Department of Molecular, Cell, and Cancer Biology; Department of Biochemistry and Molecular Pharmacology
Amino Acids, Peptides, and Proteins | Cells | Genetic Phenomena | Investigative Techniques | Molecular Biology
The repeating subunit of chromatin, the nucleosome, includes two copies of each of the four core histones, and several recent studies have reported that asymmetrically modified nucleosomes occur at regulatory elements in vivo. To probe the mechanisms by which histone modifications are read out, we designed an obligate pair of H3 heterodimers, termed H3X and H3Y, which we validated genetically and biochemically. Comparing the effects of asymmetric histone tail point mutants with those of symmetric double mutants revealed that a single methylated H3K36 per nucleosome was sufficient to silence cryptic transcription in vivo. We also demonstrate the utility of this system for analysis of histone modification crosstalk, using mass spectrometry to separately identify modifications on both H3 molecules within asymmetric nucleosomes. The ability to generate asymmetric nucleosomes in vivo provides a powerful tool to probe the mechanism by which H3 tails are read out by effector proteins in the cell.
molecular biology, nucleosome symmetry, histones, heterodimers, asymmetric nucleosomes
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The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It is made available under a CC-BY-NC-ND 4.0 International license.
DOI of Published Version
bioRxiv 170811; doi: https://doi.org/10.1101/170811. Link to preprint on bioRxiv service.
Now published in eLife doi: 10.7554/eLife.28836.
Ichikawa Y, Chen Y, Bajaj V, Connolly CM, Chou H, Sharma U, Chen HV, Bolon DN, Rando OJ, Kaufman PD. (2017). A synthetic biology approach to probing nucleosome symmetry [preprint]. University of Massachusetts Medical School Faculty Publications. https://doi.org/10.1101/170811. Retrieved from https://escholarship.umassmed.edu/faculty_pubs/1557
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