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Chromatin relaxation in response to DNA double-strand breaks is modulated by a novel ATM- and KAP-1 dependent pathway

Abstract

The cellular DNA-damage response is a signaling network that is vigorously activated by cytotoxic DNA lesions, such as double-strand breaks (DSBs)1. The DSB response is mobilized by the nuclear protein kinase ATM, which modulates this process by phosphorylating key players in these pathways2. A long-standing question in this field is whether DSB formation affects chromatin condensation. Here, we show that DSB formation is followed by ATM-dependent chromatin relaxation. ATM's effector in this pathway is the protein KRAB-associated protein (KAP-1, also known as TIF1β, KRIP-1 or TRIM28), previously known as a corepressor of gene transcription3,4. In response to DSB induction, KAP-1 is phosphorylated in an ATM-dependent manner on Ser 824. KAP-1 is phosphorylated exclusively at the damage sites, from which phosphorylated KAP-1 spreads rapidly throughout the chromatin. Ablation of the phosphorylation site of KAP-1 leads to loss of DSB-induced chromatin decondensation and renders the cells hypersensitive to DSB-inducing agents. Knocking down KAP-1, or mimicking a constitutive phosphorylation of this protein, leads to constitutive chromatin relaxation. These results suggest that chromatin relaxation is a fundamental pathway in the DNA-damage response and identify its primary mediators.

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Figure 1: KAP-1 is a novel ATM substrate in the DNA-damage response.
Figure 2: Functional significance of ATM-mediated phosphorylation of KAP-1 in the DNA-damage response.
Figure 3: KAP-1 is phosphorylated at the DSB sites by chromatin bound ATM and the phosphorylated protein rapidly spreads throughout the nucleus.
Figure 4: Chromatin relaxation in response to DSBs.

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Acknowledgements

We thank M. Oren for alerting us to the DSB response detected by the anti-pS407–Mdm2 antibody, R. Agami for the pRETRO-SUPER vector and one of the sequences used to knock down ATM, Y. Lerenthal for establishing the ATM knockdown HEK293, S. Biton for establishing the ATM knockdown LA-N-5 cells, T. Halazonetis for a gift of anti-53BP1 antibodies and L. Mittelman for expert assistance with confocal microscopy. This work was supported by research grants from the A-T Children's Project, The A-T Medical Research Foundation, The National Institutes of Health (NS31763), the A-T Medical Research Trust, The A-T Ease Foundation, The European Union, the Danish National Research Foundation, and the John and Birthe Meyer Foundation. This work was carried out in partial fulfillment of the requirements for the Ph.D. degree of D.B.

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Correspondence to Yael Ziv or Yosef Shiloh.

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Supplementary Figures S1, S2, S3, S4, S5, S6, S7 and S8 (PDF 3288 kb)

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Ziv, Y., Bielopolski, D., Galanty, Y. et al. Chromatin relaxation in response to DNA double-strand breaks is modulated by a novel ATM- and KAP-1 dependent pathway. Nat Cell Biol 8, 870–876 (2006). https://doi.org/10.1038/ncb1446

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