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Huang, H. M. et al. Phosphorylation sites in BubR1 that regulate kinetochore attachment, tension, and mitotic exit. J. Cell Biol. 183, 667-680

Fox Chase Cancer Center, Philadelphia, PA 19111, USA.
The Journal of Cell Biology (Impact Factor: 9.69). 12/2008; 183(4):667-80. DOI: 10.1083/jcb.200805163
Source: PubMed

ABSTRACT BubR1 kinase is essential for the mitotic checkpoint and also for kinetochores to establish microtubule attachments. In this study, we report that BubR1 is phosphorylated in mitosis on four residues that differ from sites recently reported to be phosphorylated by Plk1 (Elowe, S., S. Hummer, A. Uldschmid, X. Li, and E.A. Nigg. 2007. Genes Dev. 21:2205-2219; Matsumura, S., F. Toyoshima, and E. Nishida. 2007. J. Biol. Chem. 282:15217-15227). S670, the most conserved residue, is phosphorylated at kinetochores at the onset of mitosis and dephosphorylated before anaphase onset. Unlike the Plk1-dependent S676 phosphorylation, S670 phosphorylation is sensitive to microtubule attachments but not to kinetochore tension. Functionally, phosphorylation of S670 is essential for error correction and for kinetochores with end-on attachments to establish tension. Furthermore, in vitro data suggest that the phosphorylation status of BubR1 is important for checkpoint inhibition of the anaphase-promoting complex/cyclosome. Finally, RNA interference experiments show that Mps1 is a major but not the exclusive kinase that specifies BubR1 phosphorylation in vivo. The combined data suggest that BubR1 may be an effector of multiple kinases that are involved in discrete aspects of kinetochore attachments and checkpoint regulation.

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Available from: Tim J Yen, May 06, 2015
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    • "670–720) are required for binding B56-PP2A. BUBR1 is hyperphosphorylated during mitosis, and mitotic kinases Cdk1, Plk1 and Mps1 have been shown to phosphorylate S670 (Elowe et al., 2007; Huang et al., 2008) (in response to lack of kinetochore–microtubule attachment) and S676 (Elowe et al., 2007) (in response to lack of tension) within this evolutionary conserved second motif of BUBR1. During the preparation of this manuscript, Kops and his colleague show that phosphorylation within this second motif by Plk1 promotes interaction of BUBR1 with B56α-PP2A to counter excessive Aurora B activity at the kinetochores (Suijkerbuijk et al., 2012). "
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    ABSTRACT: BUBR1 is a mitotic phosphoprotein essential for the maintenance of chromosome stability by promoting chromosome congression and proper kinetochore-microtubule (K-fiber) attachment, but the underlying mechanism(s) has remained elusive. Here we identify BUBR1 as a binding partner of the B56 family of Protein Phosphatase 2A regulatory subunits. The interaction between BUBR1 and the B56 family is required for chromosome congression, since point mutations in BUBR1 that block B56 binding abolish chromosome congression. The BUBR1:B56-PP2A complex opposes Aurora B kinase activity, since loss of the complex can be reverted by inhibiting Aurora B. Importantly, we show that the failure of BUBR1 to recruit B56-PP2A also contributes to the chromosome congression defects found in cells derived from patients with the Mosaic Variegated Aneuploidy (MVA) syndrome. Together, we propose that B56-PP2A is a key mediator of BUBR1's role in chromosome congression and functions by antagonizing Aurora B activity at the kinetochore for establishing stable kinetochore-microtubule attachment at the metaphase plate.
    Biology Open 05/2013; 2(5):479-486. DOI:10.1242/bio.20134051 · 2.42 Impact Factor
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    • "BUB1B/Phos:4 does not exist in frogs because only one of the phosphorylation sites (human Ser-670, frog Ser-649) is conserved (Figure 3; BUB1B/Phos:4, red residues). However, it is noteworthy that mutation of Ser-670 alone in the human BUB1B protein produced phenotypes nearly as severe as mutating all of the BUB1B/Phos:4 sites, indicating that Ser-670 is a critical phosphorylation site (Huang et al., 2008; Elowe et al., 2010). Thus, it is possible that frog has a BUB1B form phosphorylated on Ser-649 that plays a similar role to BUB1B/Phos:4 in humans. "
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    ABSTRACT: As a member of the Open Biomedical Ontologies (OBO) foundry, the Protein Ontology (PRO) provides an ontological representation of protein forms and complexes and their relationships. Annotations in PRO can be assigned to individual protein forms and complexes, each distinguishable down to the level of post-translational modification, thereby allowing for a more precise depiction of protein function than is possible with annotations to the gene as a whole. Moreover, PRO is fully interoperable with other OBO ontologies and integrates knowledge from other protein-centric resources such as UniProt and Reactome. Here we demonstrate the value of the PRO framework in the investigation of the spindle checkpoint, a highly conserved biological process that relies extensively on protein modification and protein complex formation. The spindle checkpoint maintains genomic integrity by monitoring the attachment of chromosomes to spindle microtubules and delaying cell cycle progression until the spindle is fully assembled. Using PRO in conjunction with other bioinformatics tools, we explored the cross-species conservation of spindle checkpoint proteins, including phosphorylated forms and complexes; studied the impact of phosphorylation on spindle checkpoint function; and examined the interactions of spindle checkpoint proteins with the kinetochore, the site of checkpoint activation. Our approach can be generalized to any biological process of interest.
    Frontiers in Genetics 04/2013; 4:62. DOI:10.3389/fgene.2013.00062
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    • "The two groups identified a highly phosphorylated conserved domain of BubR1 (residues 665–682), referred to as KARD (Suijkerbuijk et al., 2012b), whose integrity is necessary for the ability of BubR1 to promote stable K–MT attachments. Within the KARD domain, there are at least three phosphorylation sites: two (S670 and S676) described above (Elowe et al., 2007, 2010; Huang et al., 2008) plus a third site (T680), also phosphorylated by Plk1 on tensionless kinetochores, as is S676 (Suijkerbuijk et al., 2012b). Expressing the triple S670A, S676A, and T680A BubR1 mutant in HeLa cells abolished chromosome alignment, but chemically inhibiting Aurora B activity largely restored proper chromosome alignment in these cells. "
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    ABSTRACT: BubR1 is a critical component of the spindle assembly checkpoint, the surveillance mechanism that helps maintain the high fidelity of mitotic chromosome segregation by preventing cells from initiating anaphase if one or more kinetochores are not attached to the spindle. BubR1 also helps promote the establishment of stable kinetochore-microtubule attachments during prometaphase. In this chapter, we review the structure, functions, and regulation of BubR1 in these "classical roles" at the kinetochore. We discuss its recruitment to kinetochores, its assembly into the inhibitor of anaphase progression, and the importance of its posttranslational modifications. We also consider the evidence for its participation in other roles beyond mitosis, such as the meiosis-specific processes of recombination and prophase arrest of the first meiotic division, the cellular response to DNA damage, and in the regulation of centrosome and basal body function. Finally, studies are presented linking BubR1 dysfunction or misregulation to aging and human disease, particularly cancer.
    International review of cell and molecular biology 01/2013; 306:223-73. DOI:10.1016/B978-0-12-407694-5.00006-7 · 4.52 Impact Factor
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