1. Chromosomes and Gene Expression
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Recombination occurs within minutes of replication blockage by RTS1 producing restarted forks that are prone to collapse

  1. Michael O Nguyen
  2. Manisha Jalan
  3. Carl A Morrow
  4. Fekret Osman
  5. Matthew C Whitby  Is a corresponding author
  1. University of Oxford, United Kingdom
Research Article
  • Cited 26
  • Views 2,030
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Cite this article as: eLife 2015;4:e04539 doi: 10.7554/eLife.04539

Abstract

The completion of genome duplication during the cell cycle is threatened by the presence of replication fork barriers (RFBs). Following collision with a RFB replication proteins can dissociate from the stalled fork (fork collapse) rendering it incapable of further DNA synthesis unless recombination intervenes to restart replication. We use time-lapse microscopy and genetic assays to show that recombination is initiated within ~10 minutes of replication fork blockage at a site-specific barrier in fission yeast, leading to a restarted fork within ~60 minutes, which is only prevented/curtailed by the arrival of the opposing replication fork. The restarted fork is susceptible to further collapse causing hyper-recombination downstream of the barrier. Surprisingly, in our system fork restart is unnecessary for maintaining cell viability. Seemingly the risk of failing to complete replication prior to mitosis is sufficient to warrant the induction of recombination even though it can cause deleterious genetic change.

Article and author information

Author details

  1. Michael O Nguyen

    Department of Biochemistry, University of Oxford, Oxford, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  2. Manisha Jalan

    Department of Biochemistry, University of Oxford, Oxford, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  3. Carl A Morrow

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

    Department of Biochemistry, University of Oxford, Oxford, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  5. Matthew C Whitby

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

Reviewing Editor

  1. Stephen C Kowalczykowski, University of California, Davis, United States

Publication history

  1. Received: August 28, 2014
  2. Accepted: March 24, 2015
  3. Accepted Manuscript published: March 25, 2015 (version 1)
  4. Version of Record published: April 23, 2015 (version 2)

Copyright

© 2015, Nguyen 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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Further reading

    1. Chromosomes and Gene Expression
    Manisha Jalan et al.
    Research Advance Updated

    Homologous recombination helps ensure the timely completion of genome duplication by restarting collapsed replication forks. However, this beneficial function is not without risk as replication restarted by homologous recombination is prone to template switching (TS) that can generate deleterious genome rearrangements associated with diseases such as cancer. Previously we established an assay for studying TS in Schizosaccharomyces pombe (Nguyen et al., 2015). Here, we show that TS is detected up to 75 kb downstream of a collapsed replication fork and can be triggered by head-on collision between the restarted fork and RNA Polymerase III transcription. The Pif1 DNA helicase, Pfh1, promotes efficient restart and also suppresses TS. A further three conserved helicases (Fbh1, Rqh1 and Srs2) strongly suppress TS, but there is no change in TS frequency in cells lacking Fml1 or Mus81. We discuss how these factors likely influence TS.

    1. Chromosomes and Gene Expression
    2. Structural Biology and Molecular Biophysics
    Samuel Bowerman et al.
    Research Article

    Eukaryotes and many archaea package their DNA with histones. While the four eukaryotic histones wrap ~147 DNA base pairs into nucleosomes, archaeal histones form 'nucleosome-like' complexes that continuously wind between 60 - 500 base pairs of DNA ('archaeasomes'), suggested by crystal contacts and analysis of cellular chromatin. Solution structures of large archaeasomes (>90 DNA base pairs) have never been directly observed. Here, we utilize molecular dynamics simulations, analytical ultracentrifugation, and cryoEM to structurally characterize the solution state of archaeasomes on longer DNA. Simulations reveal dynamics of increased accessibility without disruption of DNA-binding or tetramerization interfaces. Mg2+ concentration influences compaction, and cryoEM densities illustrate that DNA is wrapped in consecutive substates arranged 90o out-of-plane with one another. Without ATP-dependent remodelers, archaea may leverage these inherent dynamics to balance chromatin packing and accessibility.