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Interaction between transcribing RNA polymerase and topoisomerase I prevents R-loop formation in E. coli

Author

Listed:
  • Dmitry Sutormin

    (Skolkovo Institute of Science and Technology
    Institute of Gene Biology RAS)

  • Alina Galivondzhyan

    (Skolkovo Institute of Science and Technology
    Lomonosov Moscow States University)

  • Olga Musharova

    (Skolkovo Institute of Science and Technology
    Institute of Molecular Genetics, National Research Centre “Kurchatov Institute”)

  • Dmitrii Travin

    (Skolkovo Institute of Science and Technology
    Institute of Gene Biology RAS)

  • Anastasiia Rusanova

    (Institute of Gene Biology RAS)

  • Kseniya Obraztsova

    (Rowan University School of Osteopathic Medicine
    University of Pennsylvania, Perelman School of Medicine, Department of Medicine)

  • Sergei Borukhov

    (Rowan University School of Osteopathic Medicine)

  • Konstantin Severinov

    (Skolkovo Institute of Science and Technology
    Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Institute of Gene Biology, RAS
    Waksman University for Microbiology)

Abstract

Bacterial topoisomerase I (TopoI) removes excessive negative supercoiling and is thought to relax DNA molecules during transcription, replication and other processes. Using ChIP-Seq, we show that TopoI of Escherichia coli (EcTopoI) is colocalized, genome-wide, with transcribing RNA polymerase (RNAP). Treatment with transcription elongation inhibitor rifampicin leads to EcTopoI relocation to promoter regions, where RNAP also accumulates. When a 14 kDa RNAP-binding EcTopoI C-terminal domain (CTD) is overexpressed, colocalization of EcTopoI and RNAP along the transcription units is reduced. Pull-down experiments directly show that the two enzymes interact in vivo. Using ChIP-Seq and Topo-Seq, we demonstrate that EcTopoI is enriched upstream (within up to 12-15 kb) of highly-active transcription units, indicating that EcTopoI relaxes negative supercoiling generated by transcription. Uncoupling of the RNAP:EcTopoI interaction by either overexpression of EcTopoI competitor (CTD or inactive EcTopoI Y319F mutant) or deletion of EcTopoI domains involved in the interaction is toxic for cells and leads to excessive negative plasmid supercoiling. Moreover, uncoupling of the RNAP:EcTopoI interaction leads to R-loops accumulation genome-wide, indicating that this interaction is required for prevention of R-loops formation.

Suggested Citation

  • Dmitry Sutormin & Alina Galivondzhyan & Olga Musharova & Dmitrii Travin & Anastasiia Rusanova & Kseniya Obraztsova & Sergei Borukhov & Konstantin Severinov, 2022. "Interaction between transcribing RNA polymerase and topoisomerase I prevents R-loop formation in E. coli," Nature Communications, Nature, vol. 13(1), pages 1-19, December.
  • Handle: RePEc:nat:natcom:v:13:y:2022:i:1:d:10.1038_s41467-022-32106-5
    DOI: 10.1038/s41467-022-32106-5
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    References listed on IDEAS

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    1. Britney Martinez & Binod K. Bharati & Vitaly Epshtein & Evgeny Nudler, 2022. "Pervasive Transcription-coupled DNA repair in E. coli," Nature Communications, Nature, vol. 13(1), pages 1-10, December.
    2. Gareth Butland & José Manuel Peregrín-Alvarez & Joyce Li & Wehong Yang & Xiaochun Yang & Veronica Canadien & Andrei Starostine & Dawn Richards & Bryan Beattie & Nevan Krogan & Michael Davey & John Par, 2005. "Interaction network containing conserved and essential protein complexes in Escherichia coli," Nature, Nature, vol. 433(7025), pages 531-537, February.
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    1. Maxime Mistretta & Mena Cimino & Pascal Campagne & Stevenn Volant & Etienne Kornobis & Olivier Hebert & Christophe Rochais & Patrick Dallemagne & Cédric Lecoutey & Camille Tisnerat & Alban Lepailleur , 2024. "Dynamic microfluidic single-cell screening identifies pheno-tuning compounds to potentiate tuberculosis therapy," Nature Communications, Nature, vol. 15(1), pages 1-22, December.

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