ParB spreading on DNA requires cytidine triphosphate in vitro
- John Innes Centre, United Kingdom
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.
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Author details
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.
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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ParB spreading on DNA requires cytidine triphosphate in vitro
eLife 9:e53515.
https://doi.org/10.7554/eLife.53515
