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In situ chromatin interactomics using a chemical bait and trap approach

Abstract

Elucidating the physiological binding partners of histone post-translational modifications (hPTMs) is key to understanding fundamental epigenetic regulatory pathways. Determining such interactomes will enable the study of how perturbations of these interactions affect disease. Here we use a synthetic biology approach to set a series of hPTM-controlled photo-affinity traps in native chromatin. Using quantitative proteomics, the local interactomes of these chemically customized chromatin landscapes are determined. We show that the approach captures transiently interacting factors such as methyltransferases and demethylases, as well as previously reported and novel hPTM reader proteins. We also apply this in situ proteomics approach to a recently disclosed cancer-associated histone mutation, H3K4M, revealing a number of perturbed interactions with the mutated tail. Collectively our studies demonstrate that modifying and interrogating native chromatin with chemical precision is a powerful tool for exploring epigenetic regulation and dysregulation at the molecular level.

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Fig. 1: Schematic for the in situ chromatin interactomics approach.
Fig. 2: In situ installation of hPTMs with ultra-fast split inteins.
Fig. 3: Determining the in situ interactome of H3K9me3.
Fig. 4: Determining the interactome of H3K4 as a function of hPTMs.

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Data availability

All relevant data are included in the manuscript and supplementary information. Mass spectrometry data files have been uploaded to the PRIDE proteomics database (PXD017447).

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Acknowledgements

We thank F. Wojcik and R. Thompson and other members of the Allis and Muir laboratories for valuable discussions. We thank T. Srikumar, S. Kyin and H. Shwe from the Princeton Proteomics and Mass Spectrometry Core. A.J.B. is a Damon Runyon Fellow of the Damon Runyon Cancer Research Foundation (DRG-2283-17). J.D.B. was funded by a postdoctoral fellowship from the US National Institute of Health (GM123659). This work was supported by the US National Institutes of Health (NIH grants R37-GM086868 to T.W.M. and PO1-CA196539 to C.D.A. and T.W.M.).

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Contributions

A.J.B. generated the modified histones in isolated nuclei and performed the cross-linking workflows. A.J.B. and M.H. generated the SILAC cell lines and performed fluorescence anisotropy experiments. A.J.B. and J.D.B. analysed the proteomics data. L.A.G. and C.D.A. contributed reagents and to data analysis. A.J.B. and T.W.M. conceived the project, analysed all data and wrote the manuscript with input from the other authors.

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Correspondence to Tom W. Muir.

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Supplementary information

Supplementary Information

Supplementary figures and Tables 1–28 (including uncropped gels displayed in Figs. 2–4).

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Supplementary Data

Combined mass spectrometry datasets for the SILAC-based quantitative proteomics performed in this study.

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Burton, A.J., Haugbro, M., Gates, L.A. et al. In situ chromatin interactomics using a chemical bait and trap approach. Nat. Chem. 12, 520–527 (2020). https://doi.org/10.1038/s41557-020-0474-8

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