Novel method for site-specific induction of oxidative DNA damage reveals differences in recruitment of repair proteins to heterochromatin and euchromatin

Li Lan, Satoshi Nakajima, Leizhen Wei, Luxi Sun, Ching Lung Hsieh, Robert W. Sobol, Marcel Bruchez, Bennett Van Houten, Akira Yasui, Arthur S. Levine

    Research output: Contribution to journalArticle

    50 Citations (Scopus)

    Abstract

    Reactive oxygen species (ROS)-induced DNA damage is repaired by the base excision repair pathway. However, the effect of chromatin structure on BER protein recruitment to DNA damage sites in living cells is poorly understood. To address this problem, we developed a method to specifically produce ROS-induced DNA damage by fusing KillerRed (KR), a light-stimulated ROS-inducer, to a tet-repressor (tetR-KR) or a transcription activator (TA-KR). TetR-KR or TA-KR, bound to a TRE cassette (∼90 kb) integrated at a defined genomic locus in U2OS cells, was used to induce ROS damage in hetero-or euchromatin, respectively. We found that DNA glycosylases were efficiently recruited to DNA damage in heterochromatin, as well as in euchromatin. PARP1 was recruited to DNA damage within condensed chromatin more efficiently than in active chromatin. In contrast, recruitment of FEN1 was highly enriched at sites of DNA damage within active chromatin in a PCNA-and transcription activation-dependent manner. These results indicate that oxidative DNA damage is differentially processed within hetero or euchromatin.

    Original languageEnglish
    Pages (from-to)2330-2345
    Number of pages16
    JournalNucleic acids research
    Volume42
    Issue number4
    DOIs
    Publication statusPublished - 2014 Feb

    ASJC Scopus subject areas

    • Genetics

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  • Cite this

    Lan, L., Nakajima, S., Wei, L., Sun, L., Hsieh, C. L., Sobol, R. W., Bruchez, M., Van Houten, B., Yasui, A., & Levine, A. S. (2014). Novel method for site-specific induction of oxidative DNA damage reveals differences in recruitment of repair proteins to heterochromatin and euchromatin. Nucleic acids research, 42(4), 2330-2345. https://doi.org/10.1093/nar/gkt1233