Abstract

In all living organisms, it is essential to transmit genetic information faithfully to the next generation. The SMC-ParAB-parS system is widely employed for chromosome segregation in bacteria. A DNA-binding protein ParB nucleates on parS sites and must associate with neighboring DNA, a process known as spreading, to enable efficient chromosome segregation. Despite its importance, how the initial few ParB molecules nucleating at parS sites recruit hundreds of further ParB to spread is not fully understood. Here, we reconstitute a parS-dependent ParB spreading event using purified proteins from Caulobacter crescentus and show that CTP is required for spreading. We further show that ParB spreading requires a closed DNA substrate, and a DNA-binding transcriptional regulator can act as a roadblock to attenuate spreading unidirectionally in vitro. Our biochemical reconstitutions recapitulate many observed in vivo properties of ParB and opens up avenues to investigate the interactions between ParB-parS with ParA and SMC.

Data availability

No deep sequencing data or X-ray crystallography data were generated during this study. All other data (BLI, uncropped gel images etc...) are included in the manuscript, figures, and source data files.

Article and author information

Author details

  1. Adam SB Jalal

    Department of Molecular Microbiology, John Innes Centre, Norwich, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  2. Ngat T Tran

    Department of Molecular Microbiology, John Innes Centre, Norwich, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  3. Tung BK Le

    Department of Molecular Microbiology, John Innes Centre, Norwich, United Kingdom
    For correspondence
    tung.le@jic.ac.uk
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-4764-8851

Funding

Biotechnology and Biological Sciences Research Council (BB/P018165/1)

  • Tung BK Le

Royal Society (UF140053)

  • Tung BK Le

Biotechnology and Biological Sciences Research Council (BBS/E/J/000PR9791)

  • Ngat T Tran
  • Tung BK Le

Royal Society (RG150448)

  • Adam SB Jalal
  • Tung BK Le

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

Reviewing Editor

  1. Anthony G Vecchiarelli, University of Michigan, United States

Version history

  1. Received: November 11, 2019
  2. Accepted: February 19, 2020
  3. Accepted Manuscript published: February 20, 2020 (version 1)
  4. Version of Record published: March 3, 2020 (version 2)

Copyright

© 2020, Jalal 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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  1. Adam SB Jalal
  2. Ngat T Tran
  3. Tung BK Le
(2020)
ParB spreading on DNA requires cytidine triphosphate in vitro
eLife 9:e53515.
https://doi.org/10.7554/eLife.53515

Share this article

https://doi.org/10.7554/eLife.53515

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