Abstract
Error-free chromosome segregation depends on the precise regulation of phosphorylation to stabilize kinetochore–microtubule attachments (K-fibres) on sister chromatids that have attached to opposite spindle poles (bi-oriented)1. In many instances, phosphorylation correlates with K-fibre destabilization2,3,4,5,6,7. Consistent with this, multiple kinases, including Aurora B and Plk1, are enriched at kinetochores of mal-oriented chromosomes when compared with bi-oriented chromosomes, which have stable attachments2,8. Paradoxically, however, these kinases also target to prometaphase chromosomes that have not yet established spindle attachments and it is therefore unclear how kinetochore–microtubule interactions can be stabilized when kinase levels are high. Here we show that the generation of stable K-fibres depends on the B56-PP2A phosphatase, which is enriched at centromeres/kinetochores of unattached chromosomes. When B56-PP2A is depleted, K-fibres are destabilized and chromosomes fail to align at the spindle equator. Strikingly, B56-PP2A depletion increases the level of phosphorylation of Aurora B and Plk1 kinetochore substrates as well as Plk1 recruitment to kinetochores. Consistent with increased substrate phosphorylation, we find that chemical inhibition of Aurora or Plk1 restores K-fibres in B56-PP2A-depleted cells. Our findings reveal that PP2A, an essential tumour suppressor9, tunes the balance of phosphorylation to promote chromosome–spindle interactions during cell division.
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References
Musacchio, A. & Salmon, E. D. The spindle-assembly checkpoint in space and time. Nat. Rev. Mol. Cell Biol. 8, 379–393 (2007).
Ahonen, L. J. et al. Polo-like kinase 1 creates the tension-sensing 3F3/2 phosphoepitope and modulates the association of spindle-checkpoint proteins at kinetochores. Curr. Biol. 15, 1078–1089 (2005).
Cheeseman, I. M. et al. Phospho-regulation of kinetochore-microtubule attachments by the Aurora kinase Ipl1p. Cell 111, 163–172 (2002).
Lampson, M. A., Renduchitala, K., Khodjakov, A. & Kapoor, T. M. Correcting improper chromosome-spindle attachments during cell division. Nat. Cell Biol. 6, 232–237 (2004).
Pinsky, B. A., Kung, C., Shokat, K. M. & Biggins, S. The Ipl1-Aurora protein kinase activates the spindle checkpoint by creating unattached kinetochores. Nat. Cell Biol. 8, 78–83 (2006).
Welburn, J. P. et al. Aurora B phosphorylates spatially distinct targets to differentially regulate the kinetochore-microtubule interface. Mol. Cell 38, 383–392 (2010).
Wong, O. K. & Fang, G. Plx1 is the 3F3/2 kinase responsible for targeting spindle checkpoint proteins to kinetochores. J. Cell Biol. 170, 709–719 (2005).
Salimian, K. J. et al. Feedback control in sensing chromosome biorientation by the aurora B kinase. Curr. Biol. 21, 1158–1165 (2011).
Westermarck, J. & Hahn, W. C. Multiple pathways regulated by the tumor suppressor PP2A in transformation. Trends Mol. Med. 14, 152–160 (2008).
Ubersax, J. A. & Ferrell, J. E. Jr Mechanisms of specificity in protein phosphorylation. Nat. Rev. Mol. Cell Biol. 8, 530–541 (2007).
Trinkle-Mulcahy, L. et al. Time-lapse imaging reveals dynamic relocalization of PP1γ throughout the mammalian cell cycle. Mol. Biol. Cell 14, 107–117 (2003).
Kitajima, T. S. et al. Shugoshin collaborates with protein phosphatase 2A to protect cohesin. Nature 441, 46–52 (2006).
Riedel, C. G. et al. Protein phosphatase 2A protects centromeric sister chromatid cohesion during meiosis I. Nature 441, 53–61 (2006).
Tang, Z. et al. PP2A is required for centromeric localization of Sgo1 and proper chromosome segregation. Dev. Cell 10, 575–585 (2006).
Liu, D. et al. Regulated targeting of protein phosphatase 1 to the outer kinetochore by KNL1 opposes Aurora B kinase. J. Cell Biol. 188, 809–820 (2010).
Posch, M. et al. Sds22 regulates aurora B activity and microtubule-kinetochore interactions at mitosis. J. Cell Biol. 191, 61–74 (2010).
Janssens, V. & Goris, J. Protein phosphatase 2A: a highly regulated family of serine/threonine phosphatases implicated in cell growth and signalling. Biochem. J. 353, 417–439 (2001).
Baharians, Z. & Schonthal, A. H. Autoregulation of protein phosphatase type 2A expression. J. Biol. Chem. 273, 19019–19024 (1998).
Hutchins, J. R. et al. Systematic analysis of human protein complexes identifies chromosome segregation proteins. Science 328, 593–599 (2010).
Hoffman, D. B., Pearson, C. G., Yen, T. J., Howell, B. J. & Salmon, E. D. Microtubule-dependent changes in assembly of microtubule motor proteins and mitotic spindle checkpoint proteins at PtK1 kinetochores. Mol. Biol. Cell 12, 1995–2009 (2001).
Waters, J. C., Chen, R. H., Murray, A. W. & Salmon, E. D. Localization of Mad2 to kinetochores depends on microtubule attachment, not tension. J. Cell Biol. 141, 1181–1191 (1998).
Neumann, B. et al. Phenotypic profiling of the human genome by time-lapse microscopy reveals cell division genes. Nature 464, 721–727 (2010).
Cho, U. S. & Xu, W. Crystal structure of a protein phosphatase 2A heterotrimeric holoenzyme. Nature 445, 53–57 (2007).
Xu, Y. et al. Structure of the protein phosphatase 2A holoenzyme. Cell 127, 1239–1251 (2006).
Rieder, C. L. The structure of the cold-stable kinetochore fiber in metaphase PtK1 cells. Chromosoma 84, 145–158 (1981).
Kitajima, T. S., Kawashima, S. A. & Watanabe, Y. The conserved kinetochore protein shugoshin protects centromeric cohesion during meiosis. Nature 427, 510–517 (2004).
Rabitsch, K. P. et al. Two fission yeast homologs of Drosophila Mei-S332 are required for chromosome segregation during meiosis I and II. Curr. Biol. 14, 287–301 (2004).
Salic, A., Waters, J. C. & Mitchison, T. J. Vertebrate shugoshin links sister centromere cohesion and kinetochore microtubule stability in mitosis. Cell 118, 567–578 (2004).
Cheeseman, I. M., Chappie, J. S., Wilson-Kubalek, E. M. & Desai, A. The conserved KMN network constitutes the core microtubule-binding site of the kinetochore. Cell 127, 983–997 (2006).
Lampson, M. A. & Cheeseman, I. M. Sensing centromere tension: Aurora B and the regulation of kinetochore function. Trends Cell Biol. 21, 133–140 (2010).
Ditchfield, C. et al. Aurora B couples chromosome alignment with anaphase by targeting BubR1, Mad2, and Cenp-E to kinetochores. J. Cell Biol. 161, 267–280 (2003).
Hauf, S. et al. The small molecule Hesperadin reveals a role for Aurora B in correcting kinetochore-microtubule attachment and in maintaining the spindle assembly checkpoint. J. Cell Biol. 161, 281–294 (2003).
Lenart, P. et al. The small-molecule inhibitor BI 2536 reveals novel insights into mitotic roles of polo-like kinase 1. Curr. Biol. 17, 304–315 (2007).
Elowe, S., Hummer, S., Uldschmid, A., Li, X. & Nigg, E. A. Tension-sensitive Plk1 phosphorylation on BubR1 regulates the stability of kinetochore microtubule interactions. Genes Dev. 21, 2205–2219 (2007).
Elia, A. E., Cantley, L. C. & Yaffe, M. B. Proteomic screen finds pSer/pThr-binding domain localizing Plk1 to mitotic substrates. Science 299, 1228–1231 (2003).
Rieder, C. L. & Alexander, S. P. Kinetochores are transported poleward along a single astral microtubule during chromosome attachment to the spindle in newt lung cells. J. Cell Biol. 110, 81–95 (1990).
McEwen, B. F., Heagle, A. B., Cassels, G. O., Buttle, K. F. & Rieder, C. L. Kinetochore fiber maturation in PtK1 cells and its implications for the mechanisms of chromosome congression and anaphase onset. J. Cell Biol. 137, 1567–1580 (1997).
Liu, D., Vader, G., Vromans, M. J., Lampson, M. A. & Lens, S. M. Sensing chromosome bi-orientation by spatial separation of aurora B kinase from kinetochore substrates. Science 323, 1350–1353 (2009).
Rosenberg, J. S., Cross, F. R. & Funabiki, H. KNL1/Spc105 recruits PP1 to silence the spindle assembly checkpoint. Curr. Biol. 21, 942–947 (2011).
Ruediger, R., Pham, H. T. & Walter, G. Disruption of protein phosphatase 2A subunit interaction in human cancers with mutations in the A α subunit gene. Oncogene 20, 10–15 (2001).
Lengauer, C., Kinzler, K. W. & Vogelstein, B. Genetic instabilities in human cancers. Nature 396, 643–649 (1998).
Kline, S. L., Cheeseman, I. M., Hori, T., Fukagawa, T. & Desai, A. The human Mis12 complex is required for kinetochore assembly and proper chromosome segregation. J. Cell Biol. 173, 9–17 (2006).
Acknowledgements
We thank I. Cheeseman (MIT, Cambridge, Massachusetts, USA), A. Desai (University of California San Diego, San Diego, California, USA), S. Elowe (Centre de Recherche du CHUQ, Quebec, Canada) and H. Yu (UT Southwestern Medical Center, Dallas, Texas, USA) for antibodies and W. Harper for plasmids. E.A.F. was supported by a Damon Runyon Cancer Research postdoctoral fellowship (DRG 1936-07) and the Charles H. Revson Foundation. This work was supported by NIH GM65933 (T.M.K.).
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E.A.F. and T.M.K. designed the experiments and wrote the paper. E.A.F. carried out essentially all of the experiments. M.M. contributed to the live-cell imaging.
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Foley, E., Maldonado, M. & Kapoor, T. Formation of stable attachments between kinetochores and microtubules depends on the B56-PP2A phosphatase. Nat Cell Biol 13, 1265–1271 (2011). https://doi.org/10.1038/ncb2327
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DOI: https://doi.org/10.1038/ncb2327
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