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DNA binding by PHF1 prolongs PRC2 residence time on chromatin and thereby promotes H3K27 methylation

Nature Structural & Molecular Biology volume 24, pages 10391047 (2017) | Download Citation

Abstract

Polycomb repressive complex 2 (PRC2) trimethylates histone H3 at lysine 27 to mark genes for repression. We measured the dynamics of PRC2 binding on recombinant chromatin and free DNA at the single-molecule level using total internal reflection fluorescence (TIRF) microscopy. PRC2 preferentially binds free DNA with multisecond residence time and midnanomolar affinity. PHF1, a PRC2 accessory protein of the Polycomblike family, extends PRC2 residence time on DNA and chromatin. Crystallographic and functional studies reveal that Polycomblike proteins contain a winged-helix domain that binds DNA in a sequence-nonspecific fashion. DNA binding by this winged-helix domain accounts for the prolonged residence time of PHF1–PRC2 on chromatin and makes it a more efficient H3K27 methyltranferase than PRC2 alone. Together, these studies establish that interactions with DNA provide the predominant binding affinity of PRC2 for chromatin. Moreover, they reveal the molecular basis for how Polycomblike proteins stabilize PRC2 on chromatin and stimulate its activity.

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Acknowledgements

We thank the MPI Biochemistry Facility and the Crystallization Facility for support with the biophysical and structural experiments and the beamline scientists at the Swiss Light Source for excellent assistance with data collection. We thank T. Cech and his lab members X. Wang and R.D. Paucek (University of Colorado Boulder) for sharing unpublished data and discussions. We thank S. Kilic and M. Tobler for reagents. This work was supported by the European Commission Seventh Framework Program 4DCellFate (grant number 277899), the Max Planck Society (J.M.), the Swiss National Science Foundation (grant number 31003A_173169) and EPFL (B.F.).

Author information

Author notes

    • Jeongyoon Choi
    •  & Andreas Linus Bachmann

    These authors contributed equally to this work.

Affiliations

  1. Max Planck Institute of Biochemistry, Laboratory of Chromatin Biology, Martinsried, Germany.

    • Jeongyoon Choi
    • , Katharina Tauscher
    •  & Jürg Müller
  2. Ecole Polytechnique Fédérale de Lausanne (EPFL), Institute of Chemical Sciences and Engineering (ISIC), Laboratory of Biophysical Chemistry of Macromolecules, Lausanne, Switzerland.

    • Andreas Linus Bachmann
    •  & Beat Fierz
  3. Max Planck Institute of Biochemistry, Department of Structural Cell Biology, Martinsried, Germany.

    • Christian Benda

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Contributions

J.C. and J.M. conceived the project. J.C., C.B., B.F. and J.M. designed the experiments. J.C. performed protein purification, biophysical experiments, crystallization and HMTase assays. J.C. and A.L.B. performed smTIRFM. J.C., A.L.B., C.B., B.F. and J.M. discussed and interpreted the data. J.C., B.F. and J.M. wrote the manuscript. K.T. provided technical support.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Beat Fierz or Jürg Müller.

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https://doi.org/10.1038/nsmb.3488

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