Structural basis for recognition of H3K56-acetylated histone H3–H4 by the chaperone Rtt106


Dynamic variations in the structure of chromatin influence virtually all DNA-related processes in eukaryotes and are controlled in part by post-translational modifications of histones1,2,3. One such modification, the acetylation of lysine 56 (H3K56ac) in the amino-terminal α-helix (αN) of histone H3, has been implicated in the regulation of nucleosome assembly during DNA replication and repair, and nucleosome disassembly during gene transcription4,5,6,7,8,9,10. In Saccharomyces cerevisiae, the histone chaperone Rtt106 contributes to the deposition of newly synthesized H3K56ac-carrying H3–H4 complex on replicating DNA5, but it is unclear how Rtt106 binds H3–H4 and specifically recognizes H3K56ac as there is no apparent acetylated lysine reader domain in Rtt106. Here, we show that two domains of Rtt106 are involved in a combinatorial recognition of H3–H4. An N-terminal domain homodimerizes and interacts with H3–H4 independently of acetylation while a double pleckstrin-homology (PH) domain binds the K56-containing region of H3. Affinity is markedly enhanced upon acetylation of K56, an effect that is probably due to increased conformational entropy of the αN helix of H3. Our data support a mode of interaction where the N-terminal homodimeric domain of Rtt106 intercalates between the two H3–H4 components of the (H3–H4)2 tetramer while two double PH domains in the Rtt106 dimer interact with each of the two H3K56ac sites in (H3–H4)2. We show that the Rtt106–(H3–H4)2 interaction is important for gene silencing and the DNA damage response.

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Figure 1: 3D structures of Rtt106 dimeric and double PH domains and their interaction with histones.
Figure 2: Identification of a K56ac-binding cleft in Rtt106 and model of Rtt106 in complex with K56-acetylated (H3–H4)2.
Figure 3: Effects of Rtt106PH mutations on H3K56ac interaction.
Figure 4: Effects of Rtt106 mutations on HMR silencing and genome stability.

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Biological Magnetic Resonance Data Bank

Protein Data Bank

Data deposits

The atomic coordinates and structure factors orNMR restraints of Rtt106DD, Rtt106PH, Rtt106PHL and Rtt106PH–acetyl-histamine have been deposited with the Protein Data Bank under accession codes 2LH0, 3FSS, 3TVV and 3TW1, respectively. The NMR chemical shifts of Rtt106 (residues 1–67) have been deposited in the Biological Magnetic Resonance Bank with the accession code 17832.


  1. 1

    Luger, K. Dynamic nucleosomes. Chromosome Res. 14, 5–16 (2006)

    CAS  Article  Google Scholar 

  2. 2

    Kouzarides, T. Chromatin modifications and their function. Cell 128, 693–705 (2007)

    CAS  Article  Google Scholar 

  3. 3

    Corpet, A. & Almouzni, G. Making copies of chromatin: the challenge of nucleosomal organization and epigenetic information. Trends Cell Biol. 19, 29–41 (2009)

    CAS  Article  Google Scholar 

  4. 4

    Chen, C. C. et al. Acetylated lysine 56 on histone H3 drives chromatin assembly after repair and signals for the completion of repair. Cell 134, 231–243 (2008)

    CAS  Article  Google Scholar 

  5. 5

    Li, Q. et al. Acetylation of histone H3 lysine 56 regulates replication-coupled nucleosome assembly. Cell 134, 244–255 (2008)

    CAS  Article  Google Scholar 

  6. 6

    Williams, S. K., Truong, D. & Tyler, J. K. Acetylation in the globular core of histone H3 on lysine-56 promotes chromatin disassembly during transcriptional activation. Proc. Natl Acad. Sci. USA 105, 9000–9005 (2008)

    ADS  CAS  Article  Google Scholar 

  7. 7

    Xie, W. et al. Histone H3 lysine 56 acetylation is linked to the core transcriptional network in human embryonic stem cells. Mol. Cell 33, 417–427 (2009)

    CAS  Article  Google Scholar 

  8. 8

    Das, C., Lucia, M. S., Hansen, K. C. & Tyler, J. K. CBP/p300-mediated acetylation of histone H3 on lysine 56. Nature 459, 113–117 (2009)

    ADS  CAS  Article  Google Scholar 

  9. 9

    Tjeertes, J. V., Miller, K. M. & Jackson, S. P. Screen for DNA-damage-responsive histone modifications identifies H3K9Ac and H3K56Ac in human cells. EMBO J. 28, 1878–1889 (2009)

    CAS  Article  Google Scholar 

  10. 10

    Yuan, J., Pu, M., Zhang, Z. & Lou, Z. Histone H3–K56 acetylation is important for genomic stability in mammals. Cell Cycle 8, 1747–1753 (2009)

    CAS  Article  Google Scholar 

  11. 11

    VanDemark, A. P. et al. The structure of the yFACT Pob3-M domain, its interaction with the DNA replication factor RPA, and a potential role in nucleosome deposition. Mol. Cell 22, 363–374 (2006)

    CAS  Article  Google Scholar 

  12. 12

    Huang, R. et al. Site-specific introduction of an acetyl-lysine mimic into peptides and proteins by cysteine alkylation. J. Am. Chem. Soc. 132, 9986–9987 (2010)

    CAS  Article  Google Scholar 

  13. 13

    Blomberg, N., Baraldi, E., Nilges, M. & Saraste, M. The PH superfold: a structural scaffold for multiple functions. Trends Biochem. Sci. 24, 441–445 (1999)

    CAS  Article  Google Scholar 

  14. 14

    Bowman, A., Ward, R., El-Mkami, H., Owen-Hughes, T. & Norman, D. G. Probing the (H3–H4)2 histone tetramer structure using pulsed EPR spectroscopy combined with site-directed spin labeling. Nucleic Acids Res. 38, 695–707 (2010)

    CAS  Article  Google Scholar 

  15. 15

    Liu, Y. et al. Structural analysis of Rtt106p reveals a DNA binding role required for heterochromatin silencing. J. Biol. Chem. 285, 4251–4262 (2010)

    CAS  Article  Google Scholar 

  16. 16

    White, C. L., Suto, R. K. & Luger, K. Structure of the yeast nucleosome core particle reveals fundamental changes in internucleosome interactions. EMBO J. 20, 5207–5218 (2001)

    CAS  Article  Google Scholar 

  17. 17

    Huang, S. et al. Rtt106p is a histone chaperone involved in heterochromatin-mediated silencing. Proc. Natl Acad. Sci. USA 102, 13410–13415 (2005)

    ADS  CAS  Article  Google Scholar 

  18. 18

    Huang, S., Zhou, H., Tarara, J. & Zhang, Z. A novel role for histone chaperones CAF-1 and Rtt106p in heterochromatin silencing. EMBO J. 26, 2274–2283 (2007)

    CAS  Article  Google Scholar 

  19. 19

    Dhalluin, C. et al. Structure and ligand of a histone acetyltransferase bromodomain. Nature 399, 491–496 (1999)

    ADS  CAS  Article  Google Scholar 

  20. 20

    Luger, K., Rechsteiner, T. J. & Richmond, T. J. Preparation of nucleosome core particle from recombinant histones. Methods Enzymol. 304, 3–19 (1999)

    CAS  Article  Google Scholar 

  21. 21

    Botuyan, M. V. et al. Structural basis of BACH1 phosphopeptide recognition by BRCA1 tandem BRCT domains. Structure 12, 1137–1146 (2004)

    CAS  Article  Google Scholar 

  22. 22

    Hendrickson, W. A., Horton, J. R. & LeMaster, D. M. Selenomethionyl proteins produced for analysis by multiwavelength anomalous diffraction (MAD): a vehicle for direct determination of three-dimensional structure. EMBO J. 9, 1665–1672 (1990)

    CAS  Article  Google Scholar 

  23. 23

    Minor, W., Cymborowski, M., Otwinowski, Z. & Chruszcz, M. HKL-3000: the integration of data reduction and structure solution–from diffraction images to an initial model in minutes. Acta Crystallogr. D 62, 859–866 (2006)

    Article  Google Scholar 

  24. 24

    Sheldrick, G. M. A short history of SHELX. Acta Crystallogr. A 64, 112–122 (2008)

    ADS  CAS  Article  Google Scholar 

  25. 25

    Terwilliger, T. C. SOLVE and RESOLVE: automated structure solution and density modification. Methods Enzymol. 374, 22–37 (2003)

    CAS  Article  Google Scholar 

  26. 26

    Perrakis, A., Morris, R. & Lamzin, V. S. Automated protein model building combined with iterative structure refinement. Nature Struct. Biol. 6, 458–463 (1999)

    CAS  Article  Google Scholar 

  27. 27

    Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. D 60, 2126–2132 (2004)

    Article  Google Scholar 

  28. 28

    Murshudov, G. N., Vagin, A. A. & Dodson, E. J. Refinement of macromolecular structures by the maximum-likelihood method. Acta Crystallogr. D 53, 240–255 (1997)

    CAS  Article  Google Scholar 

  29. 29

    Otwinowski, Z. & Minor, W. Processing of X-ray diffraction data collected in oscillation mode. Methods Enzymol. 276, 307–326 (1997)

    CAS  Article  Google Scholar 

  30. 30

    Storoni, L. C., McCoy, A. J. & Read, R. J. Likelihood-enhanced fast rotation functions. Acta Crystallogr. D 60, 432–438 (2004)

    Article  Google Scholar 

  31. 31

    Adams, P. D. et al. PHENIX: building new software for automated crystallographic structure determination. Acta Crystallogr. D 58, 1948–1954 (2002)

    Article  Google Scholar 

  32. 32

    Ferentz, A. E. & Wagner, G. NMR spectroscopy: a multifaceted approach to macromolecular structure. Q. Rev. Biophys. 33, 29–65 (2000)

    CAS  Article  Google Scholar 

  33. 33

    Delaglio, F. et al. NMRPipe: a multidimensional spectral processing system based on UNIX pipes. J. Biomol. NMR 6, 277–293 (1995)

    CAS  Article  Google Scholar 

  34. 34

    Brünger, A. T. et al. Crystallography & NMR system: A new software suite for macromolecular structure determination. Acta Crystallogr. D 54, 905–921 (1998)

    Article  Google Scholar 

  35. 35

    Koradi, R., Billeter, M. & Wüthrich, K. MOLMOL: a program for display and analysis of macromolecular structures. J. Mol. Graph. 14, 51–55 (1996)

    CAS  Article  Google Scholar 

  36. 36

    Puig, O. et al. The tandem affinity purification (TAP) method: a general procedure of protein complex purification. Methods 24, 218–229 (2001)

    CAS  Article  Google Scholar 

  37. 37

    Thomas, B. J. & Rothstein, R. Elevated recombination rates in transcriptionally active DNA. Cell 56, 619–630 (1989)

    CAS  Article  Google Scholar 

  38. 38

    Zhou, H., Madden, B. J., Muddiman, D. C. & Zhang, Z. Chromatin assembly factor 1 interacts with histone H3 methylated at lysine 79 in the processes of epigenetic silencing and DNA repair. Biochemistry 45, 2852–2861 (2006)

    CAS  Article  Google Scholar 

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We are grateful to N. Juranić, S. Macura and T. Burghardt for experimental advice, Y. Kim for assistance with X-ray data collection, Z. Zhang for suggestions on chemical synthesis, and K. Luger and A. Hieb for advice on the preparation of histones. We acknowledge the use of synchrotron beamlines 19BM and 19ID of the Structural Biology Center at Argonne National Laboratory’s APS, supported by the US Department of Energy, Basic Energy Sciences, Office of Science, under contract no. W-31-109-ENG-38. This work was funded in part by National Institutes of Health grants to Z.Z. and G.M.

Author information




Q.H. carried out the NMR spectroscopy experiments, prepared methylthiocarbonyl-aziridine and acetylated H3–H4, and performed the ITC measurements and analysis with assistance from G.M.; D.S. obtained crystals of the Rtt106 constructs, performed the X-ray diffraction measurements and solved the structures. Q.L. did the in vivo experiments. G.C. helped with the NMR experiments and ITC data analysis, A.F. and B.A.D. purified Rtt106 for initial crystal screening. J.R.T. worked on the refinement of crystal structures, M.V.B. contributed extensively to plasmid design, mutagenesis and protein preparations. G.M. and Z.Z. supervised the research. G.M. wrote the manuscript. All authors contributed in editing the manuscript.

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Correspondence to Zhiguo Zhang or Georges Mer.

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The authors declare no competing financial interests.

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This file contains a Supplementary Discussion, Supplementary Figures 1-14 with legends, Supplementary Tables 1-4 and Supplementary References. (PDF 14606 kb)

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Su, D., Hu, Q., Li, Q. et al. Structural basis for recognition of H3K56-acetylated histone H3–H4 by the chaperone Rtt106. Nature 483, 104–107 (2012).

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