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ATRX ADD domain links an atypical histone methylation recognition mechanism to human mental-retardation syndrome

Nature Structural & Molecular Biology volume 18, pages 769–776 (2011)Cite this article

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Abstract

ATR-X (alpha-thalassemia/mental retardation, X-linked) syndrome is a human congenital disorder that causes severe intellectual disabilities. Mutations in the ATRX gene, which encodes an ATP-dependent chromatin-remodeler, are responsible for the syndrome. Approximately 50% of the missense mutations in affected persons are clustered in a cysteine-rich domain termed ADD (ATRX-DNMT3-DNMT3L, ADDATRX), whose function has remained elusive. Here we identify ADDATRX as a previously unknown histone H3–binding module, whose binding is promoted by lysine 9 trimethylation (H3K9me3) but inhibited by lysine 4 trimethylation (H3K4me3). The cocrystal structure of ADDATRX bound to H31–15K9me3 peptide reveals an atypical composite H3K9me3-binding pocket, which is distinct from the conventional trimethyllysine-binding aromatic cage. Notably, H3K9me3-pocket mutants and ATR-X syndrome mutants are defective in both H3K9me3 binding and localization at pericentromeric heterochromatin; thus, we have discovered a unique histone-recognition mechanism underlying the ATR-X etiology.

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Figure 1: Methylation at K4 and K9 inversely regulates interaction between ADDATRX and histone H3.
Figure 2: Atrx is recruited to PCH by Suv39h1- and Suv39h2-mediated H3K9 trimethylation.
Figure 3: Molecular basis for H3K9me3 recognition by ADDATRX.
Figure 4: Effect of H3 sequence context on ADDATRX binding.
Figure 5: Mutations in ADDATRX compromise histone binding and targeting of ATRX onto PCH.
Figure 6: Model of the mechanism underlying ATR-X syndrome that is caused by the mutations in ADDATRX.

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Acknowledgements

The authors are grateful to the staffs of beamlines X-29 at the Brookhaven National Laboratory, 24IDC at the Advanced Photon Source and 3W1A at the Beijing Synchrotron Radiation Facility (BSRF) for their assistance in the synchrotron data collection, with special thanks to Y. Dong of BSRF. We thank A. Vaquero (Bellvitge Institute for Biomedical Research) and S. Minucci (European Institute of Oncology) for providing the Suv39h1 and Suv39h2 double knockout cells and SUV39H1 expression vectors. Anti-MeCP2 antiserum was a generous gift from M. Greenberg (Harvard Medical School, HMS). We are grateful to A. Ruthenburg (Rockefeller University) for providing histone H3 peptides and to L. Liang (Memorial Sloan-Kettering Cancer Center), Y.Y. Chen (Institute of Biophysics, Chinese Academy of Sciences) and M. Geigges (Children's Hospital Boston) for experimental help. We express great appreciation to C. Richardson, M. Takahashi at HMS and individual members of the Shi, Li and Patel laboratories for helpful discussions. S.I. was a Jane Coffin Childs Memorial Fund Postdoctoral Fellowship Award recipient and is currently supported by the Japan Society for the Promotion of Science Postdoctoral Fellowship for Research Abroad. This work was supported by grants GM071004 and GM058012 from the US National Institutes of Health (Y.S.) and by the Starr Foundation, the Leukemia and Lymphoma Society, the Abby Rockefeller Mauze Trust and the Maloris Foundation (D.J. Patel); the Canadian Institutes of Health Research (D.J. Picketts); the Major State Basic Research Development Program in China (grant # 2011CB965300, H.L.) and by Tsinghua University 985 Phase II funds (H.L.).

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Authors and Affiliations

  1. Division of Newborn Medicine, Department of Medicine, Children's Hospital Boston, Harvard Medical School, Boston, Massachusetts, USA

    Shigeki Iwase, Ting Ren & Yang Shi

  2. Department of Medicine, Epigenetics Program, Children's Hospital Boston, Harvard Medical School, Boston, Massachusetts, USA

    Shigeki Iwase, Ting Ren & Yang Shi

  3. Center for Structural Biology, School of Life Sciences, Tsinghua University, Beijing, China

    Bin Xiang & Haitao Li

  4. School of Medicine, Tsinghua University, Beijing, China

    Bin Xiang & Haitao Li

  5. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts, USA

    Sharmistha Ghosh

  6. Laboratory of Chromatin Biology and Epigenetics, Rockefeller University, New York, New York, USA

    Peter W Lewis & C David Allis

  7. Department of Molecular Biology, Massachusetts General Hospital, Boston, Massachusetts, USA

    Jesse C Cochrane

  8. Regenerative Medicine Program, Ottawa Hospital Research Institute, Ottawa, Ontario, Canada

    David J Picketts

  9. Structural Biology Program, Memorial Sloan-Kettering Cancer Center, New York, New York, USA

    Dinshaw J Patel

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  1. Shigeki Iwase
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Contributions

S.I., H.L., D.J. Patel and Y.S. participated in the experimental design, contributed to the concept and wrote the paper. S.I. did pulldown assays, SPR assays and localization analysis. B.X. crystallized the ADDATRX–H3K9me3 complex and participated in calorimetric assays. H.L. carried out crystallographic and calorimetric studies. P.W.L., T.R. and J.C.C. carried out or helped with pulldown assays. S.G. helped with measuring SPR. C.D.A. supervised pulldown assays. D.J. Picketts provided important reagents and participated in manuscript preparation.

Corresponding authors

Correspondence to Dinshaw J Patel, Haitao Li or Yang Shi.

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Competing interests

Y.S. is a cofounder of Constellation Pharmaceuticals. D.J. Patel is on the Epigenetics Advisory Board of Epinova-Glaxo. The remaining authors declare no competing financial interests.

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Iwase, S., Xiang, B., Ghosh, S. et al. ATRX ADD domain links an atypical histone methylation recognition mechanism to human mental-retardation syndrome. Nat Struct Mol Biol 18, 769–776 (2011). https://doi.org/10.1038/nsmb.2062

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