Poly-ADP-ribosylation is a post-translational modification catalyzed by PARP enzymes with roles in transcription and chromatin biology. Here we show that distinct macrodomains, including those of histone macroH2A1.1, are recruited to sites of PARP1 activation induced by laser-generated DNA damage. Chemical PARP1 inhibitors, PARP1 knockdown and mutation of ADP-ribose–binding residues in macroH2A1.1 abrogate macrodomain recruitment. Notably, histone macroH2A1.1 senses PARP1 activation, transiently compacts chromatin, reduces the recruitment of DNA damage factor Ku70–Ku80 and alters γ-H2AX patterns, whereas the splice variant macroH2A1.2, which is deficient in poly-ADP-ribose binding, does not mediate chromatin rearrangements upon PARP1 activation. The structure of the macroH2A1.1 macrodomain in complex with ADP-ribose establishes a poly-ADP-ribose cap-binding function and reveals conformational changes in the macrodomain upon ligand binding. We thus identify macrodomains as modules that directly sense PARP activation in vivo and establish macroH2A histones as dynamic regulators of chromatin plasticity.
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We thank M. Gossen (Max-Delbrück-Centrum for Molecular Medicine) for the HeLa TetOn system, S. Jackson (The Gurdon Institute) and KuDos Pharmaceuticals for purified DNA-PK holo-complex, A. Cohen and H. Stunnenberg for MS, V. Rybin in the European Molecular Biology Laboratory (EMBL) Protein Expression and Purification Facility, V. Benes and the EMBL GeneCore Facility, Olympus Europe for supporting EMBL's Advanced Light Microscopy Facility, J. Ellenberg for discussion of chromatin compaction and A. Akhtar, C. Häring, J. Conaway, I. Mattaj, V. Sartorelli and C. Wu for advice. This work was supported by funding from EMBL, EU Network of Excellence “The Epigenome”, the Peter and Traudl Engelhorn Stiftung (M.H.), EU Marie Curie Research Training Network “Chromatin Plasticity” and the Human Frontiers Science Program.
Supplementary Figures 1–10 and Supplementary Tables 1 and 2 (PDF 3841 kb)
Recruitment kinetics of the archaebacterial Af1521 macrodomain to microirradiated DNA sites in vivo. (MOV 3396 kb)
Recruitment kinetics of the histone macroH2A1.1 macrodomain (left) and PARP1 (middle) to micro-irradiated DNA sites in vivo. The right panel shows the merged channels. (MOV 5541 kb)
Recruitment kinetics of the human macroD2 macrodomain to micro-irradiated DNA sites in vivo. (MOV 9891 kb)
Comparison of the recruitment levels for fluorescently-tagged PARP1 (left) with fluorescently-tagged PARP2 (right) at a laser micro-irradiation site in HeLa cells. (MOV 8784 kb)
Confocal stacks showing the enrichment of Hoechst staining at laser micro-irradiation site in HeLa cells expressing the tagged macroH2A1.1 histone variant. The movie shows two nuclei in each panel. Hoechst staining (left), macroH2A1.1 antibody staining (middle) and phosphorylated H2AX histone staining (right). (MOV 1041 kb)
Confocal stacks showing the enrichment of Hoechst staining at a laser micro-irradiation site in HeLa cells that do not express the macroH2A1.1 histone transgene. Hoechst staining (left) and phosphorylated H2AX histone staining (right). (MOV 984 kb)
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Timinszky, G., Till, S., Hassa, P. et al. A macrodomain-containing histone rearranges chromatin upon sensing PARP1 activation. Nat Struct Mol Biol 16, 923–929 (2009). https://doi.org/10.1038/nsmb.1664
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