Mad2 binding to Mad1 and Cdc20, rather than oligomerization, is required for the spindle checkpoint.
Mad2 is a key component of the spindle checkpoint, a device that controls the fidelity of chromosome segregation in mitosis. The ability of Mad2 to form oligomers in vitro has been correlated with its ability to block the cell cycle upon injection into Xenopus embryos. Here we show that Mad2 forms incompatible complexes with Mad1 and Cdc20, neither of which requires Mad2 oligomerization. A monomeric point mutant of Mad2 can sustain a cell cycle arrest of comparable strength to that of the wild-type protein. We show that the interaction of Mad2 with Mad1 is crucial for the localization of Mad2 to kinetochores, where Mad2 interacts with Cdc20. We propose a model that features the kinetochore as a 'folding factory' for the formation of a Mad2-Cdc20 complex endowed with inhibitory activity on the anaphase promoting complex.
Sironi
L
L
Department of Experimental Oncology, European Institute of Oncology, Via Ripamonti 435, 20141 Milan, Italy.
Melixetian
M
M
Faretta
M
M
Prosperini
E
E
Helin
K
K
Musacchio
A
A
D000705:Anaphase; D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D001665:Binding Sites; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002352:Carrier Proteins; D064199:Cdc20 Proteins; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000378:metabolism*; D002850:Chromatography, Gel; D018076:DNA, Complementary / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D005656:Fungal Proteins / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D018386:Kinetochores / Q000378:metabolism; D064246:Mad2 Proteins; D051379:Mice; D008856:Microscopy, Fluorescence; D008938:Mitosis; D008954:Models, Biological; D016297:Mutagenesis, Site-Directed; D009687:Nuclear Proteins; D010455:Peptides / Q000737:chemistry; D010750:Phosphoproteins / Q000378:metabolism*; D010957:Plasmids / Q000378:metabolism; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D011233:Precipitin Tests; D011485:Protein Binding; D011994:Recombinant Proteins / Q000378:metabolism; D012097:Repressor Proteins / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D012965:Sodium Chloride / Q000494:pharmacology; D014162:Transfection; D014508:Urea / Q000494:pharmacology
D000911:Antibodies, Monoclonal; C577495:CDC20 protein, S cerevisiae; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D064199:Cdc20 Proteins; D018797:Cell Cycle Proteins; D018076:DNA, Complementary; D005656:Fungal Proteins; C501382:MAD1 protein, S cerevisiae; C501384:MAD1L1 protein, human; C577695:MAD2 protein, S cerevisiae; C501385:Mad1l1 protein, mouse; D064246:Mad2 Proteins; C577698:Mad2l1 protein, mouse; C577699:Mad2l2 protein, mouse; D009687:Nuclear Proteins; D010455:Peptides; D010750:Phosphoproteins; D011994:Recombinant Proteins; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D012965:Sodium Chloride; D014508:Urea