Ultra-soft X-ray system for imaging the early cellular responses to X-ray induced DNA damage

Nucleic Acids Res. 2019 Sep 26;47(17):e100. doi: 10.1093/nar/gkz609.


The majority of the proteins involved in processing of DNA double-strand breaks (DSBs) accumulate at the damage sites. Real-time imaging and analysis of these processes, triggered by the so-called microirradiation using UV lasers or heavy particle beams, yielded valuable insights into the underlying DSB repair mechanisms. To study the temporal organization of DSB repair responses triggered by a more clinically-relevant DNA damaging agent, we developed a system coined X-ray multi-microbeam microscope (XM3), capable of simultaneous high dose-rate (micro)irradiation of large numbers of cells with ultra-soft X-rays and imaging of the ensuing cellular responses. Using this setup, we analyzed the changes in real-time kinetics of MRE11, MDC1, RNF8, RNF168 and 53BP1-proteins involved in the signaling axis of mammalian DSB repair-in response to X-ray and UV laser-induced DNA damage, in non-cancerous and cancer cells and in the presence or absence of a photosensitizer. Our results reveal, for the first time, the kinetics of DSB signaling triggered by X-ray microirradiation and establish XM3 as a powerful platform for real-time analysis of cellular DSB repair responses.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptor Proteins, Signal Transducing / metabolism
  • Animals
  • Cell Cycle Proteins / metabolism
  • Cell Line
  • DNA Breaks, Double-Stranded*
  • DNA Repair*
  • DNA-Binding Proteins / metabolism*
  • Humans
  • MRE11 Homologue Protein
  • Microscopy, Electron, Scanning
  • Osteosarcoma / metabolism
  • Pigment Epithelium of Eye / metabolism
  • Signal Transduction
  • Time-Lapse Imaging / methods*
  • Tumor Suppressor p53-Binding Protein 1 / metabolism
  • Ubiquitin-Protein Ligases / metabolism
  • Ultraviolet Rays
  • X-Rays*


  • Adaptor Proteins, Signal Transducing
  • Cell Cycle Proteins
  • DNA-Binding Proteins
  • MDC1 protein, human
  • MRE11 protein, human
  • RNF8 protein, human
  • Tumor Suppressor p53-Binding Protein 1
  • RNF168 protein, human
  • Ubiquitin-Protein Ligases
  • MRE11 Homologue Protein