Rapid reconstitution of ubiquitinated nucleosome using a non-denatured histone octamer ubiquitylation approach

Weijie Li; Peirong Cao; Pengqi Xu; Fahui Sun; Chi Wang; Jiale Zhang; Shuqi Dong; Jon R. Wilson; Difei Xu; Hengxin Fan; Zhenhuan Feng; Xiaofei Zhang; Qingjun Zhu; Yingzhi Fan; Nick Brown; Neil Justin; Steven J. Gamblin; He Li; Ying Zhang; Jun He

doi:10.1186/s13578-024-01265-x

Rapid reconstitution of ubiquitinated nucleosome using a non-denatured histone octamer ubiquitylation approach

Weijie Li, Peirong Cao, Pengqi Xu, Fahui Sun, Chi Wang, Jiale Zhang, Shuqi Dong, Jon R. Wilson, Difei Xu, Hengxin Fan, Zhenhuan Feng, Xiaofei Zhang, Qingjun Zhu, Yingzhi Fan, Nick Brown, Neil Justin, Steven J. Gamblin, He Li, Ying Zhang^*, Jun He^*

^*Corresponding author for this work

Research output: Contribution to journal › Journal article › peer-review

1 Citation (Scopus)

Abstract

Background: Histone ubiquitination modification is emerging as a critical epigenetic mechanism involved in a range of biological processes. In vitro reconstitution of ubiquitinated nucleosomes is pivotal for elucidating the influence of histone ubiquitination on chromatin dynamics. Results: In this study, we introduce a Non-Denatured Histone Octamer Ubiquitylation (NDHOU) approach for generating ubiquitin or ubiquitin-like modified histone octamers. The method entails the co-expression and purification of histone octamers, followed by their chemical cross-linking to ubiquitin using 1,3-dibromoacetone. We demonstrate that nucleosomes reconstituted with these octamers display a high degree of homogeneity, rendering them highly compatible with in vitro biochemical assays. These ubiquitinated nucleosomes mimic physiological substrates in function and structure. Additionally, we have extended this method to cross-linking various histone octamers and three types of ubiquitin-like proteins. Conclusions: Overall, our findings offer an efficient strategy for producing ubiquitinated nucleosomes, advancing biochemical and biophysical studies in the field of chromatin biology.

Original language	English
Article number	81
Number of pages	15
Journal	Cell and Bioscience
Volume	14
Issue number	1
DOIs	https://doi.org/10.1186/s13578-024-01265-x
Publication status	Published - 17 Jun 2024
Externally published	Yes

Scopus Subject Areas

General Biochemistry,Genetics and Molecular Biology

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.1186/s13578-024-01265-x

Cite this

Li, W., Cao, P., Xu, P., Sun, F., Wang, C., Zhang, J., Dong, S., Wilson, J. R., Xu, D., Fan, H., Feng, Z., Zhang, X., Zhu, Q., Fan, Y., Brown, N., Justin, N., Gamblin, S. J., Li, H., Zhang, Y., & He, J. (2024). Rapid reconstitution of ubiquitinated nucleosome using a non-denatured histone octamer ubiquitylation approach. Cell and Bioscience, 14(1), Article 81. https://doi.org/10.1186/s13578-024-01265-x

@article{bef7554cbfe84ad8af4e9bec27dc0b23,

title = "Rapid reconstitution of ubiquitinated nucleosome using a non-denatured histone octamer ubiquitylation approach",

abstract = "Background: Histone ubiquitination modification is emerging as a critical epigenetic mechanism involved in a range of biological processes. In vitro reconstitution of ubiquitinated nucleosomes is pivotal for elucidating the influence of histone ubiquitination on chromatin dynamics. Results: In this study, we introduce a Non-Denatured Histone Octamer Ubiquitylation (NDHOU) approach for generating ubiquitin or ubiquitin-like modified histone octamers. The method entails the co-expression and purification of histone octamers, followed by their chemical cross-linking to ubiquitin using 1,3-dibromoacetone. We demonstrate that nucleosomes reconstituted with these octamers display a high degree of homogeneity, rendering them highly compatible with in vitro biochemical assays. These ubiquitinated nucleosomes mimic physiological substrates in function and structure. Additionally, we have extended this method to cross-linking various histone octamers and three types of ubiquitin-like proteins. Conclusions: Overall, our findings offer an efficient strategy for producing ubiquitinated nucleosomes, advancing biochemical and biophysical studies in the field of chromatin biology.",

author = "Weijie Li and Peirong Cao and Pengqi Xu and Fahui Sun and Chi Wang and Jiale Zhang and Shuqi Dong and Wilson, {Jon R.} and Difei Xu and Hengxin Fan and Zhenhuan Feng and Xiaofei Zhang and Qingjun Zhu and Yingzhi Fan and Nick Brown and Neil Justin and Gamblin, {Steven J.} and He Li and Ying Zhang and Jun He",

note = "This work was supported by the National Natural Science Foundation of China (31900429), Shenzhen Science and Technology Innovation Committee (grant no. JCYJ20190809151611269), Shenzhen Sanming Project of Medicine (SZSM201911003), Shenzhen Science Technology and Innovation Commission (SZSTI) Basic Research Program (JCYJ20190809165417340) and by 100 Top Talents Program of Sun Yat-sen University (grant no. 392009), Natural Science Foundation of Guangdong Province, China (2021A1515011070); Guang Dong Basic and Applied Basic Research Foundation (2019A1515110812); Shenzhen Science and Technology Innovation Committee (JCYJ20210324120601005); Guang Dong Basic and Applied Basic Research Foundation (2020A1515110060);.Funding information National Natural Science Foundation of China (grant No. U20A2013, 32241021, 32170189); China Postdoctoral Science Foundation (grant No. 2023M743512); Nation-sponsored Postdoctoral Researcher program (grant No. GZC20232691); Guangzhou Science and Technology Plan Project Basic Research Plan (grant No. 2023A04J1076,202201011516); International (Regional) Cooperation and Exchange (ICE) Projects of the National Natural Science Foundation of China (NSFC) (grant No. 32361163669); R&D Program of Guangzhou Laboratory (award No. SRPG22-003 ); Science and Technology Planning Project of Guangdong Province (grant No. 2023B1212060050). Publisher Copyright: {\textcopyright} The Author(s) 2024.",

year = "2024",

month = jun,

day = "17",

doi = "10.1186/s13578-024-01265-x",

language = "English",

volume = "14",

journal = "Cell and Bioscience",

issn = "2045-3701",

publisher = "BioMed Central Ltd.",

number = "1",

}

Li, W, Cao, P, Xu, P, Sun, F, Wang, C, Zhang, J, Dong, S, Wilson, JR, Xu, D, Fan, H, Feng, Z, Zhang, X, Zhu, Q, Fan, Y, Brown, N, Justin, N, Gamblin, SJ, Li, H, Zhang, Y & He, J 2024, 'Rapid reconstitution of ubiquitinated nucleosome using a non-denatured histone octamer ubiquitylation approach', Cell and Bioscience, vol. 14, no. 1, 81. https://doi.org/10.1186/s13578-024-01265-x

TY - JOUR

T1 - Rapid reconstitution of ubiquitinated nucleosome using a non-denatured histone octamer ubiquitylation approach

AU - Li, Weijie

AU - Cao, Peirong

AU - Xu, Pengqi

AU - Sun, Fahui

AU - Wang, Chi

AU - Zhang, Jiale

AU - Dong, Shuqi

AU - Wilson, Jon R.

AU - Xu, Difei

AU - Fan, Hengxin

AU - Feng, Zhenhuan

AU - Zhang, Xiaofei

AU - Zhu, Qingjun

AU - Fan, Yingzhi

AU - Brown, Nick

AU - Justin, Neil

AU - Gamblin, Steven J.

AU - Li, He

AU - Zhang, Ying

AU - He, Jun

N1 - This work was supported by the National Natural Science Foundation of China (31900429), Shenzhen Science and Technology Innovation Committee (grant no. JCYJ20190809151611269), Shenzhen Sanming Project of Medicine (SZSM201911003), Shenzhen Science Technology and Innovation Commission (SZSTI) Basic Research Program (JCYJ20190809165417340) and by 100 Top Talents Program of Sun Yat-sen University (grant no. 392009), Natural Science Foundation of Guangdong Province, China (2021A1515011070); Guang Dong Basic and Applied Basic Research Foundation (2019A1515110812); Shenzhen Science and Technology Innovation Committee (JCYJ20210324120601005); Guang Dong Basic and Applied Basic Research Foundation (2020A1515110060);.Funding information National Natural Science Foundation of China (grant No. U20A2013, 32241021, 32170189); China Postdoctoral Science Foundation (grant No. 2023M743512); Nation-sponsored Postdoctoral Researcher program (grant No. GZC20232691); Guangzhou Science and Technology Plan Project Basic Research Plan (grant No. 2023A04J1076,202201011516); International (Regional) Cooperation and Exchange (ICE) Projects of the National Natural Science Foundation of China (NSFC) (grant No. 32361163669); R&D Program of Guangzhou Laboratory (award No. SRPG22-003 ); Science and Technology Planning Project of Guangdong Province (grant No. 2023B1212060050). Publisher Copyright: © The Author(s) 2024.

PY - 2024/6/17

Y1 - 2024/6/17

N2 - Background: Histone ubiquitination modification is emerging as a critical epigenetic mechanism involved in a range of biological processes. In vitro reconstitution of ubiquitinated nucleosomes is pivotal for elucidating the influence of histone ubiquitination on chromatin dynamics. Results: In this study, we introduce a Non-Denatured Histone Octamer Ubiquitylation (NDHOU) approach for generating ubiquitin or ubiquitin-like modified histone octamers. The method entails the co-expression and purification of histone octamers, followed by their chemical cross-linking to ubiquitin using 1,3-dibromoacetone. We demonstrate that nucleosomes reconstituted with these octamers display a high degree of homogeneity, rendering them highly compatible with in vitro biochemical assays. These ubiquitinated nucleosomes mimic physiological substrates in function and structure. Additionally, we have extended this method to cross-linking various histone octamers and three types of ubiquitin-like proteins. Conclusions: Overall, our findings offer an efficient strategy for producing ubiquitinated nucleosomes, advancing biochemical and biophysical studies in the field of chromatin biology.

AB - Background: Histone ubiquitination modification is emerging as a critical epigenetic mechanism involved in a range of biological processes. In vitro reconstitution of ubiquitinated nucleosomes is pivotal for elucidating the influence of histone ubiquitination on chromatin dynamics. Results: In this study, we introduce a Non-Denatured Histone Octamer Ubiquitylation (NDHOU) approach for generating ubiquitin or ubiquitin-like modified histone octamers. The method entails the co-expression and purification of histone octamers, followed by their chemical cross-linking to ubiquitin using 1,3-dibromoacetone. We demonstrate that nucleosomes reconstituted with these octamers display a high degree of homogeneity, rendering them highly compatible with in vitro biochemical assays. These ubiquitinated nucleosomes mimic physiological substrates in function and structure. Additionally, we have extended this method to cross-linking various histone octamers and three types of ubiquitin-like proteins. Conclusions: Overall, our findings offer an efficient strategy for producing ubiquitinated nucleosomes, advancing biochemical and biophysical studies in the field of chromatin biology.

UR - http://www.scopus.com/inward/record.url?scp=85196087820&partnerID=8YFLogxK

U2 - 10.1186/s13578-024-01265-x

DO - 10.1186/s13578-024-01265-x

M3 - Journal article

AN - SCOPUS:85196087820

SN - 2045-3701

VL - 14

JO - Cell and Bioscience

JF - Cell and Bioscience

IS - 1

M1 - 81

ER -

Rapid reconstitution of ubiquitinated nucleosome using a non-denatured histone octamer ubiquitylation approach

Abstract

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