Tight bending of the Ndc80 complex provides intrinsic regulation of its binding to microtubules

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Abstract

Regulation of the outer kinetochore complex Ndc80 is essential to ensure correct kinetochore-microtubule attachments during mitosis. Here we present a novel mechanism of regulation that is intrinsic to its structure; tight bending of the Ndc80 complex inhibits its microtubule binding. Using single molecule Förster resonance energy transfer (FRET), we show that the S. cerevisiae Ndc80 complex can fluctuate between straight and bent forms, and that binding of the complex to microtubules selects for straightened forms. The loop region of the complex enables its bent conformation, as deletion of the loop promotes straightening. In addition, the kinetochore complex MIND enhances microtubule binding by opposing the tightly bent, auto-inhibited conformation of the Ndc80 complex. We suggest that prior to its assembly at the kinetochore, the Ndc80 complex interchanges between bent (auto-inhibited) and open conformations. Once assembled, its association with MIND stabilizes the Ndc80 complex in a straightened form for higher affinity microtubule binding.

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All data generated or analysed during this study are included in the manuscript and supporting files. Source data files have been provided for Figures 1, 2, 3, 4 and 5, as well as Figure 1-figure supplement 1, Figure 3-figure supplement 1, Figure 4-figure supplement 1 and Figure 5-figure supplement 1.

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Emily Anne Scarborough

Department of Biochemistry, University of Washington, Seattle, United States

Competing interests
The authors declare that no competing interests exist.
Trisha N Davis

Department of Biochemistry, University of Washington, Seattle, United States

For correspondence
tdavis@uw.edu

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0003-4797-3152
Charles L Asbury

Department of Physiology and Biophysics, University of Washington, Seattle, United States

For correspondence
casbury@uw.edu

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-0143-5394

Funding

National Institute of General Medical Sciences (T32 GM007270)

Emily Anne Scarborough

National Institute of General Medical Sciences (R01 GM040506)

Trisha N Davis

National Institute of General Medical Sciences (R01 GM079373)

Charles L Asbury

National Institute of General Medical Sciences (P01 GM105537)

Charles L Asbury

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

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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.