Analysis of protein-DNA interactions in chromatin by UV induced cross-linking and mass spectrometry.
Journal
Nature communications
ISSN: 2041-1723
Titre abrégé: Nat Commun
Pays: England
ID NLM: 101528555
Informations de publication
Date de publication:
16 10 2020
16 10 2020
Historique:
received:
06
09
2019
accepted:
24
09
2020
entrez:
17
10
2020
pubmed:
18
10
2020
medline:
18
11
2020
Statut:
epublish
Résumé
Protein-DNA interactions are key to the functionality and stability of the genome. Identification and mapping of protein-DNA interaction interfaces and sites is crucial for understanding DNA-dependent processes. Here, we present a workflow that allows mass spectrometric (MS) identification of proteins in direct contact with DNA in reconstituted and native chromatin after cross-linking by ultraviolet (UV) light. Our approach enables the determination of contact interfaces at amino-acid level. With the example of chromatin-associated protein SCML2 we show that our technique allows differentiation of nucleosome-binding interfaces in distinct states. By UV cross-linking of isolated nuclei we determined the cross-linking sites of several factors including chromatin-modifying enzymes, demonstrating that our workflow is not restricted to reconstituted materials. As our approach can distinguish between protein-RNA and DNA interactions in one single experiment, we project that it will be possible to obtain insights into chromatin and its regulation in the future.
Identifiants
pubmed: 33067435
doi: 10.1038/s41467-020-19047-7
pii: 10.1038/s41467-020-19047-7
pmc: PMC7567871
doi:
Substances chimiques
Chromatin
0
Nucleosomes
0
Polycomb-Group Proteins
0
Proteins
0
SCML2 protein, human
0
DNA
9007-49-2
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
5250Subventions
Organisme : Wellcome Trust
Pays : United Kingdom
Organisme : Medical Research Council
Pays : United Kingdom
Organisme : Wellcome Trust
ID : 109854/Z/15/Z
Pays : United Kingdom
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