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

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

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Department of Medical Biology, Amsterdam University Medical Centers (location AMC), Cancer Center Amsterdam, Meibergdreef 15, 1105 AZ Amsterdam, The Netherlands.
Department of Cell Biochemistry, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, 30-387 Krakow, Poland.
Erasmus MC, University Medical Center Rotterdam, Department of Molecular Genetics, Oncode Institute, Wytemaweg 80, 3015 CN, Rotterdam, The Netherlands.


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.

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