Article | Published:

Dynamics of yeast histone H2A and H2B phosphorylation in response to a double-strand break

Nature Structural & Molecular Biology volume 21, pages 103109 (2014) | Download Citation

Abstract

In budding yeast, a single double-strand break (DSB) triggers extensive Tel1 (ATM)- and Mec1 (ATR)-dependent phosphorylation of histone H2A around the DSB, to form γ-H2AX. We describe Mec1- and Tel1-dependent phosphorylation of histone H2B at T129. γ-H2B formation is impaired by γ-H2AX and its binding partner Rad9. High-density microarray analyses show similar γ-H2AX and γ-H2B distributions, but γ-H2B is absent near telomeres. Both γ-H2AX and γ-H2B are strongly diminished over highly transcribed regions. When transcription of GAL7, GAL10 and GAL1 genes is turned off, γ-H2AX is restored within 5 min, in a Mec1-dependent manner; after reinduction of these genes, γ-H2AX is rapidly lost. Moreover, when a DSB is induced near CEN2, γ-H2AX spreads to all other pericentromeric regions, again depending on Mec1. Our data provide new insights in the function and establishment of phosphorylation events occurring on chromatin after DSB induction.

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Acknowledgements

We thanks V. Benes and T. Ivacevic from the European Molecular Biology Laboratory Genomic Core facility for hybridization with Affymetrix arrays. Funding in the Legube laboratory was provided by grants from the Association Contre le Cancer (ARC), Agence Nationale pour la Recherche (ANR-09-JCJC-0138), Canceropole Grand Sud-Ouest and Research Innovation Therapeutic Cancerologie (RITC). Research in the Haber lab was supported by US National Institutes of Health grants GM61766, GM20056 and GM76020.

Author information

Author notes

    • Cheng-Sheng Lee
    •  & Kihoon Lee

    These authors contributed equally to this work.

Affiliations

  1. Department of Biology, Brandeis University, Waltham, Massachusetts, USA.

    • Cheng-Sheng Lee
    • , Kihoon Lee
    •  & James E Haber
  2. Rosenstiel Basic Medical Sciences Research Center, Brandeis University, Waltham, Massachusetts, USA.

    • Cheng-Sheng Lee
    • , Kihoon Lee
    •  & James E Haber
  3. Université de Toulouse, Université Paul Sabatier, Laboratoire de Biologie Cellulaire et Moléculaire du Contrôle de la Proliferation (LBCMCP), Toulouse, France.

    • Gaëlle Legube
  4. Centre National de la Recherche Scientifique (CNRS), LBCMCP, Toulouse, France.

    • Gaëlle Legube

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Contributions

C.-S.L. designed, executed and analyzed experiments shown in Figures 5,6,7 and related supplementary information and prepared ChIP samples for experiments shown in Figures 2,3,4,5,6. K.L. designed, executed and analyzed experiments shown in Figure 1 and related Supplementary Information and prepared ChIP samples for experiments shown in Figures 2,3,4,5,6. G.L. performed ChIP-chip analysis and analyzed the data shown in Figures 2,3,4,5,6 and related supplementary information. J.E.H. designed experiments and analyzed data and was the principal author of the manuscript, with contributions from all other coauthors.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Gaëlle Legube or James E Haber.

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DOI

https://doi.org/10.1038/nsmb.2737

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