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Positional stability of single double-strand breaks in mammalian cells

Nature Cell Biology volume 9, pages 675–682 (2007)Cite this article

Abstract

Formation of cancerous translocations requires the illegitimate joining of chromosomes containing double-strand breaks (DSBs). It is unknown how broken chromosome ends find their translocation partners within the cell nucleus. Here, we have visualized and quantitatively analysed the dynamics of single DSBs in living mammalian cells. We demonstrate that broken ends are positionally stable and unable to roam the cell nucleus. Immobilization of broken chromosome ends requires the DNA-end binding protein Ku80, but is independent of DNA repair factors, H2AX, the MRN complex and the cohesion complex. DSBs preferentially undergo translocations with neighbouring chromosomes and loss of local positional constraint correlates with elevated genomic instability. These results support a contact-first model in which chromosome translocations predominantly form among spatially proximal DSBs.

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Figure 1: An experimental system to visualize single broken DNA ends.
Figure 2: Analysis of the positional and local movement of broken DNA ends.
Figure 3: Separation of broken DNA ends in the absence of Ku80.
Figure 4: Spatial proximity of preferential translocation partners.

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Acknowledgements

We are grateful to: T. Karpova and M. Kruhlak for help with the microscopy; E. Martinez for providing reagents; S. Mabon for technical assistance; E. Brunet for the help with LMPCR; T. Voss, K. Meaburn and all members of the Misteli laboratory for discussions. Imaging was performed at the National Cancer Institute (NCI) Fluorescence Imaging Facility. E.S. was supported by a fellowship from the Human Frontiers Science Program (HFSP). J.D. is a fellow of the Roche Research Foundation. This research was supported in part by the Intramural Research Program of the National Institutes of Health (NIH), NCI, Center for Cancer Research and by the NIH grant GM 68956.

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Authors and Affiliations

  1. National Cancer Institute, NIH, Bethesda, 20892, MD, USA

    Evi Soutoglou & Tom Misteli

  2. The Scripps Research Institute, La Jolla, 92037, CA, USA

    Jonas F. Dorn & Gaudenz Danuser

  3. Genetics Branch, National Cancer Institute, NIH, Bethesda, 20892, MD, USA

    Kundan Sengupta & Thomas Ried

  4. Memorial Sloan Kettering Cancer Center, New York, 10021, NY, USA

    Maria Jasin

  5. Experimental Immunology Branch, National Cancer Institute, NIH, Bethesda, 20892, MD, USA

    Andre Nussenzweig

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  1. Evi Soutoglou
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Contributions

E.S. and T.M. designed the study, E.S., J.F.D. and K.S. performed the experiments, J.M., A.N. and T.R. provided reagents and advice, and E.S. and T.M. wrote the manuscript.

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Correspondence to Tom Misteli.

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The authors declare no competing financial interests.

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Soutoglou, E., Dorn, J., Sengupta, K. et al. Positional stability of single double-strand breaks in mammalian cells. Nat Cell Biol 9, 675–682 (2007). https://doi.org/10.1038/ncb1591

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