1. Structural Biology and Molecular Biophysics

The structure of the yeast Ctf3 complex

  1. Stephen M Hinshaw  Is a corresponding author
  2. Andrew N Dates
  3. Stephen C Harrison  Is a corresponding author
  1. Howard Hughes Medical Institute, Harvard Medical School, United States
  2. Harvard University, United States
https://doi.org/10.7554/eLife.48215
Share this article
Cite this article
  1. Stephen M Hinshaw
  2. Andrew N Dates
  3. Stephen C Harrison
(2019)
The structure of the yeast Ctf3 complex
eLife 8:e48215.
https://doi.org/10.7554/eLife.48215
  2. Download BibTeX
  3. Download .RIS

Abstract

Kinetochores are the chromosomal attachment points for spindle microtubules. They are also signaling hubs that control major cell cycle transitions and coordinate chromosome folding. Most well-studied eukaryotes rely on a conserved set of factors, which are divided among two loosely-defined groups, for these functions. Outer kinetochore proteins contact microtubules or regulate this contact directly. Inner kinetochore proteins designate the kinetochore assembly site by recognizing a specialized nucleosome containing the H3 variant Cse4/CENP-A. We previously determined the structure, resolved by cryo-electron microscopy (cryo-EM), of the yeast Ctf19 complex (Ctf19c, homologous to the vertebrate CCAN), providing a high-resolution view of inner kinetochore architecture (Hinshaw and Harrison, 2019). We now extend these observations by reporting a near-atomic model of the Ctf3 complex, the outermost Ctf19c sub-assembly seen in our original cryo-EM density. The model is sufficiently well-determined by the new data to enable molecular interpretation of Ctf3 recruitment and function.

Data availability

The final Ctf3c cryo-EM map and the soft mask used for three-dimensional refinements of fully unbinned particles have been deposited with accession code EMD-20200. The refined Ctf3c model has been deposited with accession code 6OUA.

The following previously published data sets were used

Article and author information

Author details

  1. Stephen M Hinshaw

    Howard Hughes Medical Institute, Harvard Medical School, Boston, United States
    For correspondence
    hinshaw@crystal.harvard.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-4215-5206

  2. Andrew N Dates

    Program in Chemical Biology, Harvard University, Cambridge, United States
    Competing interests
    The authors declare that no competing interests exist.

  3. Stephen C Harrison

    Howard Hughes Medical Institute, Harvard Medical School, Boston, United States
    For correspondence
    harrison@crystal.harvard.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7215-9393

Funding

Howard Hughes Medical Institute

  • Stephen M Hinshaw

Howard Hughes Medical Institute

  • Stephen C Harrison

Helen Hay Whitney Foundation

  • Stephen M Hinshaw

National Institutes of Health (5T32GM095450-08)

  • Andrew N Dates

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

Reviewing Editor

  1. Andrea Musacchio, Max Planck Institute of Molecular Physiology, Germany

Version history

  1. Received: May 9, 2019
  2. Accepted: June 12, 2019
  3. Accepted Manuscript published: June 13, 2019 (version 1)
  4. Version of Record published: July 1, 2019 (version 2)

Copyright

© 2019, Hinshaw 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.

Metrics

  • 1,831
    views
  • 309
    downloads
  • 15
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Stephen M Hinshaw
  2. Andrew N Dates
  3. Stephen C Harrison
(2019)
The structure of the yeast Ctf3 complex
eLife 8:e48215.
https://doi.org/10.7554/eLife.48215

Share this article

https://doi.org/10.7554/eLife.48215

Categories and tags

Research organism

Further reading

    1. Structural Biology and Molecular Biophysics

    The structure of the Ctf19c/CCAN from budding yeast

    Stephen M Hinshaw, Stephen C Harrison
    Research Article Updated

    Eukaryotic kinetochores connect spindlemicrotubules to chromosomal centromeres. A group of proteins called the Ctf19 complex (Ctf19c) in yeast and the constitutive centromere associated network (CCAN) in other organisms creates the foundation of a kinetochore. The Ctf19c/CCAN influences the timing of kinetochore assembly, sets its location by associating with a specialized nucleosome containing the histone H3 variant Cse4/CENP-A, and determines the organization of the microtubule attachment apparatus. We present here the structure of a reconstituted 13-subunit Ctf19c determined by cryo-electron microscopy at ~4 Å resolution. The structure accounts for known and inferred contacts with the Cse4 nucleosome and for an observed assembly hierarchy. We describe its implications for establishment of kinetochores and for their regulation by kinases throughout the cell cycle.

    1. Structural Biology and Molecular Biophysics

    Plasticity of the proteasome-targeting signal Fat10 enhances substrate degradation

    Hitendra Negi, Aravind Ravichandran ... Ranabir Das
    Research Article

    The proteasome controls levels of most cellular proteins, and its activity is regulated under stress, quiescence, and inflammation. However, factors determining the proteasomal degradation rate remain poorly understood. Proteasome substrates are conjugated with small proteins (tags) like ubiquitin and Fat10 to target them to the proteasome. It is unclear if the structural plasticity of proteasome-targeting tags can influence substrate degradation. Fat10 is upregulated during inflammation, and its substrates undergo rapid proteasomal degradation. We report that the degradation rate of Fat10 substrates critically depends on the structural plasticity of Fat10. While the ubiquitin tag is recycled at the proteasome, Fat10 is degraded with the substrate. Our results suggest significantly lower thermodynamic stability and faster mechanical unfolding in Fat10 compared to ubiquitin. Long-range salt bridges are absent in the Fat10 structure, creating a plastic protein with partially unstructured regions suitable for proteasome engagement. Fat10 plasticity destabilizes substrates significantly and creates partially unstructured regions in the substrate to enhance degradation. NMR-relaxation-derived order parameters and temperature dependence of chemical shifts identify the Fat10-induced partially unstructured regions in the substrate, which correlated excellently to Fat10-substrate contacts, suggesting that the tag-substrate collision destabilizes the substrate. These results highlight a strong dependence of proteasomal degradation on the structural plasticity and thermodynamic properties of the proteasome-targeting tags.

    1. Biochemistry and Chemical Biology
    2. Structural Biology and Molecular Biophysics

    Special Issue: Allosteric regulation of kinase activity

    Amy H Andreotti, Volker Dötsch
    Editorial

    The articles in this special issue highlight how modern cellular, biochemical, biophysical and computational techniques are allowing deeper and more detailed studies of allosteric kinase regulation.