Approaches to Study Native Chromatin-Modifying Complex Activities and Functions.
chromatin
histone modification
native protein complexes
nuclear protein depletion
recombinant nucleosomes
Journal
Frontiers in cell and developmental biology
ISSN: 2296-634X
Titre abrégé: Front Cell Dev Biol
Pays: Switzerland
ID NLM: 101630250
Informations de publication
Date de publication:
2021
2021
Historique:
received:
23
06
2021
accepted:
24
08
2021
entrez:
4
10
2021
pubmed:
5
10
2021
medline:
5
10
2021
Statut:
epublish
Résumé
The modification of histones-the structural components of chromatin-is a central topic in research efforts to understand the mechanisms regulating genome expression and stability. These modifications frequently occur through associations with multisubunit complexes, which contain active enzymes and additional components that orient their specificity and read the histone modifications that comprise epigenetic signatures. To understand the functions of these modifications it is critical to study the enzymes and substrates involved in their native contexts. Here, we describe experimental approaches to purify native chromatin modifiers complexes from mammalian cells and to produce recombinant nucleosomes that are used as substrates to determine the activity of the complex. In addition, we present a novel approach, similar to the yeast anchor-away system, to study the functions of essential chromatin modifiers by quickly inducing their depletion from the nucleus. The step-by-step protocols included will help standardize these approaches in the research community, enabling convincing conclusions about the specificities and functions of these crucial regulators of the eukaryotic genome.
Identifiants
pubmed: 34604228
doi: 10.3389/fcell.2021.729338
pmc: PMC8481805
doi:
Types de publication
Journal Article
Langues
eng
Pagination
729338Informations de copyright
Copyright © 2021 Galloy, Lachance, Cheng, Distéfano-Gagné, Côté and Fradet-Turcotte.
Déclaration de conflit d'intérêts
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
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