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Dissection of DNA damage and repair pathways in live cells by femtosecond laser microirradiation and free-electron modeling.
Schmalz, Michael; Liang, Xiao-Xuan; Wieser, Ines; Gruschel, Caroline; Muskalla, Lukas; Stöckl, Martin Thomas; Nitschke, Roland; Linz, Norbert; Leitenstorfer, Alfred; Vogel, Alfred; Ferrando-May, Elisa.
Afiliação
  • Schmalz M; Department of Physics, University of Konstanz, 78457 Konstanz, Germany.
  • Liang XX; Department of Biology, University of Konstanz, 78457 Konstanz, Germany.
  • Wieser I; Center for Applied Photonics, University of Konstanz, 78457 Konstanz, Germany.
  • Gruschel C; Institute of Biomedical Optics, University of Lübeck, 23562 Lübeck, Germany.
  • Muskalla L; Department of Biology, University of Konstanz, 78457 Konstanz, Germany.
  • Stöckl MT; Department of Biology, University of Konstanz, 78457 Konstanz, Germany.
  • Nitschke R; Department of Biology, University of Konstanz, 78457 Konstanz, Germany.
  • Linz N; Department of Biology, University of Konstanz, 78457 Konstanz, Germany.
  • Leitenstorfer A; Life Imaging Center and Centre for Biological Signalling Studies, University of Freiburg, 79104 Freiburg, Germany.
  • Vogel A; Institute of Biomedical Optics, University of Lübeck, 23562 Lübeck, Germany.
  • Ferrando-May E; Department of Physics, University of Konstanz, 78457 Konstanz, Germany.
Proc Natl Acad Sci U S A ; 120(25): e2220132120, 2023 06 20.
Article em En | MEDLINE | ID: mdl-37307476
Understanding and predicting the outcome of the interaction of light with DNA has a significant impact on the study of DNA repair and radiotherapy. We report on a combination of femtosecond pulsed laser microirradiation at different wavelengths, quantitative imaging, and numerical modeling that yields a comprehensive picture of photon-mediated and free-electron-mediated DNA damage pathways in live cells. Laser irradiation was performed under highly standardized conditions at four wavelengths between 515 nm and 1,030 nm, enabling to study two-photon photochemical and free-electron-mediated DNA damage in situ. We quantitatively assessed cyclobutane pyrimidine dimer (CPD) and γH2AX-specific immunofluorescence signals to calibrate the damage threshold dose at these wavelengths and performed a comparative analysis of the recruitment of DNA repair factors xeroderma pigmentosum complementation group C (XPC) and Nijmegen breakage syndrome 1 (Nbs1). Our results show that two-photon-induced photochemical CPD generation dominates at 515 nm, while electron-mediated damage dominates at wavelengths ≥620 nm. The recruitment analysis revealed a cross talk between nucleotide excision and homologous recombination DNA repair pathways at 515 nm. Numerical simulations predicted electron densities and electron energy spectra, which govern the yield functions of a variety of direct electron-mediated DNA damage pathways and of indirect damage by •OH radicals resulting from laser and electron interactions with water. Combining these data with information on free electron-DNA interactions gained in artificial systems, we provide a conceptual framework for the interpretation of the wavelength dependence of laser-induced DNA damage that may guide the selection of irradiation parameters in studies and applications that require the selective induction of DNA lesions.
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Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Dano ao DNA / Elétrons Tipo de estudo: Prognostic_studies Idioma: En Revista: Proc Natl Acad Sci U S A Ano de publicação: 2023 Tipo de documento: Article País de afiliação: Alemanha

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Dano ao DNA / Elétrons Tipo de estudo: Prognostic_studies Idioma: En Revista: Proc Natl Acad Sci U S A Ano de publicação: 2023 Tipo de documento: Article País de afiliação: Alemanha