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Molecular basis for site-specific read-out of histone H3K4me3 by the BPTF PHD finger of NURF

Nature volume 442, pages 91–95 (2006)Cite this article

Abstract

Mono-, di- and trimethylated states of particular histone lysine residues are selectively found in different regions of chromatin, thereby implying specialized biological functions for these marks ranging from heterochromatin formation to X-chromosome inactivation and transcriptional regulation1,2,3. A major challenge in chromatin biology has centred on efforts to define the connection between specific methylation states and distinct biological read-outs impacting on function4. For example, histone H3 trimethylated at lysine 4 (H3K4me3) is associated with transcription start sites of active genes5,6,7, but the molecular ‘effectors’ involved in specific recognition of H3K4me3 tails remain poorly understood. Here we demonstrate the molecular basis for specific recognition of H3(1–15)K4me3 (residues 1–15 of histone H3 trimethylated at K4) by a plant homeodomain (PHD) finger of human BPTF (bromodomain and PHD domain transcription factor), the largest subunit of the ATP-dependent chromatin-remodelling complex, NURF (nucleosome remodelling factor). We report on crystallographic and NMR structures of the bromodomain-proximal PHD finger of BPTF in free and H3(1–15)K4me3-bound states. H3(1–15)K4me3 interacts through anti-parallel β-sheet formation on the surface of the PHD finger, with the long side chains of arginine 2 (R2) and K4me3 fitting snugly in adjacent pre-formed surface pockets, and bracketing an invariant tryptophan. The observed stapling role by non-adjacent R2 and K4me3 provides a molecular explanation for H3K4me3 site specificity. Binding studies establish that the BPTF PHD finger exhibits a modest preference for K4me3- over K4me2-containing H3 peptides, and discriminates against monomethylated and unmodified counterparts. Furthermore, we identified key specificity-determining residues from binding studies of H3(1–15)K4me3 with PHD finger point mutants. Our findings call attention to the PHD finger as a previously uncharacterized chromatin-binding module found in a large number of chromatin-associated proteins.

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Figure 1: NMR, calorimetry and surface plasmon resonance-based studies of the binding of the BPTF PHD finger by H3(1–15)K4 peptides as a function of methylation state.
Figure 2: Crystal structures of human BPTF PHD finger-linker-bromodomain in the free state and bound to H3(1–15)K4me3.
Figure 3: Details of the intermolecular contacts in the H3(1–15)K4me3 peptide–BPTF PHD finger complex and comparison with its H3(1–15)K4me2-bound counterpart.
Figure 4: NMR-based structures of free and H3(1–15)K4me3-bound BPTF PHD finger, together with binding studies of PHD finger mutants with H3(1–15)K4me3 peptide.

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Acknowledgements

D.J.P. is supported by funds from the Abby Rockefeller Mauze Trust, and the Dewitt Wallace and Maloris Foundations. C.D.A. is supported by an NIH MERIT award and funds from Rockefeller University. S.I. holds a Ruth Kirschstein NIH postdoctoral fellowship and J.W. holds a Damon-Runyon CRF Fellowship. We thank the Peptide Core Facilities at Sloan-Kettering (S. S. Yi at Microchemistry and Proteomics) and Rockefeller University for the synthesis and purification of K4-methylated H3 peptides. We would like to thank the staff at beam line X25 at the Brookhaven National Laboratory and beam lines 23ID-D and 24ID-C of the Advanced Photon Source at the Argonne National Laboratory, supported by the US Department of Energy, for assistance with data collection. D.J.P. is a member of the New York Structural Biology Center, supported in part by funds from the NIH. Author Contributions H.L. is responsible for the X-ray and surface plasmon resonance studies, S.I. for the NMR studies, W.W. for the calorimetric studies, B.D. for fluorescence polarization studies and J.W. for identification and functional characterization of the BPTF PHD finger as a H3K4me3 reader. D.J.P. and C.D.A. supervised the structural and functional (see companion paper) aspects of the project, respectively, and take overall responsibility for their joint research. All authors discussed the results and commented on the manuscript.

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Authors and Affiliations

  1. Structural Biology Program, Memorial Sloan-Kettering Cancer Center, New York, New York, 10021, USA

    Haitao Li, Serge Ilin, Wooikoon Wang & Dinshaw J. Patel

  2. Laboratory of Chromatin Biology, Rockefeller University, New York, New York, 10021, USA

    Elizabeth M. Duncan, Joanna Wysocka & C. David Allis

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  1. Haitao Li
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Correspondence to C. David Allis or Dinshaw J. Patel.

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Coordinates of the X-ray structures of the BPTF PHD finger-linker-bromodomain in the free state and when bound to H3(1–15)K4me3 and H3(1–15)K4me2 peptides have been deposited in the RCSB Protein Data Bank under accession codes 2F6N, 2F6J and 2FSA, respectively. Coordinates of the NMR structures of the BPTF PHD finger in the free state and H3(1–15)K4me3 peptide-bound state have been deposited in the RCSB Protein Data Bank under accession codes 2FUI and 2FUU, respectively. Reprints and permissions information is available at npg.nature.com/reprintsandpermissions. The authors declare no competing financial interests.

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This file contains Supplementary Methods, Supplementary Notes, Supplementary Figures 1–13 and Supplementary Tables 1–3. (PDF 2841 kb)

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Li, H., Ilin, S., Wang, W. et al. Molecular basis for site-specific read-out of histone H3K4me3 by the BPTF PHD finger of NURF. Nature 442, 91–95 (2006). https://doi.org/10.1038/nature04802

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