in soft-tissue sarcoma. Pathology international 28 18477210
2007 APC inactivation associates with abnormal mitosis completion and concomitant BUB1B/MAD2L1 up-regulation. Gastroenterology 27 17570218
2006 Depression of MAD2 inhibits apoptosis of gastric cancer cells by upregulating Bcl-2 and interfering mitochondrion pathway. Biochemical and biophysical research communications 27 16714000
2020 miR-139-5p Inhibits Lung Adenocarcinoma Cell Proliferation, Migration, and Invasion by Targeting MAD2L1. Computational and mathematical methods in medicine 26 33204298
2005 Checkpoint signalling: Mad2 conformers and signal propagation. Current biology : CB 25 15723780
2019 Too MAD or not MAD enough: The duplicitous role of the spindle assembly checkpoint protein MAD2 in cancer. Cancer letters 23 31593803
2004 MAD2 dependent mitotic checkpoint defects in tumorigenesis and tumor cell death: a double edged sword. Cell cycle (Georgetown, Tex.) 23 15254432
2011 Closed MAD2 (C-MAD2) is selectively incorporated into the mitotic checkpoint complex (MCC). Cell cycle (Georgetown, Tex.) 22 22037211
2010 Recruitment of Cdc20 to the kinetochore requires BubR1 but not Mad2 in Drosophila melanogaster. Molecular and cellular biology 22 20421417
2009 p31comet Induces cellular senescence through p21 accumulation and Mad2 disruption. Molecular cancer research : MCR 22 19276188
2009 Expression of mitotic checkpoint proteins BUB1B and MAD2L1 in salivary duct carcinomas. Journal of oral pathology & medicine : official publication of the International Association of Oral Pathologists and the American Academy of Oral Pathology 22 20040022
2015 MAD1L1 Arg558His and MAD2L1 Leu84Met interaction with smoking increase the risk of colorectal cancer. Scientific reports 21 26183163
2013 Chk1-Mad2 interaction: a crosslink between the DNA damage checkpoint and the mitotic spindle checkpoint. Cell cycle (Georgetown, Tex.) 21 23454898
2007 RNA Interference as a tool to study the function of MAD2 in mouse oocyte meiotic maturation. Molecular reproduction and development 21 16924662
2023 MAD2L1 is transcriptionally regulated by TEAD4 and promotes cell proliferation and migration in colorectal cancer. Cancer gene therapy 20 36599972