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Uncovering global SUMOylation signaling networks in a site-specific manner

Nature Structural & Molecular Biology volume 21, pages 927936 (2014) | Download Citation

Abstract

SUMOylation is a reversible post-translational modification essential for genome stability. Using high-resolution MS, we have studied global SUMOylation in human cells in a site-specific manner, identifying a total of >4,300 SUMOylation sites in >1,600 proteins. To our knowledge, this is the first time that >1,000 SUMOylation sites have been identified under standard growth conditions. We quantitatively studied SUMOylation dynamics in response to SUMO protease inhibition, proteasome inhibition and heat shock. Many SUMOylated lysines have previously been reported to be ubiquitinated, acetylated or methylated, thus indicating cross-talk between SUMO and other post-translational modifications. We identified 70 phosphorylation and four acetylation events in proximity to SUMOylation sites, and we provide evidence for acetylation-dependent SUMOylation of endogenous histone H3. SUMOylation regulates target proteins involved in all nuclear processes including transcription, DNA repair, chromatin remodeling, precursor-mRNA splicing and ribosome assembly.

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Acknowledgements

The authors are grateful for support from the European Research Council, grant 310913 (A.C.O.V.), the Netherlands Organization for Scientific Research (NWO), grants 70058425, 93511037 and 70059006 (A.C.O.V.), and the Max-Planck Society for the Advancement of Science (M.M.). We would like to acknowledge J. Cox for his help with MaxQuant and K. Sharma for helpful discussions. We would like to acknowledge our colleagues A.G. Jochemsen and D. Baker (both at Leiden University Medical Center) for reagents and helpful discussions.

Author information

Affiliations

  1. Department of Molecular Cell Biology, Leiden University Medical Center, Leiden, the Netherlands.

    • Ivo A Hendriks
    • , Bing Yang
    • , Matty Verlaan-de Vries
    •  & Alfred C O Vertegaal
  2. Department of Proteomics and Signal Transduction, Max Planck Institute for Biochemistry, Martinsried, Germany.

    • Rochelle C J D'Souza
    •  & Matthias Mann

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Contributions

A.C.O.V. and I.A.H. conceived the biochemical methodology and designed the experiments. I.A.H. optimized the biochemical methodology and prepared all MS samples. M.M. supervised the initial MS experiments and the optimization of the MS configuration performed by R.C.J.D. and I.A.H.; B.Y. and I.A.H. performed further optimization of the MS configuration. B.Y. and R.C.J.D. operated Q-Exactive machines. I.A.H. processed the MS data and performed bioinformatics analysis. I.A.H. and M.V.V. prepared biochemical samples and performed immunoblotting experiments. A.C.O.V. conceived the project. A.C.O.V. and M.M. supervised the project. I.A.H. and A.C.O.V. wrote the manuscript with input from all authors.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Alfred C O Vertegaal.

Integrated supplementary information

Supplementary information

PDF files

  1. 1.

    Supplementary Text and Figures

    Supplementary Figures 1–7 and Supplementary Note

  2. 2.

    Supplementary Data Set 1

    A compilation of all uncropped images corresponding to all scans of gels, membranes and films displayed throughout this manuscript

Excel files

  1. 1.

    Supplementary Table 1

    A complete list of all (4,361) identified SUMO sites and all (5,339) identified SUMOylated peptides

  2. 2.

    Supplementary Table 2

    A complete list of all identified SUMOylated proteins and a list of putative SUMOylated proteins identified by only non-SUMOylated peptides

  3. 3.

    Supplementary Table 3

    Label-free quantification of SUMOylated proteins in response to MG-132, PR-619 or heat shock

  4. 4.

    Supplementary Table 4

    An overlap matrix of all SUMOylated proteins identified in this work, as compared to SUMOylated proteins previously identified in other studies

  5. 5.

    Supplementary Table 5

    An overlap matrix of all identified SUMO sites, along with previously identified SUMO sites, and all known ubiquitination, acetylation and lysine-methylation sites

  6. 6.

    Supplementary Table 6

    A list of all SUMOylated enzymes involved in phosphorylation, ubiquitination, methylation and acetylation

  7. 7.

    Supplementary Table 7

    A list of SUMOylated peptides exclusively or nonexclusively detected in the presence of acetylation or phosphorylation

  8. 8.

    Supplementary Table 8

    A complete term enrichment analysis on all SUMOylated proteins, as compared to the human proteome

  9. 9.

    Supplementary Table 9

    A fully annotated list of all SUMOylated proteins

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DOI

https://doi.org/10.1038/nsmb.2890

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