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RAG2 PHD finger couples histone H3 lysine 4 trimethylation with V(D)J recombination

Nature volume 450, pages 1106–1110 (2007)Cite this article

Abstract

Nuclear processes such as transcription, DNA replication and recombination are dynamically regulated by chromatin structure. Eukaryotic transcription is known to be regulated by chromatin-associated proteins containing conserved protein domains that specifically recognize distinct covalent post-translational modifications on histones. However, it has been unclear whether similar mechanisms are involved in mammalian DNA recombination. Here we show that RAG2—an essential component of the RAG1/2 V(D)J recombinase, which mediates antigen-receptor gene assembly1—contains a plant homeodomain (PHD) finger that specifically recognizes histone H3 trimethylated at lysine 4 (H3K4me3). The high-resolution crystal structure of the mouse RAG2 PHD finger bound to H3K4me3 reveals the molecular basis of H3K4me3-recognition by RAG2. Mutations that abrogate RAG2’s recognition of H3K4me3 severely impair V(D)J recombination in vivo. Reducing the level of H3K4me3 similarly leads to a decrease in V(D)J recombination in vivo. Notably, a conserved tryptophan residue (W453) that constitutes a key structural component of the K4me3-binding surface and is essential for RAG2’s recognition of H3K4me3 is mutated in patients with immunodeficiency syndromes. Together, our results identify a new function for histone methylation in mammalian DNA recombination. Furthermore, our results provide the first evidence indicating that disrupting the read-out of histone modifications can cause an inherited human disease.

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Figure 1: The RAG2 PHD finger is a novel H3K4me3-binding module.
Figure 2: The molecular basis of H3K4me3-recognition by RAG2PHD.
Figure 3: Recognition of H3K4me3 by RAG2 PHD is dispensable for RAG2 in vitro enzymatic activity, but essential for RAG2 binding to native histones.
Figure 4: Recognition of H3K4me3 is crucial for RAG1/2 recombinase activity in vivo.

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References

  1. Gellert, M. V(D)J recombination: RAG proteins, repair factors, and regulation. Annu. Rev. Biochem. 71, 101–132 (2002)

    Article  CAS  Google Scholar 

  2. Oettinger, M. A. How to keep V(D)J recombination under control. Immunol. Rev. 200, 165–181 (2004)

    Article  CAS  Google Scholar 

  3. Chowdhury, D. & Sen, R. Stepwise activation of the immunoglobulin μ heavy chain gene locus. EMBO J. 20, 6394–6403 (2001)

    Article  CAS  Google Scholar 

  4. Johnson, K., Angelin-Duclos, C., Park, S. & Calame, K. L. Changes in histone acetylation are associated with differences in accessibility of VH gene segments to V-DJ recombination during B-cell ontogeny and development. Mol. Cell. Biol. 23, 2438–2450 (2003)

    Article  CAS  Google Scholar 

  5. Morshead, K. B., Ciccone, D. N., Taverna, S. D., Allis, C. D. & Oettinger, M. A. Antigen receptor loci poised for V(D)J rearrangement are broadly associated with BRG1 and flanked by peaks of histone H3 dimethylated at lysine 4. Proc. Natl Acad. Sci. USA 100, 11577–11582 (2003)

    Article  CAS  ADS  Google Scholar 

  6. Callebaut, I. & Mornon, J. P. The V(D)J recombination activating protein RAG2 consists of a six-bladed propeller and a PHD fingerlike domain, as revealed by sequence analysis. Cell. Mol. Life Sci. 54, 880–891 (1998)

    Article  CAS  Google Scholar 

  7. Elkin, S. K. et al. A PHD finger motif in the C terminus of RAG2 modulates recombination activity. J. Biol. Chem. 280, 28701–28710 (2005)

    Article  CAS  Google Scholar 

  8. Ragvin, A. et al. Nucleosome binding by the bromodomain and PHD finger of the transcriptional cofactor p300. J. Mol. Biol. 337, 773–788 (2004)

    Article  CAS  Google Scholar 

  9. Shi, X. et al. ING2 PHD domain links histone H3 lysine 4 methylation to active gene repression. Nature 442, 96–99 (2006)

    Article  CAS  ADS  Google Scholar 

  10. Wysocka, J. et al. A PHD finger of NURF couples histone H3 lysine 4 trimethylation with chromatin remodelling. Nature 442, 86–90 (2006)

    Article  CAS  ADS  Google Scholar 

  11. West, K. L. et al. A direct interaction between the RAG2 C terminus and the core histones is required for efficient V(D)J recombination. Immunity 23, 203–212 (2005)

    Article  CAS  Google Scholar 

  12. Li, H. et al. Molecular basis for site-specific read-out of histone H3K4me3 by the BPTF PHD finger of NURF. Nature 442, 91–95 (2006)

    Article  CAS  ADS  Google Scholar 

  13. Pena, P. V. et al. Molecular mechanism of histone H3K4me3 recognition by plant homeodomain of ING2. Nature 442, 100–103 (2006)

    Article  CAS  ADS  Google Scholar 

  14. Taverna, S. D. et al. Yng1 PHD finger binding to H3 trimethylated at K4 promotes NuA3 HAT activity at K14 of H3 and transcription at a subset of targeted ORFs. Mol. Cell 24, 785–796 (2006)

    Article  CAS  Google Scholar 

  15. Ruthenburg, A. J., Allis, C. D. & Wysocka, J. Methylation of lysine 4 on histone H3: intricacy of writing and reading a single epigenetic mark. Mol. Cell 25, 15–30 (2007)

    Article  CAS  Google Scholar 

  16. Ramon-Maiques, S. et al. The PHD finger of RAG2 recognizes histone H3 methylated at both lysine-4 and arginine-2. Proc. Natl Acad. Sci. USA doi: 10.1073/pnas.0709170104 (in the press)

  17. Shi, X. et al. Proteome-wide analysis in Saccharomyces cerevisiae identifies several PHD fingers as novel direct and selective binding modules of histone H3 methylated at either lysine 4 or lysine 36. J. Biol. Chem. 282, 2450–2455 (2007)

    Article  CAS  Google Scholar 

  18. Gomez, C. A. et al. Mutations in conserved regions of the predicted RAG2 kelch repeats block initiation of V(D)J recombination and result in primary immunodeficiencies. Mol. Cell. Biol. 20, 5653–5664 (2000)

    Article  CAS  Google Scholar 

  19. Villa, A. et al. Partial V(D)J recombination activity leads to Omenn syndrome. Cell 93, 885–896 (1998)

    Article  CAS  Google Scholar 

  20. Baumann, M., Mamais, A., McBlane, F., Xiao, H. & Boyes, J. Regulation of V(D)J recombination by nucleosome positioning at recombination signal sequences. EMBO J. 22, 5197–5207 (2003)

    Article  CAS  Google Scholar 

  21. Sims, R. J. & Reinberg, D. Histone H3 Lys 4 methylation: caught in a bind? Genes Dev. 20, 2779–2786 (2006)

    Article  CAS  Google Scholar 

  22. Iwase, S. et al. The X-linked mental retardation gene SMCX/JARID1C defines a family of histone H3 lysine 4 demethylases. Cell 128, 1077–1088 (2007)

    Article  CAS  Google Scholar 

  23. Perkins, E. J., Kee, B. L. & Ramsden, D. A. Histone 3 lysine 4 methylation during the pre-B to immature B-cell transition. Nucleic Acids Res. 32, 1942–1947 (2004)

    Article  CAS  Google Scholar 

  24. Akamatsu, Y. et al. Deletion of the RAG2 C terminus leads to impaired lymphoid development in mice. Proc. Natl Acad. Sci. USA 100, 1209–1214 (2003)

    Article  CAS  ADS  Google Scholar 

  25. Corneo, B. et al. Rag mutations reveal robust alternative end joining. Nature 449, 483–486 (2007)

    Article  CAS  ADS  Google Scholar 

  26. Kirch, S. A., Rathbun, G. A. & Oettinger, M. A. Dual role of RAG2 in V(D)J recombination: catalysis and regulation of ordered Ig gene assembly. EMBO J. 17, 4881–4886 (1998)

    Article  CAS  Google Scholar 

  27. Liang, H. E. et al. The “dispensable” portion of RAG2 is necessary for efficient V-to-DJ rearrangement during B and T cell development. Immunity 17, 639–651 (2002)

    Article  CAS  Google Scholar 

  28. Shi, Y. & Whetstine, J. R. Dynamic regulation of histone lysine methylation by demethylases. Mol. Cell 25, 1–14 (2007)

    Article  CAS  Google Scholar 

  29. Tenney, K. & Shilatifard, A. A. COMPASS in the voyage of defining the role of trithorax/MLL-containing complexes: linking leukemogensis to covalent modifications of chromatin. J. Cell. Biochem. 95, 429–436 (2005)

    Article  CAS  Google Scholar 

  30. Liu, Y., Subrahmanyam, R., Chakraborty, T., Sen, R. & Desiderio, S. A plant homeodomain in Rag-2 that binds hypermethylated lysine 4 of histone H3 is necessary for efficient antigen-receptor-gene rearrangement. Immunity 4, 561–571 (2007)

    Article  Google Scholar 

  31. Matthews, A. G., Elkin, S. K. & Oettinger, M. A. Ordered DNA release and target capture in RAG transposition. EMBO J. 23, 1198–1206 (2004)

    Article  CAS  Google Scholar 

  32. Luger, K., Mader, A. W., Richmond, R. K., Sargent, D. F. & Richmond, T. J. Crystal structure of the nucleosome core particle at 2.8 A resolution. Nature 389, 251–260 (1997)

    Article  CAS  ADS  Google Scholar 

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

    Article  Google Scholar 

  34. McCoy, A. J. Solving structures of protein complexes by molecular replacement with Phaser. Acta Crystallogr. D Biol. Crystallogr. 63, 32–41 (2007)

    Article  CAS  Google Scholar 

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

    Article  Google Scholar 

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

    Article  CAS  Google Scholar 

  37. Sheldrick, G. M. & Schneider, T. R. SHELXL: High-resolution refinement. Methods Enzymol. 277, 319–343 (1997)

    Article  CAS  Google Scholar 

  38. Elkin, S. K., Matthews, A. G. & Oettinger, M. A. The C-terminal portion of RAG2 protects against transposition in vitro . EMBO J. 22, 1931–1938 (2003)

    Article  CAS  Google Scholar 

  39. Dai, Y. et al. Nonhomologous end joining and V(D)J recombination require an additional factor. Proc. Natl Acad. Sci. USA 100, 2462–2467 (2003)

    Article  CAS  ADS  Google Scholar 

  40. Gauss, G. H. & Lieber, M. R. Unequal signal and coding joint formation in human V(D)J recombination. Mol. Cell. Biol. 13, 3900–3906 (1993)

    Article  CAS  Google Scholar 

  41. Schlissel, M. S., Corcoran, L. M. & Baltimore, D. Virus-transformed pre-B cells show ordered activation but not inactivation of immunoglobulin gene rearrangement and transcription. J. Exp. Med. 173, 711–720 (1991)

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank K.-J. Armache and J.-J. Song for the generous gift of recombinant mononucleosomes; R. Kingston and members of the Kingston laboratory for helpful discussions; and N. Lau, J.-J. Song, and M. Gellert for critical reading of this manuscript. This work was supported by NIH grants (M.A.O., O.G., D.I. and T.G.K.), as well as a Korea Research Foundation grant (S.H.). O.G. is a recipient of a Burroughs Wellcome Career Award in Biomedical Sciences and a Kimmel Scholar Award. S.R.-M. has been the recipient of a fellowship from the Human Frontier Science Program. A.J.K. is funded by Stanford University through a Genentech Foundation Predoctoral Fellowship. A.G.W.M. is a Howard Hughes Medical Institute Predoctoral Fellow.

Author information

Author notes

  1. Adam G. W. Matthews, Alex J. Kuo, Or Gozani and Marjorie A. Oettinger: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Molecular Biology, Massachusetts General Hospital and Department of Genetics, Harvard Medical School, Boston, Massachusetts 02114, USA,

    Adam G. W. Matthews, Sunmi Han, Mercedes Gallardo, David N. Ciccone & Marjorie A. Oettinger

  2. Department of Biological Sciences, Stanford University, Stanford, California 94305, USA,

    Alex J. Kuo, Dylan Carney, Peggie Cheung, Kay L. Walter & Or Gozani

  3. Laboratory of Molecular Biology, NIDDK, NIH, Bethesda, Maryland 20892, USA ,

    Santiago Ramón-Maiques & Wei Yang

  4. University of Colorado Health Sciences Center, Aurora, Colorado 80045, USA ,

    Karen S. Champagne & Tatiana G. Kutateladze

  5. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115, USA,

    Dmitri Ivanov

  6. Department of Medicine, Stanford University School of Medicine, Stanford, California 94305, USA,

    Paul J. Utz

  7. Department of Pathology, Harvard Medical School, Boston, Massachusetts 02115, USA,

    Yang Shi

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Correspondence to Or Gozani or Marjorie A. Oettinger.

Supplementary information

Supplementary Information

The file contains Supplementary Figures S1-S13 with Legends, Supplementary Table 1 and additional references. The Supplementary Figures support the general conclusion that the RAG2PHD-H3K4me3 interaction plays an important role in regulating V(D)J recombination, Supplementary Figure 1 providing a model for how H3K4me3-binding by RAG2 influences V(D)J recombination. (PDF 2669 kb)

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Matthews, A., Kuo, A., Ramón-Maiques, S. et al. RAG2 PHD finger couples histone H3 lysine 4 trimethylation with V(D)J recombination. Nature 450, 1106–1110 (2007). https://doi.org/10.1038/nature06431

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