1. Cell Biology
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Measuring NDC80 binding reveals the molecular basis of tension-dependent kinetochore-microtubule attachments

  1. Tae Yeon Yoo  Is a corresponding author
  2. Jeong-Mo Choi
  3. William Conway
  4. Che-Hang Yu
  5. Rohit V Pappu
  6. Daniel J Needleman
  1. Harvard University, United States
  2. Washington University in St Louis, United States
Research Article
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Cite this article as: eLife 2018;7:e36392 doi: 10.7554/eLife.36392

Abstract

Proper kinetochore-microtubule attachments, mediated by the NDC80 complex, are required for error-free chromosome segregation. Erroneous attachments are corrected by the tension dependence of kinetochore-microtubule interactions. Here, we present a method, based on fluorescence lifetime imaging microscopy and Förster resonance energy transfer, to quantitatively measure the fraction of NDC80 complexes bound to microtubules at individual kinetochores in living human cells. We found that NDC80 binding is modulated in a chromosome autonomous fashion over prometaphase and metaphase, and is predominantly regulated by centromere tension. We show that this tension dependency requires phosphorylation of the N-terminal tail of Hec1, a component of the NDC80 complex, and the proper localization of Aurora B kinase, which modulates NDC80 binding. Our results lead to a mathematical model of the molecular basis of tension-dependent NDC80 binding to kinetochore microtubules in vivo.

Data availability

- All microscopy image data and data points in the presented plots have been deposited in Dryad (DOI: https://doi.org/10.5061/dryad.14rr125)- Analysis codes are deposited in Github, where doi's are provided in the manuscript.

The following data sets were generated

Article and author information

Author details

  1. Tae Yeon Yoo

    Department of Molecular and Cellular Biology, Harvard University, Cambridge, United States
    For correspondence
    taeyeon_yoo@hms.harvard.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8145-1051
  2. Jeong-Mo Choi

    Department of Biomedical Engineering, Washington University in St Louis, St Louis, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-2656-4851
  3. William Conway

    Department of Physics, Harvard University, Cambridge, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Che-Hang Yu

    John A Paulson School of Engineering and Applied Sciences, Harvard University, Cambridge, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0353-9752
  5. Rohit V Pappu

    Department of Biomedical Engineering, Washington University in St Louis, St Louis, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-2568-1378
  6. Daniel J Needleman

    Department of Molecular and Cellular Biology, Harvard University, Cambridge, United States
    Competing interests
    The authors declare that no competing interests exist.

Funding

National Science Foundation (DBI-0959721)

  • Daniel J Needleman

National Institutes of Health (R01NS056114)

  • Rohit V Pappu

National Science Foundation (DMR-0820484)

  • Daniel J Needleman

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Trisha N Davis, University of Washington, United States

Publication history

  1. Received: March 5, 2018
  2. Accepted: July 24, 2018
  3. Accepted Manuscript published: July 25, 2018 (version 1)
  4. Version of Record published: August 13, 2018 (version 2)
  5. Version of Record updated: August 15, 2018 (version 3)
  6. Version of Record updated: August 23, 2018 (version 4)

Copyright

© 2018, Yoo et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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