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Chemical tagging and customizing of cellular chromatin states using ultrafast trans-splicing inteins

Nature Chemistry volume 7, pages 394–402 (2015)Cite this article

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Abstract

Post-translational modification of the histone proteins in chromatin plays a central role in the epigenetic control of DNA-templated processes in eukaryotic cells. Developing methods that enable the structure of histones to be manipulated is, therefore, essential to understand the biochemical mechanisms that underlie genomic regulation. Here we present a synthetic biology method to engineer histones that bear site-specific modifications on cellular chromatin using protein trans-splicing (PTS). We genetically fused the N-terminal fragment of ultrafast split intein to the C terminus of histone H2B, which, on reaction with a complementary synthetic C intein, generated labelled histone. Using this approach, we incorporated various non-native chemical modifications into chromatin in vivo with temporal control. Furthermore, the time and concentration dependence of PTS performed in nucleo enabled us to examine differences in the accessibility of the euchromatin and heterochromatin regions of the epigenome. Finally, we used PTS to semisynthesize a native histone modification, H2BK120 ubiquitination, in isolated nuclei and showed that this can trigger downstream epigenetic crosstalk of H3K79 methylation.

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Figure 1: Modification of native chromatin using PTS.
Figure 2: Modification of histone H2B in native chromatin.
Figure 3: PTS as a function of chromatin state.
Figure 4: Synthesis of IntC–H2B–K120Ub (construct 6).
Figure 5: In nucleo semisynthesis of H2B–K120Ub and its effect on H3K79 methylation.

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Acknowledgements

The authors thank the current and former members of the Muir laboratory for many valuable discussions. We further thank D. H. Perlman (Princeton Proteomics and Mass Spectrometry Core, Princeton University) for the mass spectrometry data and G. Laevsky (Confocal Core Facility, Princeton University) for help with the microscopy experiments. We also thank W. Wang, D. Storton and J. Miller (Microarray Facility, Princeton University) for help with the qPCR and RNA-seq experiments. Finally, we thank S. Josefowicz, C. Li and D. Allis (Rockefeller University) for help with the ChIP assays. This research was supported by the US National Institutes of Health (grants R37-GM086868 and R01 GM107047).

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Authors and Affiliations

  1. Department of Chemistry, Princeton University, Frick Laboratory, Princeton, 08544, New Jersey, USA

    Yael David, Miquel Vila-Perelló, Shivam Verma & Tom W. Muir

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  1. Yael David
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  2. Miquel Vila-Perelló
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Contributions

Y.D., M.V-P. and T.W.M. conceived and designed the research. Y.D., M.V-P. and S.V. prepared the reagents and performed the experiments. Y.D. and T.W.M. wrote the manuscript.

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Correspondence to Tom W. Muir.

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David, Y., Vila-Perelló, M., Verma, S. et al. Chemical tagging and customizing of cellular chromatin states using ultrafast trans-splicing inteins. Nature Chem 7, 394–402 (2015). https://doi.org/10.1038/nchem.2224

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