Three-dimensional structure of kinetochore-fibers in human mitotic spindles
- Technische Universität Dresden, Germany
- Harvard University, United States
- Zuse Institute Berlin, Germany
Abstract
During cell division, kinetochore microtubules (KMTs) provide a physical linkage between the chromosomes and the rest of the spindle. KMTs in mammalian cells are organized into bundles, so-called kinetochore-fibers (k-fibers), but the ultrastructure of these fibers is currently not well characterized. Here we show by large-scale electron tomography that each k-fiber in HeLa cells in metaphase is composed of approximately nine KMTs, only half of which reach the spindle pole. Our comprehensive reconstructions allowed us to analyze the three-dimensional (3D) morphology of k-fibers and their surrounding MTs in detail. We found that k-fibers exhibit remarkable variation in circumference and KMT density along their length, with the pole-proximal side showing a broadening. Extending our structural analysis then to other MTs in the spindle, we further observed that the association of KMTs with non-KMTs predominantly occurs in the spindle pole regions. Our 3D reconstructions have implications for KMT growth and k-fiber self-organization models as covered in a parallel publication applying complementary live-cell imaging in combination with biophysical modeling (Conway et al., 2022). Finally, we also introduce a new visualization tool allowing an interactive display of our 3D spindle data that will serve as a resource for further structural studies on mitosis in human cells.
Data availability
All Datasets were uploaded and ara available in OpaRA server:http://doi.org/10.25532/OPARA-128; http://doi.org/10.25532/OPARA-177he code used to perform quantitative analysis and visualization of MT organization in spindles has been uploaded to the GitHub repository and is available as open access under the GPL v3.0 license:https://github.com/RRobert92/ASGA; https://github.com/RRobert92/ASGA_3DViewer
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HeLa metaphase spindle tomographic data sets and analysisOpaRA, doi.org/10.25532/OPARA-128.
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HeLa metaphase spindle tomographic data sets and analysis for z-expansion dataOpaRA, doi.org/10.25532/OPARA-177.
Article and author information
Author details
Thomas Müller-Reichert
Funding
Deutsche Forschungsgemeinschaft (MU 1423/8-2)
- Robert Kiewisz
- Gunar Fabig
- Thomas Müller-Reichert
Horizon 2020 Framework Programme (675737)
- Robert Kiewisz
- Thomas Müller-Reichert
Harvard University
- William Conway
- Daniel J Needleman
Nick Simons Foundation (1764269)
- William Conway
- 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
- Adèle L Marston, University of Edinburgh, United Kingdom
Version history
- Received: November 10, 2021
- Preprint posted: November 13, 2021 (view preprint)
- Accepted: July 24, 2022
- Accepted Manuscript published: July 27, 2022 (version 1)
- Version of Record published: August 10, 2022 (version 2)
- Version of Record updated: August 15, 2022 (version 3)
Copyright
© 2022, Kiewisz 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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Three-dimensional structure of kinetochore-fibers in human mitotic spindles
eLife 11:e75459.
https://doi.org/10.7554/eLife.75459
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