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Monoubiquitinated H2B is associated with the transcribed region of highly expressed genes in human cells

Nature Cell Biology volume 10, pages 483–488 (2008)Cite this article

Abstract

Histone modifications have emerged as important regulators of transcription1,2. Histone H2B monoubiquitination has also been implicated in transcription3,4; however, better understanding of the biological significance of this modification in mammalian cells has been hindered by the lack of suitable reagents, particularly antibodies capable of specifically recognizing ubiquitinated H2B (ubH2B). Here, we report the generation of anti-ubH2B monoclonal antibodies using a branched peptide as immunogen. These antibodies provide a powerful tool for exploring the biochemical functions of H2B monoubiquitination at both a genome-wide and gene-specific level. Application of these antibodies in high resolution chromatin immunoprecipitation (ChIP)-chip experiments in human cells, using tiling arrays, revealed preferential association of ubiquitinated H2B with the transcribed regions of highly expressed genes. Unlike dimethylated H3K4, ubH2B was not associated with distal promoter regions. Furthermore, experimental modulation of the transcriptional activity of the tumour suppressor p53 was accompanied by rapid changes in the H2B ubiquitination status of its p21 target gene, attesting to the dynamic nature of this process. It has recently been demonstrated that the apparent extent of gene expression often reflects elongation rather than initiation rates5; thus, our findings suggest that H2B ubiquitination is intimately linked with global transcriptional elongation in mammalian cells.

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Figure 1: Generation and validation of anti-ubH2B antibodies.
Figure 2: Analysis of ubH2B and H3K4me2 patterns around transcription start sites.
Figure 3: ubH2B is associated with the transcribed region of highly expressed genes.
Figure 4: H2B ubiquitination of the p21 gene correlates with p53-dependent transcriptional activation.

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References

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

    Article  CAS  PubMed  Google Scholar 

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

    Article  CAS  PubMed  Google Scholar 

  3. Osley, M. A. Regulation of histone H2A and H2B ubiquitylation. Brief Funct. Genomic Proteomic 5, 179–189 (2006).

    Article  CAS  PubMed  Google Scholar 

  4. Shilatifard, A. Chromatin modifications by methylation and ubiquitination: implications in the regulation of gene expression. Annu. Rev. Biochem. 75, 243–269 (2006).

    Article  CAS  PubMed  Google Scholar 

  5. Guenther, M. G., Levine, S. S., Boyer, L. A., Jaenisch, R. & Young, R. A. A chromatin landmark and transcription initiation at most promoters in human cells. Cell 130, 77–88 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Davie, J. R. & Murphy, L. C. Inhibition of transcription selectively reduces the level of ubiquitinated histone H2B in chromatin. Biochem. Biophys. Res. Commun. 203, 344–350 (1994).

    Article  CAS  PubMed  Google Scholar 

  7. Briggs, S. D. et al. Histone H3 lysine 4 methylation is mediated by Set1 and required for cell growth and rDNA silencing in Saccharomyces cerevisiae. Genes Dev. 15, 3286–3295 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Lee, K. K., Florens, L., Swanson, S. K., Washburn, M. P. & Workman, J. L. The deubiquitylation activity of Ubp8 is dependent upon Sgf11 and its association with the SAGA complex. Mol. Cell Biol. 25, 1173–1182 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Mutiu, A. I., Hoke, S. M., Genereaux, J., Liang, G. & Brandl, C. J. The role of histone ubiquitylation and deubiquitylation in gene expression as determined by the analysis of an HTB1(K123R) Saccharomyces cerevisiae strain. Mol. Genet. Genomics 277, 491–506 (2007).

    Article  CAS  PubMed  Google Scholar 

  10. Sun, Z. W. & Allis, C. D. Ubiquitination of histone H2B regulates H3 methylation and gene silencing in yeast. Nature 418, 104–108 (2002).

    Article  CAS  PubMed  Google Scholar 

  11. Henry, K. W. et al. Transcriptional activation via sequential histone H2B ubiquitylation and deubiquitylation, mediated by SAGA-associated Ubp8. Genes Dev. 17, 2648–2663 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Shukla, A., Stanojevic, N., Duan, Z., Shadle, T. & Bhaumik, S.R. Functional analysis of H2B-Lys-123 ubiquitination in regulation of H3-Lys-4 methylation and recruitment of RNA polymerase II at the coding sequences of several active genes in vivo. J. Biol. Chem. 281, 19045–19054 (2006).

    Article  CAS  PubMed  Google Scholar 

  13. Xiao, T. et al. Histone H2B ubiquitylation is associated with elongating RNA polymerase II. Mol. Cell Biol. 25, 637–651 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Kao, C. F. et al. Rad6 plays a role in transcriptional activation through ubiquitylation of histone H2B. Genes Dev. 18, 184–195 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Wyce, A. et al. H2B ubiquitylation acts as a barrier to Ctk1 nucleosomal recruitment prior to removal by Ubp8 within a SAGA-related complex. Mol. Cell 27, 275–288 (2007).

    Article  CAS  PubMed  Google Scholar 

  16. Emre, N. C. et al. Maintenance of low histone ubiquitylation by Ubp10 correlates with telomere-proximal Sir2 association and gene silencing. Mol. Cell 17, 585–594 (2005).

    Article  CAS  PubMed  Google Scholar 

  17. Marzluff, W. F., Gongidi, P., Woods, K. R., Jin, J. & Maltais, L. J. The human and mouse replication-dependent histone genes. Genomics 80, 487–498 (2002).

    Article  CAS  PubMed  Google Scholar 

  18. Pavri, R. et al. Histone H2B monoubiquitination functions cooperatively with FACT to regulate elongation by RNA polymerase II. Cell 125, 703–717 (2006).

    Article  CAS  PubMed  Google Scholar 

  19. Thorne, A. W., Sautiere, P., Briand, G. & Crane-Robinson, C. The structure of ubiquitinated histone H2B. EMBO J. 6, 1005–1010 (1987).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Kim, J., Hake, S. B. & Roeder, R. G. The human homolog of yeast BRE1 functions as a transcriptional coactivator through direct activator interactions. Mol. Cell 20, 759–770 (2005).

    Article  CAS  PubMed  Google Scholar 

  21. Plaue, S., Muller, S. & van Regenmortel, M. H. A branched, synthetic octapeptide of ubiquitinated histone H2A as target of autoantibodies. J. Exp. Med. 169, 1607–1617 (1989).

    Article  CAS  PubMed  Google Scholar 

  22. Vassilev, A. P., Rasmussen, H. H., Christensen, E. I., Nielsen, S. & Celis, J. E. The levels of ubiquitinated histone H2A are highly upregulated in transformed human cells: partial colocalization of uH2A clusters and PCNA/cyclin foci in a fraction of cells in S-phase. J. Cell Sci. 108, 1205–1215 (1995).

    CAS  PubMed  Google Scholar 

  23. Mimnaugh, E. G., Chen, H. Y., Davie, J. R., Celis, J. E. & Neckers, L. Rapid deubiquitination of nucleosomal histones in human tumor cells caused by proteasome inhibitors and stress response inducers: effects on replication, transcription, translation, and the cellular stress response. Biochemistry 36, 14418–14429 (1997).

    Article  CAS  PubMed  Google Scholar 

  24. Lis, E. T. & Romesberg, F. E. Role of Doa1 in the Saccharomyces cerevisiae DNA damage response. Mol. Cell Biol. 26, 4122–4133 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Shahbazian, M. D., Zhang, K. & Grunstein, M. Histone H2B ubiquitylation controls processive methylation but not monomethylation by Dot1 and Set1. Mol. Cell 19, 271–277 (2005).

    Article  CAS  PubMed  Google Scholar 

  26. Raver-Shapira, N. et al. Transcriptional activation of miR-34a contributes to p53-mediated apoptosis. Mol. Cell 26, 731–743 (2007).

    Article  CAS  PubMed  Google Scholar 

  27. West, M. H. & Bonner, W. M. Histone 2B can be modified by the attachment of ubiquitin. Nucleic Acids Res. 8, 4671–4680 (1980).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Nicassio, F. et al. Human USP3 is a chromatin modifier required for S phase progression and genome stability. Curr. Biol. 17, 1972–1977 (2007).

    Article  CAS  PubMed  Google Scholar 

  29. Sridhar, V. V. et al. Control of DNA methylation and heterochromatic silencing by histone H2B deubiquitination. Nature 447, 735–738 (2007).

    Article  CAS  PubMed  Google Scholar 

  30. Minsky, N. & Oren, M. The RING domain of Mdm2 mediates histone ubiquitylation and transcriptional repression. Mol. Cell 16, 631–639 (2004).

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

We are grateful to M. Fridkin for peptide design and synthesis, O. Leitner and A. Bren for immunizations and hybridoma fusion and growth, and S. Horn-Saban for DNA amplification and array hybridization. N.M. thanks R. Roeder for his support and encouragement. Supported in part by grant R37 CA40099 from the National Cancer Institute, a Prostate Cancer Foundation (Israel) Center of Excellence, the Dr. Miriam and Sheldon Adelson Medical Research Foundation, and the Yad Abraham Center for Cancer Diagnosis and Therapy.

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  1. Neri Minsky

    Present address: Current address: Laboratory of Biochemistry and Molecular Biology, The Rockefeller University, New York, NY 10021, USA.,

Authors and Affiliations

  1. Department of Molecular Cell Biology, The Weizmann Institute of Science, 76100, Rehovot, Israel

    Neri Minsky, Efrat Shema & Moshe Oren

  2. Department of Computer Science and Applied Mathematics, The Weizmann Institute of Science, 76100, Rehovot, Israel

    Yair Field, Meromit Schuster & Eran Segal

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Contributions

N. M. and E, Sh. performed the experiments; Y. F. and M. S. performed he computational analysis; M. O. and E. Se. supervised the work. All authors contributed to the writing of the manuscript.

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Correspondence to Moshe Oren.

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Minsky, N., Shema, E., Field, Y. et al. Monoubiquitinated H2B is associated with the transcribed region of highly expressed genes in human cells. Nat Cell Biol 10, 483–488 (2008). https://doi.org/10.1038/ncb1712

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