title: Bimodal activation of BubR1 by Bub3 sustains mitotic checkpoint signaling.
abstract: The mitotic checkpoint (also known as the spindle assembly checkpoint) prevents premature anaphase onset through generation of an inhibitor of the E3 ubiquitin ligase APC/C, whose ubiquitination of cyclin B and securin targets them for degradation. Combining in vitro reconstitution and cell-based assays, we now identify dual mechanisms through which Bub3 promotes mitotic checkpoint signaling. Bub3 enhances signaling at unattached kinetochores not only by facilitating binding of BubR1 but also by enhancing Cdc20 recruitment to kinetochores mediated by BubR1's internal Cdc20 binding site. Downstream of kinetochore-produced complexes, Bub3 promotes binding of BubR1's conserved, amino terminal Cdc20 binding domain to a site in Cdc20 that becomes exposed by initial Mad2 binding. This latter Bub3-stimulated event generates the final mitotic checkpoint complex of Bub3-BubR1-Cdc20 that selectively inhibits ubiquitination of securin and cyclin B by APC/C(Cdc20). Thus, Bub3 promotes two distinct BubR1-Cdc20 interactions, involving each of the two Cdc20 binding sites of BubR1 and acting at unattached kinetochores or cytoplasmically, respectively, to facilitate production of the mitotic checkpoint inhibitor.
authors.affiliation: Ludwig Institute for Cancer Research and Departments of Cellular and Molecular Medicine and dcleveland@ucsd.edu j3han@ucsd.edu.
authors.forename: Joo Seok
authors.initials: JS
authors.lastname: Han
authors.affiliation: Ludwig Institute for Cancer Research and Departments of Cellular and Molecular Medicine and.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Vitre
authors.affiliation: Ludwig Institute for Cancer Research and Departments of Cellular and Molecular Medicine and.
authors.forename: Daniele
authors.initials: D
authors.lastname: Fachinetti
authors.affiliation: Ludwig Institute for Cancer Research and Departments of Cellular and Molecular Medicine and Medicine, University of California, San Diego, La Jolla, CA 92093 dcleveland@ucsd.edu j3han@ucsd.edu.
authors.forename: Don W
authors.initials: DW
authors.lastname: Cleveland
mesh_terms: D064173:Anaphase-Promoting Complex-Cyclosome / Q000378:metabolism; D001665:Binding Sites / Q000235:genetics; D064199:Cdc20 Proteins / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D056744:Cyclin B1 / Q000378:metabolism; D036681:Fluorescence Recovery After Photobleaching; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D018386:Kinetochores / Q000378:metabolism; D059566:M Phase Cell Cycle Checkpoints / Q000235:genetics / Q000502:physiology*; D008856:Microscopy, Fluorescence; D009154:Mutation; D000075223:Poly-ADP-Ribose Binding Proteins; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology*; D059008:Time-Lapse Imaging; D054875:Ubiquitination
chemical_list: C489444:BUB3 protein, human; D064199:Cdc20 Proteins; D018797:Cell Cycle Proteins; D056744:Cyclin B1; D000075223:Poly-ADP-Ribose Binding Proteins; D049452:Green Fluorescent Proteins; C577488:CDC20 protein, human; D064173:Anaphase-Promoting Complex-Cyclosome; C584277:BUB1 protein, human; D017346:Protein Serine-Threonine Kinases