Nucleosome stability measured in situ by automated quantitative imaging

Abstract Current approaches have limitations in providing insight into the functional properties of particular nucleosomes in their native molecular environment. Here we describe a simple and powerful method involving elution of histones using intercalators or salt, to assess stability features depe...

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Autores principales:	László Imre, Zoltán Simándi, Attila Horváth, György Fenyőfalvi, Péter Nánási, Erfaneh Firouzi Niaki, Éva Hegedüs, Zsolt Bacsó, Urbain Weyemi, Rebekka Mauser, Juan Ausio, Albert Jeltsch, William Bonner, László Nagy, Hiroshi Kimura, Gábor Szabó
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2017
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/196732ab0b2f4c0c8ac935e97496a41e
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spelling	oai:doaj.org-article:196732ab0b2f4c0c8ac935e97496a41e2021-12-02T15:04:56ZNucleosome stability measured in situ by automated quantitative imaging10.1038/s41598-017-12608-92045-2322https://doaj.org/article/196732ab0b2f4c0c8ac935e97496a41e2017-10-01T00:00:00Zhttps://doi.org/10.1038/s41598-017-12608-9https://doaj.org/toc/2045-2322Abstract Current approaches have limitations in providing insight into the functional properties of particular nucleosomes in their native molecular environment. Here we describe a simple and powerful method involving elution of histones using intercalators or salt, to assess stability features dependent on DNA superhelicity and relying mainly on electrostatic interactions, respectively, and measurement of the fraction of histones remaining chromatin-bound in the individual nuclei using histone type- or posttranslational modification- (PTM-) specific antibodies and automated, quantitative imaging. The method has been validated in H3K4me3 ChIP-seq experiments, by the quantitative assessment of chromatin loop relaxation required for nucleosomal destabilization, and by comparative analyses of the intercalator and salt induced release from the nucleosomes of different histones. The accuracy of the assay allowed us to observe examples of strict association between nucleosome stability and PTMs across cell types, differentiation state and throughout the cell-cycle in close to native chromatin context, and resolve ambiguities regarding the destabilizing effect of H2A.X phosphorylation. The advantages of the in situ measuring scenario are demonstrated via the marked effect of DNA nicking on histone eviction that underscores the powerful potential of topological relaxation in the epigenetic regulation of DNA accessibility.László ImreZoltán SimándiAttila HorváthGyörgy FenyőfalviPéter NánásiErfaneh Firouzi NiakiÉva HegedüsZsolt BacsóUrbain WeyemiRebekka MauserJuan AusioAlbert JeltschWilliam BonnerLászló NagyHiroshi KimuraGábor SzabóNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 7, Iss 1, Pp 1-15 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q László Imre Zoltán Simándi Attila Horváth György Fenyőfalvi Péter Nánási Erfaneh Firouzi Niaki Éva Hegedüs Zsolt Bacsó Urbain Weyemi Rebekka Mauser Juan Ausio Albert Jeltsch William Bonner László Nagy Hiroshi Kimura Gábor Szabó Nucleosome stability measured in situ by automated quantitative imaging
description	Abstract Current approaches have limitations in providing insight into the functional properties of particular nucleosomes in their native molecular environment. Here we describe a simple and powerful method involving elution of histones using intercalators or salt, to assess stability features dependent on DNA superhelicity and relying mainly on electrostatic interactions, respectively, and measurement of the fraction of histones remaining chromatin-bound in the individual nuclei using histone type- or posttranslational modification- (PTM-) specific antibodies and automated, quantitative imaging. The method has been validated in H3K4me3 ChIP-seq experiments, by the quantitative assessment of chromatin loop relaxation required for nucleosomal destabilization, and by comparative analyses of the intercalator and salt induced release from the nucleosomes of different histones. The accuracy of the assay allowed us to observe examples of strict association between nucleosome stability and PTMs across cell types, differentiation state and throughout the cell-cycle in close to native chromatin context, and resolve ambiguities regarding the destabilizing effect of H2A.X phosphorylation. The advantages of the in situ measuring scenario are demonstrated via the marked effect of DNA nicking on histone eviction that underscores the powerful potential of topological relaxation in the epigenetic regulation of DNA accessibility.
format	article
author	László Imre Zoltán Simándi Attila Horváth György Fenyőfalvi Péter Nánási Erfaneh Firouzi Niaki Éva Hegedüs Zsolt Bacsó Urbain Weyemi Rebekka Mauser Juan Ausio Albert Jeltsch William Bonner László Nagy Hiroshi Kimura Gábor Szabó
author_facet	László Imre Zoltán Simándi Attila Horváth György Fenyőfalvi Péter Nánási Erfaneh Firouzi Niaki Éva Hegedüs Zsolt Bacsó Urbain Weyemi Rebekka Mauser Juan Ausio Albert Jeltsch William Bonner László Nagy Hiroshi Kimura Gábor Szabó
author_sort	László Imre
title	Nucleosome stability measured in situ by automated quantitative imaging
title_short	Nucleosome stability measured in situ by automated quantitative imaging
title_full	Nucleosome stability measured in situ by automated quantitative imaging
title_fullStr	Nucleosome stability measured in situ by automated quantitative imaging
title_full_unstemmed	Nucleosome stability measured in situ by automated quantitative imaging
title_sort	nucleosome stability measured in situ by automated quantitative imaging
publisher	Nature Portfolio
publishDate	2017
url	https://doaj.org/article/196732ab0b2f4c0c8ac935e97496a41e
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Nucleosome stability measured in situ by automated quantitative imaging

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