• Open Access

Imaging of the DNA damage-induced dynamics of nuclear proteins via nonlinear photoperturbation

Authors

  • Martin Tomas,

    1. Department of Physics, Modern Optics and Quantum Electronics and Center for Applied Photonics, University of Konstanz, Konstanz, 78457, Germany
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  • Philipp Blumhardt,

    1. Department of Physics, Modern Optics and Quantum Electronics and Center for Applied Photonics, University of Konstanz, Konstanz, 78457, Germany
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  • Anja Deutzmann,

    1. Department of Biology, Bioimaging Center and Center for Applied Photonics, University of Konstanz, Konstanz, 78457, Germany
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  • Tobias Schwarz,

    1. Department of Biology, Bioimaging Center and Center for Applied Photonics, University of Konstanz, Konstanz, 78457, Germany
    2. Present Address: Light Microscopy Centre, ETH Zürich, Zürich, 8093, Switzerland
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  • Dimitri Kromm,

    1. Department of Physics, Modern Optics and Quantum Electronics and Center for Applied Photonics, University of Konstanz, Konstanz, 78457, Germany
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  • Alfred Leitenstorfer,

    1. Department of Physics, Modern Optics and Quantum Electronics and Center for Applied Photonics, University of Konstanz, Konstanz, 78457, Germany
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  • Elisa Ferrando-May

    Corresponding author
    1. Department of Biology, Bioimaging Center and Center for Applied Photonics, University of Konstanz, Konstanz, 78457, Germany
    • Phone: +49 7531 884054, Fax: +49 7531 884005
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Abstract

Understanding the cellular response to DNA strand breaks is crucial to decipher the mechanisms maintaining the integrity of our genome. We present a novel method to visualize how the mobility of nuclear proteins changes in response to localized DNA damage. DNA strand breaks are induced via nonlinear excitation with femtosecond laser pulses at λ = 1050 nm in a 3D-confined subnuclear volume. After a time delay of choice, protein mobility within this volume is analysed by two-photon photoactivation of PA-GFP fusion proteins at λ = 775 nm. By changing the position of the photoactivation spot with respect to the zone of lesion the influence of chromatin structure and of the distance from damage are investigated. As first applications we demonstrate a locally confined, time-dependent mobility increase of histone H1.2, and a progressive retardation of the DNA repair factor XRCC1 at damaged sites. This assay can be used to map the response of nuclear proteins to DNA damage in time and space. (© 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)

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