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Nucleosome dynamics regulates DNA processing

Nature Structural & Molecular Biology volume 20, pages 836–842 (2013)Cite this article

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Abstract

The repair of DNA double-strand breaks (DSBs) is critical for the maintenance of genome integrity. The first step in DSB repair by homologous recombination is the processing of the ends by one of two resection pathways, executed by the Saccharomyces cerevisiae Exo1 and Sgs1–Dna2 machineries. Here we report in vitro and in vivo studies that characterize the impact of chromatin on each resection pathway. We find that efficient resection by the Sgs1–Dna2–dependent machinery requires a nucleosome-free gap adjacent to the DSB. Resection by Exo1 is blocked by nucleosomes, and processing activity can be partially restored by removal of the H2A–H2B dimers. Our study also supports a role for the dynamic incorporation of the H2A.Z histone variant in Exo1 processing, and it further suggests that the two resection pathways require distinct chromatin remodeling events to navigate chromatin structure.

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Figure 1: Increasing nucleosome density inhibits resection.
Figure 2: Mononucleosomes inhibit resection pathways differentially.
Figure 3: Increasing free DNA adjacent to a nucleosome enhances the helicase activity of Sgs1.
Figure 4: Nucleosomes act as a barrier to Exo1 DNA resection.
Figure 5: Loss of H2A–H2B dimers and incorporation of H2A.Z dimers promotes chromatin resection by Exo1.
Figure 6: Swr1 stimulates DSB repair through the Exo1 pathway.
Figure 7: Depletion of Swr1 inhibits Exo1 resection.

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Acknowledgements

This work was supported by grants from the US National Institutes of Health to N.L.A. (F32 GM096701), C.L.P. (RO1 GM54096) and P.S. (RO1 ES07061). We thank Y. Kwon (Yale University, New Haven, Connecticut, USA) for purified MRX, G. Ira (Baylor University, Houston, Texas, USA) for yeast strains, V. Borde (Institute Curie, Paris, France) for RPA antibody, C. Van (University of Massachusetts Medical School, Worcester, Massachusetts, USA) for help with the degron experiments and M. Liskay (Ohio State University, Columbus, Ohio, USA) for the Exo1 clone.

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Authors and Affiliations

  1. Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, Massachusetts, USA

    Nicholas L Adkins & Craig L Peterson

  2. Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, Connecticut, USA

    Hengyao Niu & Patrick Sung

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  1. Nicholas L Adkins
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N.L.A. performed all experiments, P.S. and H.N. provided purified resection enzymes, and all authors were involved in data analysis and manuscript preparation.

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Correspondence to Craig L Peterson.

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Adkins, N., Niu, H., Sung, P. et al. Nucleosome dynamics regulates DNA processing. Nat Struct Mol Biol 20, 836–842 (2013). https://doi.org/10.1038/nsmb.2585

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