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Cooperative binding of two acetylation marks on a histone tail by a single bromodomain

Abstract

A key step in many chromatin-related processes is the recognition of histone post-translational modifications by effector modules such as bromodomains and chromo-like domains of the Royal family1,2. Whereas effector-mediated recognition of single post-translational modifications is well characterized3, how the cell achieves combinatorial readout of histones bearing multiple modifications is poorly understood. One mechanism involves multivalent binding by linked effector modules4. For example, the tandem bromodomains of human TATA-binding protein-associated factor-1 (TAF1) bind better to a diacetylated histone H4 tail than to monoacetylated tails, a cooperative effect attributed to each bromodomain engaging one acetyl-lysine mark5. Here we report a distinct mechanism of combinatorial readout for the mouse TAF1 homologue Brdt, a testis-specific member of the BET protein family6. Brdt associates with hyperacetylated histone H4 (ref. 7) and is implicated in the marked chromatin remodelling that follows histone hyperacetylation during spermiogenesis, the stage of spermatogenesis in which post-meiotic germ cells mature into fully differentiated sperm7,8,9,10. Notably, we find that a single bromodomain (BD1) of Brdt is responsible for selectively recognizing histone H4 tails bearing two or more acetylation marks. The crystal structure of BD1 bound to a diacetylated H4 tail shows how two acetyl-lysine residues cooperate to interact with one binding pocket. Structure-based mutagenesis that reduces the selectivity of BD1 towards diacetylated tails destabilizes the association of Brdt with acetylated chromatin in vivo. Structural analysis suggests that other chromatin-associated proteins may be capable of a similar mode of ligand recognition, including yeast Bdf1, human TAF1 and human CBP/p300 (also known as CREBBP and EP300, respectively). Our findings describe a new mechanism for the combinatorial readout of histone modifications in which a single effector module engages two marks on a histone tail as a composite binding epitope.

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Figure 1: Brdt recognizes a diacetylated H4 peptide through BD1.
Figure 2: Structure of the Brdt-BD1–H4K5acK8ac complex.
Figure 3: Stability of Brdt on acetylated chromatin mirrors BD1-mediated H4K5acK8ac recognition.

Accession codes

Primary accessions

Protein Data Bank

Data deposits

Atomic coordinates and structure factors have been deposited with the Protein Data Bank under accession codes 2WP2 (BD1) and 2WP1 (BD2).

References

  1. 1

    Berger, S. L. The complex language of chromatin regulation during transcription. Nature 447, 407–412 (2007)

    ADS  CAS  Article  Google Scholar 

  2. 2

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

    CAS  Article  Google Scholar 

  3. 3

    Taverna, S. D., Li, H., Ruthenburg, A. J., Allis, C. D. & Patel, D. J. How chromatin-binding modules interpret histone modifications: lessons from professional pocket pickers. Nature Struct. Mol. Biol. 14, 1025–1040 (2007)

    CAS  Article  Google Scholar 

  4. 4

    Ruthenburg, A. J., Li, H., Patel, D. J. & Allis, C. D. Multivalent engagement of chromatin modifications by linked binding modules. Nature Rev. Mol. Cell Biol. 8, 983–994 (2007)

    CAS  Article  Google Scholar 

  5. 5

    Jacobson, R. H., Ladurner, A. G., King, D. S. & Tjian, R. Structure and function of a human TAFII250 double bromodomain module. Science 288, 1422–1425 (2000)

    ADS  CAS  Article  Google Scholar 

  6. 6

    Florence, B. & Faller, D. V. You BET-cha: a novel family of transcriptional regulators. Front. Biosci. 6, d1008–d1018 (2001)

    CAS  PubMed  Google Scholar 

  7. 7

    Pivot-Pajot, C. et al. Acetylation-dependent chromatin reorganization by BRDT, a testis-specific bromodomain-containing protein. Mol. Cell. Biol. 23, 5354–5365 (2003)

    CAS  Article  Google Scholar 

  8. 8

    Shang, E., Nickerson, H. D., Wen, D., Wang, X. & Wolgemuth, D. J. The first bromodomain of Brdt, a testis-specific member of the BET sub-family of double-bromodomain-containing proteins, is essential for male germ cell differentiation. Development 134, 3507–3515 (2007)

    CAS  Article  Google Scholar 

  9. 9

    Govin, J., Caron, C., Lestrat, C., Rousseaux, S. & Khochbin, S. The role of histones in chromatin remodelling during mammalian spermiogenesis. Eur. J. Biochem. 271, 3459–3469 (2004)

    CAS  Article  Google Scholar 

  10. 10

    Rousseaux, S. et al. Establishment of male-specific epigenetic information. Gene 345, 139–153 (2005)

    CAS  Article  Google Scholar 

  11. 11

    Crowley, T. E., Kaine, E. M., Yoshida, M., Nandi, A. & Wolgemuth, D. J. Reproductive cycle regulation of nuclear import, euchromatic localization, and association with components of Pol II mediator of a mammalian double-bromodomain protein. Mol. Endocrinol. 16, 1727–1737 (2002)

    CAS  Article  Google Scholar 

  12. 12

    Leroy, G., Rickards, B. & Flint, S. J. The double bromodomain proteins Brd2 and Brd3 couple histone acetylation to transcription. Mol. Cell 30, 51–60 (2008)

    CAS  Article  Google Scholar 

  13. 13

    Ladurner, A. G., Inouye, C., Jain, R. & Tjian, R. Bromodomains mediate an acetyl-histone encoded antisilencing function at heterochromatin boundaries. Mol. Cell 11, 365–376 (2003)

    CAS  Article  Google Scholar 

  14. 14

    Matangkasombut, O. & Buratowski, S. Different sensitivities of bromodomain factors 1 and 2 to histone H4 acetylation. Mol. Cell 11, 353–363 (2003)

    CAS  Article  Google Scholar 

  15. 15

    Toyama, R., Rebbert, M. L., Dey, A., Ozato, K. & Dawid, I. B. Brd4 associates with mitotic chromosomes throughout early zebrafish embryogenesis. Dev. Dyn. 237, 1636–1644 (2008)

    CAS  Article  Google Scholar 

  16. 16

    Dey, A., Chitsaz, F., Abbasi, A., Misteli, T. & Ozato, K. The double bromodomain protein Brd4 binds to acetylated chromatin during interphase and mitosis. Proc. Natl Acad. Sci. USA 100, 8758–8763 (2003)

    ADS  CAS  Article  Google Scholar 

  17. 17

    Dey, A. et al. A bromodomain protein, MCAP, associates with mitotic chromosomes and affects G2-to-M transition. Mol. Cell. Biol. 20, 6537–6549 (2000)

    CAS  Article  Google Scholar 

  18. 18

    Mujtaba, S., Zeng, L. & Zhou, M. M. Structure and acetyl-lysine recognition of the bromodomain. Oncogene 26, 5521–5527 (2007)

    CAS  Article  Google Scholar 

  19. 19

    Owen, D. J. et al. The structural basis for the recognition of acetylated histone H4 by the bromodomain of histone acetyltransferase Gcn5p. EMBO J. 19, 6141–6149 (2000)

    CAS  Article  Google Scholar 

  20. 20

    Kanno, T. et al. Selective recognition of acetylated histones by bromodomain proteins visualized in living cells. Mol. Cell 13, 33–43 (2004)

    CAS  Article  Google Scholar 

  21. 21

    Polesskaya, A. et al. Interaction between acetylated MyoD and the bromodomain of CBP and/or p300. Mol. Cell. Biol. 21, 5312–5320 (2001)

    CAS  Article  Google Scholar 

  22. 22

    Wei, L., Jamonnak, N., Choy, J., Wang, Z. & Zheng, W. Differential binding modes of the bromodomains of CREB-binding protein (CBP) and p300 with acetylated MyoD. Biochem. Biophys. Res. Commun. 368, 279–284 (2008)

    CAS  Article  Google Scholar 

  23. 23

    Govin, J. et al. Pericentric heterochromatin reprogramming by new histone variants during mouse spermiogenesis. J. Cell Biol. 176, 283–294 (2007)

    CAS  Article  Google Scholar 

  24. 24

    Kabsch, W. Automatic processing of rotation diffraction data from crystals of initially unknown symmetry and cell constants. J. Appl. Cryst. 26, 795–800 (1993)

    CAS  Article  Google Scholar 

  25. 25

    CCP4. The CCP4 suite: programs for protein crystallography. Acta Crystallogr D 50, 760–763 (1994)

  26. 26

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

    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

    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 

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Acknowledgements

We thank the ESRF and EMBL staff for beamline assistance, the Partnership for Structural Biology (PSB) for access to technical platforms, M. Jamin for MALLS experiments, G. Natrajan and V. Rybin for help with ITC, C. Soucier for help with FRAP, D. Panne and C. Clapier for comments on the manuscript, and S. Cusack for critical support. Work in the S.K. laboratory was supported by ANR blanche ‘EpiSperm’ and ‘Empreinte’, INCa and ‘ARECA’ (ARC) research programmes. J.Ga. was supported by a Ph.D. fellowship from the Rhône-Alpes region. J.M. was supported by an ‘E-STAR’ fellowship funded by EU FP6. C.P. acknowledges support from the ANRS/Fondation de France AIJC and CNRS ATIP programmes.

Author Contributions M.S.-L., D.J.H., S.K. and C.W.M. initiated the study. S.K., C.P. and C.W.M. coordinated the entire project and specific author contributions. J.M., S.R., U.S., M.S.-L., A.-L.V. and D.J.H. prepared constructs. J.M., U.S. and M.S.-L. expressed, purified and crystallized BD1 and BD2. J.M. and U.S. performed and analysed ITC and biophysical experiments. J.M., M.S.-L. and C.P. measured diffraction data and solved the crystal structures. C.P. and C.W.M. analysed structural and biochemical data. J.Go. performed immunoblot analysis and immunofluorescence microscopy. S.C. and S.R. performed FRAP and chromatin compaction experiments. J.Ga. prepared histones for MS/MS analysis. K.S. and J.K. performed the MS/MS analysis. C.P. wrote the manuscript. All authors discussed the results and commented on the manuscript.

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Correspondence to Christoph W. Müller.

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Morinière, J., Rousseaux, S., Steuerwald, U. et al. Cooperative binding of two acetylation marks on a histone tail by a single bromodomain. Nature 461, 664–668 (2009). https://doi.org/10.1038/nature08397

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