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Acetylation limits 53BP1 association with damaged chromatin to promote homologous recombination

Nature Structural & Molecular Biology volume 20, pages 317–325 (2013)Cite this article

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Abstract

The pathogenic sequelae of BRCA1 mutation in human and mouse cells are mitigated by concomitant deletion of 53BP1, which binds histone H4 dimethylated at Lys20 (H4K20me2) to promote nonhomologous end joining, suggesting that a balance between BRCA1 and 53BP1 regulates DNA double strand–break (DSB) repair mechanism choice. Here we document that acetylation is a key determinant of this balance. TIP60 acetyltransferase deficiency reduced BRCA1 at DSB chromatin with commensurate increases in 53BP1, whereas HDAC inhibition yielded the opposite effect. TIP60-dependent H4 acetylation diminished 53BP1 binding to H4K20me2 in part through disruption of a salt bridge between H4K16 and Glu1551 in the 53BP1 Tudor domain. Moreover, TIP60 deficiency impaired homologous recombination and conferred sensitivity to PARP inhibition in a 53BP1-dependent manner. These findings demonstrate that acetylation in cis to H4K20me2 regulates relative BRCA1 and 53BP1 DSB chromatin occupancy to direct DNA repair mechanism.

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Figure 1: TIP60 and HDAC inhibition differentially affect competition between BRCA1 and 53BP1 for localization to DSBs.
Figure 2: DNA damage– and transcription–dependent acetylation regulates BRCA1 and 53BP1 DSB occupancy.
Figure 3: Histone H4K16 acetylation impairs the interaction of the 53BP1 Tudor domain with H4K20me2.
Figure 4: Solution structure of 53BP1-Tudor in association with a methylated H4 peptide.
Figure 5: 53BP1 knockdown restores BRCA1 localization to DSBs.
Figure 6: HDACi or 53BP1 deficiency restores end resection and homology-directed DSB repair in TIP60-deficient cells.
Figure 7: 53BP1 promotes genomic instability and sensitivity to PARPi in the absence of TIP60.

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Acknowledgements

We thank J. Wu for technical support, J. Chen (MD Anderson Cancer Center) for 53BP1−/− mouse embryonic fibroblasts, M. Jasin (Memorial Sloan-Kettering Cancer Center) for U2OS DR-GFP reporter cells, G. Stewart (University of Birmingham) for RIDDLE cells and S. Janicki (Wistar Institute) and D. Spector (Cold Spring Harbor Laboratories) for the U2OS 2-6-3 reporter cell line. We thank K.M. Miller (University of Texas) and A. Sfeir (New York University) for their critical reading of the manuscript and helpful comments. R.A.G. is supported by grant 1R01CA138835-01 from the National Cancer Institute (NCI), a Research Scholar Grant from the American Cancer Society, a Department of Defense Breast Cancer Idea Award, a UPENN–Fox Chase Cancer Center (FCCC) Specialized Program of Research Excellence (SPORE) Pilot Grant and funds from the Abramson Family Cancer Research Institute and Basser Research Center for BRCA. G.M. acknowledges support from NCI grant 1R01CA132878 and funds from the Mayo Clinic Breast Cancer SPORE NCI grant P50CA116201.

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Author notes

  1. Nam Woo Cho and Gaofeng Cui: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Cancer Biology, Abramson Family Cancer Research Institute, Basser Research Center for BRCA, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA

    Jiangbo Tang, Nam Woo Cho, Erica M Manion, Niraj M Shanbhag & Roger A Greenberg

  2. Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, Minnesota, USA

    Gaofeng Cui, Maria Victoria Botuyan & Georges Mer

  3. Department of Pathology, Abramson Family Cancer Research Institute, Basser Research Center for BRCA, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA

    Roger A Greenberg

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Contributions

J.T. initiated the study and performed the majority of experiments with guidance from R.A.G. N.W.C. designed and validated the TRF1-FokI DSB reporter system in conjunction with N.M.S. and R.A.G. E.M.M. provided technical assistance to J.T. G.C. prepared the isotope-enriched proteins and peptides, and performed the NMR spectroscopy experiments and structure calculations. M.V.B. made the protein expression constructs and helped in sample preparations for NMR studies. G.M. supervised the structural studies. The study was conceived by J.T. and R.A.G. Writing was performed by J.T. and R.A.G. with contributions from G.M., N.M.S. and N.W.C.

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Correspondence to Roger A Greenberg.

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Tang, J., Cho, N., Cui, G. et al. Acetylation limits 53BP1 association with damaged chromatin to promote homologous recombination. Nat Struct Mol Biol 20, 317–325 (2013). https://doi.org/10.1038/nsmb.2499

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