The Caenorhabditis elegans protein SAS-5 forms large oligomeric assemblies critical for centriole formation

  1. Kacper B Rogala
  2. Nicola J Dynes
  3. Georgios N Hatzopoulos
  4. Jun Yan
  5. Sheng Kai Pong
  6. Carol V Robinson
  7. Charlotte M Deane
  8. Pierre Gönczy
  9. Ioannis Vakonakis  Is a corresponding author
  1. University of Oxford, United Kingdom
  2. Swiss Federal Institute of Technology, Switzerland

Abstract

Centrioles are microtubule-based organelles crucial for cell division, sensing and motility. In C. elegans, the onset of centriole formation requires notably the proteins SAS-5 and SAS-6, which have functional homologs across eukaryotic evolution. Whereas the molecular architecture of SAS-6 and its role in initiating centriole formation are well understood, the mechanisms by which SAS-5 and its relatives function is unclear. Here, we combine biophysical and structural analysis to uncover the architecture of SAS-5 and examine its functional implications in vivo. Our work reveals that two distinct self-associating domains are necessary to form higher-order oligomers of SAS-5: a trimeric coiled coil and a novel globular dimeric Implico domain. Disruption of either domain leads to centriole duplication failure in worm embryos, indicating that large SAS-5 assemblies are necessary for function in vivo.

Article and author information

Author details

  1. Kacper B Rogala

    Department of Biochemistry, University of Oxford, Oxford, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  2. Nicola J Dynes

    Swiss Institute for Experimental Cancer Research, School of Life Sciences, Swiss Federal Institute of Technology, Lausanne, Switzerland
    Competing interests
    The authors declare that no competing interests exist.
  3. Georgios N Hatzopoulos

    Department of Biochemistry, University of Oxford, Oxford, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  4. Jun Yan

    Department of Chemistry, University of Oxford, Oxford, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  5. Sheng Kai Pong

    Swiss Institute for Experimental Cancer Research, School of Life Sciences, Swiss Federal Institute of Technology, Lausanne, Switzerland
    Competing interests
    The authors declare that no competing interests exist.
  6. Carol V Robinson

    Department of Chemistry, University of Oxford, Oxford, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  7. Charlotte M Deane

    Department of Statistics, University of Oxford, Oxford, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  8. Pierre Gönczy

    Swiss Institute for Experimental Cancer Research, School of Life Sciences, Swiss Federal Institute of Technology, Lausanne, Switzerland
    Competing interests
    The authors declare that no competing interests exist.
  9. Ioannis Vakonakis

    Department of Biochemistry, University of Oxford, Oxford, United Kingdom
    For correspondence
    ioannis.vakonakis@bioch.ox.ac.uk
    Competing interests
    The authors declare that no competing interests exist.

Reviewing Editor

  1. Anthony A Hyman, Max Planck Institute of Molecular Cell Biology and Genetics, Germany

Version history

  1. Received: March 10, 2015
  2. Accepted: May 28, 2015
  3. Accepted Manuscript published: May 29, 2015 (version 1)
  4. Version of Record published: June 18, 2015 (version 2)

Copyright

© 2015, Rogala 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

  • 2,192
    Page views
  • 513
    Downloads
  • 31
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, PubMed Central, Scopus.

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. Kacper B Rogala
  2. Nicola J Dynes
  3. Georgios N Hatzopoulos
  4. Jun Yan
  5. Sheng Kai Pong
  6. Carol V Robinson
  7. Charlotte M Deane
  8. Pierre Gönczy
  9. Ioannis Vakonakis
(2015)
The Caenorhabditis elegans protein SAS-5 forms large oligomeric assemblies critical for centriole formation
eLife 4:e07410.
https://doi.org/10.7554/eLife.07410

Share this article

https://doi.org/10.7554/eLife.07410

Further reading

    1. Structural Biology and Molecular Biophysics
    Parameswaran Hariharan, Yuqi Shi ... Lan Guan
    Research Article

    While many 3D structures of cation-coupled transporters have been determined, the mechanistic details governing the obligatory coupling and functional regulations still remain elusive. The bacterial melibiose transporter (MelB) is a prototype of major facilitator superfamily transporters. With a conformation-selective nanobody, we determined a low-sugar affinity inward-facing Na+-bound cryoEM structure. The available outward-facing sugar-bound structures showed that the N- and C-terminal residues of the inner barrier contribute to the sugar selectivity. The inward-open conformation shows that the sugar selectivity pocket is also broken when the inner barrier is broken. Isothermal titration calorimetry measurements revealed that this inward-facing conformation trapped by this nanobody exhibited a greatly decreased sugar-binding affinity, suggesting the mechanisms for substrate intracellular release and accumulation. While the inner/outer barrier shift directly regulates the sugar-binding affinity, it has little or no effect on the cation binding, which is supported by molecular dynamics simulations. Furthermore, the hydron/deuterium exchange mass spectrometry analyses allowed us to identify dynamic regions; some regions are involved in the functionally important inner barrier-specific salt-bridge network, which indicates their critical roles in the barrier switching mechanisms for transport. These complementary results provided structural and dynamic insights into the mobile barrier mechanism for cation-coupled symport.

    1. Structural Biology and Molecular Biophysics
    Fouad Ouasti, Maxime Audin ... Francoise Ochsenbein
    Research Article

    Genome and epigenome integrity in eukaryotes depends on the proper coupling of histone deposition with DNA synthesis. This process relies on the evolutionary conserved histone chaperone CAF-1 for which the links between structure and functions are still a puzzle. While studies of the Saccharomyces cerevisiae CAF-1 complex enabled to propose a model for the histone deposition mechanism, we still lack a framework to demonstrate its generality and in particular, how its interaction with the polymerase accessory factor PCNA is operating. Here, we reconstituted a complete SpCAF-1 from fission yeast. We characterized its dynamic structure using NMR, SAXS and molecular modeling together with in vitro and in vivo functional studies on rationally designed interaction mutants. Importantly, we identify the unfolded nature of the acidic domain which folds up when binding to histones. We also show how the long KER helix mediates DNA binding and stimulates SpCAF-1 association with PCNA. Our study highlights how the organization of CAF-1 comprising both disordered regions and folded modules enables the dynamics of multiple interactions to promote synthesis-coupled histone deposition essential for its DNA replication, heterochromatin maintenance, and genome stability functions.