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<p>Transcription-associated torsional stress presents a continuous threat to genome integrity that needs to be tightly controlled. Here we present a method for the visualization of genome maintenance events at sites of active transcription, which builds on a previously developed reporter system that allows for the detection of nascent, MS2-repeat containing transcripts with a YFP-tagged MS2 coat protein. We describe steps to monitor Topoisomerase 1 engagement as a result of transcription-associated topological stress in both S phase and non-S phase cells, and detail procedures to visualize TOP1 cleavage complex formation and concomitant DNA repair factor engagement. We expect this system to provide a powerful means to study effectors of the transcription-associated topological stress response.</p>

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Visualizing DNA repair factor recruitment at sites of transcription in single cells

  • 期刊论文

  • 出版模式：

    开放获取

  • 发表日期：

    2026年1月3日

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  • Tianyi Guan,
  • Yuepeng Shi,
  • Tae-Hee Lee,
  • Philipp Oberdoerffer

摘要

Transcription-associated torsional stress presents a continuous threat to genome integrity that needs to be tightly controlled. Here we present a method for the visualization of genome maintenance events at sites of active transcription, which builds on a previously developed reporter system that allows for the detection of nascent, MS2-repeat containing transcripts with a YFP-tagged MS2 coat protein. We describe steps to monitor Topoisomerase 1 engagement as a result of transcription-associated topological stress in both S phase and non-S phase cells, and detail procedures to visualize TOP1 cleavage complex formation and concomitant DNA repair factor engagement. We expect this system to provide a powerful means to study effectors of the transcription-associated topological stress response.