Reconstitution reveals two paths of force transmission through the kinetochore
- University of Washington, United States
- Genentech Inc, United States
Abstract
Partitioning duplicated chromosomes equally between daughter cells is a microtubule-mediated process essential to eukaryotic life. A multi-protein machine, the kinetochore, drives chromosome segregation by coupling the chromosomes to dynamic microtubule tips, even as the tips grow and shrink through the gain and loss of subunits. The kinetochore must harness, transmit, and sense mitotic forces, as a lack of tension signals incorrect chromosome-microtubule attachment and precipitates error correction mechanisms. But though the field has arrived at a 'parts list' of dozens of kinetochore proteins organized into subcomplexes, the path of force transmission through these components has remained unclear. Here we report reconstitution of functional Saccharomyces cerevisiae kinetochore assemblies from recombinantly expressed proteins. The reconstituted kinetochores are capable of self-assembling in vitro, coupling centromeric nucleosomes to dynamic microtubules, and withstanding mitotically relevant forces. They reveal two distinct pathways of force transmission and Ndc80c recruitment.
Data availability
All data analyses during this study are included in the manuscript and supporting files. Source data files have been provided for Figures 2-5 and their supplements.
Article and author information
Author details
Grace Elizabeth Hamilton
Funding
National Institutes of Health (Training Grant in Molecular Biophysics T32 GM008268)
- Grace Elizabeth Hamilton
National Institutes of Health (R01 GM040506)
- Trisha N Davis
National Institutes of Health (R35 GM130293)
- Trisha N Davis
National Institutes of Health (R01 GM079373)
- Charles L Asbury
National Institutes of Health (R35 GM134842)
- Charles L Asbury
Genentech
- Yoana N Dimitrova
Genentech
- Cameron L Noland
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2020, Hamilton 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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Reconstitution reveals two paths of force transmission through the kinetochore
eLife 9:e56582.
https://doi.org/10.7554/eLife.56582
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