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CENP-E forms a link between attachment of spindle microtubules to kinetochores and the mitotic checkpoint

Nature Cell Biology volume 2, pages 484–491 (2000)Cite this article

Abstract

Here we show that suppression of synthesis of the microtubule motor CENP-E (centromere-associated protein E), a component of the kinetochore corona fibres of mammalian centromeres, yields chromosomes that are chronically mono-orientated, with spindles that are flattened along the plane of the substrate. Despite apparently normal microtubule numbers and the continued presence at kinetochores of other microtubule motors, spindle poles fragment in the absence of CENP-E, which implicates this protein in delivery of components from kinetochores to poles. CENP-E represents a link between attachment of spindle microtubules and the mitotic checkpoint signalling cascade, as depletion of this motor leads to profound checkpoint activation, whereas immunoprecipitation reveals a nearly stoichiometric association of CENP-E with the checkpoint kinase BubR1 during mitosis.

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Figure 1: Antisense suppression of CENP-E induces long-term mitotic arrest.
Figure 2: Loss of CENP-E does not affect spindle assembly, but causes chromosome misalignment.
Figure 3: Reduced capture or stability of kinetochore-associated microtubules in the absence of CENP-E.
Figure 4: Identification of CENP-E binding partners in normal cells.
Figure 5: Suppression of CENP-E results in failure to silence the mitotic checkpoint.
Figure 6: CENP-E binding partners remain associated with the kinetochore in the absence of CENP-E.
Figure 7: Model for the mitosis-related functions of CENP-E.

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References

  1. Brinkley, B. R. & Stubblefield, E. The fine structure of the kinetochore of a mammalian cell in vitro. Chromosoma 19, 28–43 (1966).

    Article  CAS  Google Scholar 

  2. Nicklas, R. B. The motor for poleward chromosome movement in anaphase is in or near the kinetochore . J. Cell Biol. 109, 2245-2255 (1989).

    Article  Google Scholar 

  3. 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).

    Article  CAS  Google Scholar 

  4. Hyman, A. A. & Mitchison, T. J. Two different microtubule-based motor activities with opposite polarities in kinetochores. Nature 351, 206–211 ( 1991).

    Article  CAS  Google Scholar 

  5. Roos, U-P. Light and electron microscopy of rat kangaroo cells in mitosis. III. Patterns of chromosome behavior during prometaphase. J. Cell Biol. 54, 363 –385 (1976).

    CAS  Google Scholar 

  6. Bajer, A. S. Functional autonomy of monopolar spindle and evidence for oscillatory movement in mitosis. J. Cell Biol. 93, 33– 48 (1982).

    Article  CAS  Google Scholar 

  7. Alexander, S. P. & Rieder, C. L. Chromosome motion during attachment to the vertebrate spindle: initial saltatory-like behavior of chromosomes and quantitative analysis of force production by nascent kinetochore fibers. J. Cell Biol. 113, 805– 815 (1991).

    Article  CAS  Google Scholar 

  8. Skibbens, R. V., Skeen, V. P. & Salmon, E. D. Directional instability of kinetochore motility during chromosome congression and segregation in mitotic newt lung cells: a push-pull mechanism. J. Cell Biol. 122, 859– 875 (1993).

    Article  CAS  Google Scholar 

  9. Khodjakov, A. & Rieder, C. L. Kinetochores moving away from their associated pole do not exert a significant pushing force on the chromosome . J. Cell Biol. 135, 315– 327 (1996).

    Article  CAS  Google Scholar 

  10. Rieder, C. L., Cole, R. W., Khodjakov, A. & Sluder, G. The checkpoint delaying anaphase in response to chromosome monoorientation is mediated by an inhibitory signal produced by unattached kinetochores. J. Cell Biol. 130, 941–948 (1995).

    Article  CAS  Google Scholar 

  11. Li, R. & Murray, A. W. Feedback control of mitosis in budding yeast. Cell 66, 519– 531 (1991).

    Article  CAS  Google Scholar 

  12. Hoyt, M. A., Totis, L. & Roberts, B. T. S. cerevisiae genes required for cell cycle arrest in response to loss of microtubule function. Cell 66, 507–517 (1991).

    Google Scholar 

  13. Li, X. & Nicklas, R. B. Mitotic forces control a cell-cycle checkpoint. Nature 373, 630– 632 (1995).

    Article  CAS  Google Scholar 

  14. Zhang, D. & Nicklas, R. B. ‘Anaphase’ and cytokinesis in the absence of chromosomes. Nature 382, 466–468 (1996).

    Article  CAS  Google Scholar 

  15. Chen, R. H., Waters, J. C., Salmon, E. D. & Murray, A. W. Association of spindle assembly checkpoint component XMAD2 with unattached kinetochores. Science 274, 242– 246 (1996).

    Article  CAS  Google Scholar 

  16. Li, Y. & Benezra, R. Identification of a human mitotic checkpoint gene: hsMAD2. Science 274, 246 –248 (1996).

    Article  CAS  Google Scholar 

  17. Taylor, S. S., Ha, E. & McKeon, F. The human homologue of Bub3 is required for kinetochore localization of Bub1 and a Mad3/Bub1-related protein kinase. J. Cell Biol. 142, 1–11 (1998).

    Article  CAS  Google Scholar 

  18. Martinez-Exposito, M. J., Kaplan, K. B., Copeland, J. & Sorger, P. K. Retention of the BUB3 checkpoint protein on lagging chromosomes. Proc. Natl Acad. Sci. USA 96, 8493– 8498 (1999).

    Article  CAS  Google Scholar 

  19. Taylor, S. S. & McKeon, F. Kinetochore localization of murine Bub1 is required for normal mitotic timing and checkpoint response to spindle damage. Cell 89, 727–735 (1997).

    Article  CAS  Google Scholar 

  20. Chan, G. K. T., Jablonski, S. A., Sudakin, V., Hittle, J. C. & Yen, T. J. Human BUBR1 is a mitotic checkpoint kinase that monitors CENP-E functions and binds the cyclosome/APC. J. Cell Biol. 146, 941–954 (1999).

    Article  CAS  Google Scholar 

  21. Wood, K. W., Sakowicz, R., Goldstein, L. S. B. & Cleveland, D. W. CENP-E is a plus end-directed kinetochore motor required for metaphase chromosome alignment. Cell 91, 357– 366 (1997).

    Article  CAS  Google Scholar 

  22. Brown, K. D., Coulson, R. M., Yen, T. J. & Cleveland, D. W. Cyclin-like accumulation and loss of the putative kinetochore motor CENP-E results from coupling continuous synthesis with specific degradation at the end of mitosis. J. Cell Biol. 125, 1303– 1312 (1994).

    Article  CAS  Google Scholar 

  23. Lombillo, V. A., Nislow, C., Yen, T. J., Gelfand, V. I. & McIntosh, J. R. Antibodies to the kinesin motor domain and CENP-E inhibit microtubule depolymerization-dependent motion of chromosomes in vitro. J. Cell Biol. 128, 107–115 (1995).

    Article  CAS  Google Scholar 

  24. Yen, T. J. et al. CENP-E, a novel human centromere-associated protein required for progression from metaphase to anaphase. EMBO J. 10, 1245–1254 (1991).

    Article  CAS  Google Scholar 

  25. Duesbery, N. S. et al. CENP-E is an essential kinetochore motor in maturing oocytes and is masked during mos-dependent, cell cycle arrest at metaphase II. Proc. Natl Acad. Sci. USA 94, 9165– 9170 (1997).

    Article  CAS  Google Scholar 

  26. Schaar, B. T., Chan, G. K. T., Maddox, P., Salmon, E. D. & Yen, T. J. CENP-E function at kinetochores is essential for chromosome alignment. J. Cell Biol. 139 , 1373–1382 (1997).

    Article  CAS  Google Scholar 

  27. Chan, G. K. T., Schaar, B. T. & Yen, T. J. Characterization of the kinetochore binding domain of CENP-E reveals interactions with the kinetochore proteins CENP-F and hBUBR1 . J. Cell Biol. 143, 49– 63 (1998).

    Article  CAS  Google Scholar 

  28. Waters, J. C., Skibbens, R. V. & Salmon, E. D. Oscillating mitotic newt lung cell kinetochores are, on average, under tension and rarely push. J. Cell Sci. 109, 2823–2831 (1996).

    CAS  PubMed  Google Scholar 

  29. Steuer, E. R., Wordeman, L., Schroer, T. A. & Sheetz, M. P. Localization of cytoplasmic dynein to mitotic spindles and kinetochores. Nature 345, 266–268 ( 1990).

    Article  CAS  Google Scholar 

  30. Pfarr, C. M. et al. Cytoplasmic dynein is localized to kinetochores during mitosis . Nature 345, 263–265 (1990).

    Article  CAS  Google Scholar 

  31. Wordeman, L. & Mitchison, T. J. Identification and partial characterization of mitotic centromere-associated kinesin, a kinesin-related protein that associates with centromeres during mitosis. J. Cell Biol. 128, 95–104 ( 1995).

    Article  CAS  Google Scholar 

  32. Yao, X., Anderson, K. L. & Cleveland, D. W. The microtubule-dependent motor centromere-associated protein E (CENP-E) is an integral component of kinetochore corona fibers that link centromeres to spindle microtubules. J. Cell Biol. 139, 435–447 (1997).

    Article  CAS  Google Scholar 

  33. Rieder, C. L. The structure of the cold-stable kinetochore fiber in metaphase PtK 1 cells . Chromosoma 84, 145–158 (1981).

    Article  CAS  Google Scholar 

  34. Cassimeris, L., Rieder, C. L., Rupp, G. & Salmon, E. D. Stability of microtubule attachment to metaphase kinetochores in PtK1 cells. J. Cell Sci. 96, 9–15 ( 1990).

    PubMed  Google Scholar 

  35. Starr, D. A., Williams, B. C., Hays, T. S. & Goldberg, M. L. ZW10 helps recruit dynactin and dynein to the kinetochore. J. Cell Biol. 142, 763–774 ( 1998).

    Article  CAS  Google Scholar 

  36. Merdes, A., Ramyar, K., Vechio, J. D. & Cleveland, D. W. A complex of NuMA and cytoplasmic dynein is essential for mitotic spindle assembly. Cell 87, 447– 458 (1996).

    Article  CAS  Google Scholar 

  37. Merdes, A., Heald, R., Samejima, K., Earnshaw, W. C. & Cleveland, D. W. Formation of spindle poles by dynein/dynactin-dependent transport of NuMA. J. Cell Biol. (submitted).

  38. Walczak, C. E., Mitchison, T. J. & Desai, A. XKCM1: a Xenopus kinesin-related protein that regulates microtubule dynamics during mitotic spindle assembly. Cell 84, 37–47 (1996).

    Article  CAS  Google Scholar 

  39. Desai, A., Verma, S., Mitchison, T. J. & Walczak, C. E. Kin I kinesins are microtubule-destabilizing enzymes. Cell 96, 69–78 (1999).

    Article  CAS  Google Scholar 

  40. Maney, T., Hunter, A. W., Wagenbach, M. & Wordeman, L. Mitotic centromere-associated kinesin is important for anaphase chromosome segregation. J. Cell Biol. 142, 787– 801 (1998).

    Article  CAS  Google Scholar 

  41. Vaisberg, E. A., Koonce, M. P. & McIntosh, J. R. Cytoplasmic dynein plays a role in mammalian mitotic spindle formation. J. Cell Biol. 123, 849 –858 (1993).

    Article  CAS  Google Scholar 

  42. Echeverri, C. J., Paschal, B. M., Vaughan, K. T. & Vallee, R. B. Molecular characterization of the 50-kD subunit of dynactin reveals function for the complex in chromosome alignment and spindle organization during mitosis . J. Cell Biol. 132, 617– 633 (1996).

    Article  CAS  Google Scholar 

  43. Jablonski, S. A., Chan, G. K. T., Cooke, C. A., Earnshaw, W. C. & Yen, T. J. The hBUB1 and hBUBR1 kinases sequentially assemble onto kinetochores during prophase with hBUBR1 concentrating at the kinetochore plates in mitosis. Chromosoma 107, 386–396 (1998).

    Article  CAS  Google Scholar 

  44. Caceres, A. & Kosik, K. S. Inhibition of neurite polarity by tau antisense oligonucleotides in primary cerebellar neurons. Nature 343, 461–463 ( 1990).

    Article  CAS  Google Scholar 

  45. Brown, K. D., Wood, K. W. & Cleveland, D. W. The kinesin-like protein CENP-E is kinetochore-associated throughout poleward chromosome segregation during anaphase-A. J. Cell Sci. 109, 961–969 (1996).

    CAS  PubMed  Google Scholar 

  46. Kilmartin, J. V., Wright, B. & Milstein, C. Rat monoclonal antitubulin antibodies derived by using a new nonsecreting rat cell line. J. Cell Biol. 93, 576–582 (1982).

    Article  CAS  Google Scholar 

  47. Evan, G. I., Lewis, G. K., Ramsay, G. & Bishop, J. M. Isolation of monoclonal antibodies specific for human c-myc proto-oncogene product. Mol. Cell Biol. 5, 3610–3616 (1985).

    Article  CAS  Google Scholar 

  48. Fang, G., Yu, H. & Kirschner, M. W. The checkpoint protein MAD2 and the mitotic regulator CDC20 form a ternary complex with the anaphase-promoting complex to control anaphase initiation. Genes Dev. 12, 1871 –1883 (1998).

    Article  CAS  Google Scholar 

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Acknowledgements

This work was supported by a grant from the National Institute of Health (GM29513) to D.W.C. Salary support for D.W.C. is provided by the Ludwig Institute for Cancer Research. X.Y. was supported by postdoctoral fellowships from the Bank of America Giannini Foundation and the American Cancer Society (California Division), and by a Career Development Award from the Howard Hughes Medical Institute/University of Wisconsin.

Correspondence and requests for materials should be addressed to D.W.C.

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Authors and Affiliations

  1. Department of Physiology, University of Wisconsin, Madison, 53706, Wisconsin, USA

    Xuebiao Yao & Yun Zheng

  2. Ludwig Institute for Cancer Research, University of California at San Diego, 9500 Gilman Drive,, La Jolla, 92093-0660, California, USA

    Xuebiao Yao, Ariane Abrieu & Don W. Cleveland

  3. Departments of Medicine and Neuroscience, University of California at San Diego, 9500 Gilman Drive, La Jolla, 92093-0660, California, USA

    Don W. Cleveland

  4. Department of Cell Biology, Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, 92037,, California, USA

    Kevin F. Sullivan

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Correspondence to Don W. Cleveland.

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Yao, X., Abrieu, A., Zheng, Y. et al. CENP-E forms a link between attachment of spindle microtubules to kinetochores and the mitotic checkpoint. Nat Cell Biol 2, 484–491 (2000). https://doi.org/10.1038/35019518

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