A synthetic biology approach to probing nucleosome symmetry
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 d...
| Main Authors: | , , , , , , , , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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eLife Sciences Publications Ltd
2017-09-01
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| Series: | eLife |
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| Online Access: | https://elifesciences.org/articles/28836 |
| _version_ | 1811181381481398272 |
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| author | Yuichi Ichikawa Caitlin F Connelly Alon Appleboim Thomas CR Miller Hadas Jacobi Nebiyu A Abshiru Hsin-Jung Chou Yuanyuan Chen Upasna Sharma Yupeng Zheng Paul M Thomas Hsuiyi V Chen Vineeta Bajaj Christoph W Müller Neil L Kelleher Nir Friedman Daniel NA Bolon Oliver J Rando Paul D Kaufman |
| author_facet | Yuichi Ichikawa Caitlin F Connelly Alon Appleboim Thomas CR Miller Hadas Jacobi Nebiyu A Abshiru Hsin-Jung Chou Yuanyuan Chen Upasna Sharma Yupeng Zheng Paul M Thomas Hsuiyi V Chen Vineeta Bajaj Christoph W Müller Neil L Kelleher Nir Friedman Daniel NA Bolon Oliver J Rando Paul D Kaufman |
| author_sort | Yuichi Ichikawa |
| collection | DOAJ |
| description | 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 extensively 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 each H3 molecule within asymmetric nucleosomes. The ability to generate asymmetric nucleosomes in vivo and in vitro provides a powerful and generalizable tool to probe the mechanisms by which H3 tails are read out by effector proteins in the cell. |
| first_indexed | 2024-04-11T09:16:48Z |
| format | Article |
| id | doaj.art-24b6ece5ed4441b78cfba5f0e8e8b723 |
| institution | Directory Open Access Journal |
| issn | 2050-084X |
| language | English |
| last_indexed | 2024-04-11T09:16:48Z |
| publishDate | 2017-09-01 |
| publisher | eLife Sciences Publications Ltd |
| record_format | Article |
| series | eLife |
| spelling | doaj.art-24b6ece5ed4441b78cfba5f0e8e8b7232022-12-22T04:32:18ZengeLife Sciences Publications LtdeLife2050-084X2017-09-01610.7554/eLife.28836A synthetic biology approach to probing nucleosome symmetryYuichi Ichikawa0Caitlin F Connelly1Alon Appleboim2Thomas CR Miller3Hadas Jacobi4Nebiyu A Abshiru5Hsin-Jung Chou6Yuanyuan Chen7Upasna Sharma8Yupeng Zheng9Paul M Thomas10Hsuiyi V Chen11Vineeta Bajaj12Christoph W Müller13Neil L Kelleher14https://orcid.org/0000-0002-8815-3372Nir Friedman15https://orcid.org/0000-0002-9678-3550Daniel NA Bolon16Oliver J Rando17https://orcid.org/0000-0003-1516-9397Paul D Kaufman18https://orcid.org/0000-0003-3089-313XDepartment of Molecular, Cell and Cancer Biology, University of Massachusetts Medical School, Worcester, United StatesDepartment of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, United StatesSchool of Computer Science and Engineering, The Hebrew University of Jerusalem, Jerusalem, Israel; The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem, IsraelStructural and Computational Biology Unit, European Molecular Biology Laboratory, Heidelberg, GermanySchool of Computer Science and Engineering, The Hebrew University of Jerusalem, Jerusalem, Israel; The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem, IsraelNational Resource for Translational and Developmental Proteomics, Northwestern University, Evanston, United StatesDepartment of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, United StatesDepartment of Molecular, Cell and Cancer Biology, University of Massachusetts Medical School, Worcester, United StatesDepartment of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, United StatesNational Resource for Translational and Developmental Proteomics, Northwestern University, Evanston, United StatesNational Resource for Translational and Developmental Proteomics, Northwestern University, Evanston, United StatesDepartment of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, United StatesDepartment of Molecular, Cell and Cancer Biology, University of Massachusetts Medical School, Worcester, United StatesStructural and Computational Biology Unit, European Molecular Biology Laboratory, Heidelberg, GermanyNational Resource for Translational and Developmental Proteomics, Northwestern University, Evanston, United StatesSchool of Computer Science and Engineering, The Hebrew University of Jerusalem, Jerusalem, Israel; The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem, IsraelDepartment of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, United StatesDepartment of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, United StatesDepartment of Molecular, Cell and Cancer Biology, University of Massachusetts Medical School, Worcester, United StatesThe 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 extensively 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 each H3 molecule within asymmetric nucleosomes. The ability to generate asymmetric nucleosomes in vivo and in vitro provides a powerful and generalizable tool to probe the mechanisms by which H3 tails are read out by effector proteins in the cell.https://elifesciences.org/articles/28836chromatintranscriptionsynthetic biology |
| spellingShingle | Yuichi Ichikawa Caitlin F Connelly Alon Appleboim Thomas CR Miller Hadas Jacobi Nebiyu A Abshiru Hsin-Jung Chou Yuanyuan Chen Upasna Sharma Yupeng Zheng Paul M Thomas Hsuiyi V Chen Vineeta Bajaj Christoph W Müller Neil L Kelleher Nir Friedman Daniel NA Bolon Oliver J Rando Paul D Kaufman A synthetic biology approach to probing nucleosome symmetry eLife chromatin transcription synthetic biology |
| title | A synthetic biology approach to probing nucleosome symmetry |
| title_full | A synthetic biology approach to probing nucleosome symmetry |
| title_fullStr | A synthetic biology approach to probing nucleosome symmetry |
| title_full_unstemmed | A synthetic biology approach to probing nucleosome symmetry |
| title_short | A synthetic biology approach to probing nucleosome symmetry |
| title_sort | synthetic biology approach to probing nucleosome symmetry |
| topic | chromatin transcription synthetic biology |
| url | https://elifesciences.org/articles/28836 |
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