Impact of chromatin context on Cas9-induced DNA double-strand break repair pathway balance.
Base Sequence
CRISPR-Associated Protein 9
/ metabolism
Chromatin
/ metabolism
DNA Breaks, Double-Stranded
DNA End-Joining Repair
DNA Repair
Euchromatin
/ metabolism
Gene Rearrangement
Genome, Human
Heterochromatin
/ metabolism
Humans
INDEL Mutation
/ genetics
K562 Cells
Kinetics
Protein Binding
Reproducibility of Results
CRISPR
Chromatin
DNA repair
MMEJ
NHEJ
SSTR
double strand break
heterochromatin
nuclear lamina
reporter assay
Journal
Molecular cell
ISSN: 1097-4164
Titre abrégé: Mol Cell
Pays: United States
ID NLM: 9802571
Informations de publication
Date de publication:
20 05 2021
20 05 2021
Historique:
received:
04
05
2020
revised:
23
12
2020
accepted:
19
03
2021
pubmed:
14
4
2021
medline:
17
6
2021
entrez:
13
4
2021
Statut:
ppublish
Résumé
DNA double-strand break (DSB) repair is mediated by multiple pathways. It is thought that the local chromatin context affects the pathway choice, but the underlying principles are poorly understood. Using a multiplexed reporter assay in combination with Cas9 cutting, we systematically measure the relative activities of three DSB repair pathways as a function of chromatin context in >1,000 genomic locations. This reveals that non-homologous end-joining (NHEJ) is broadly biased toward euchromatin, while the contribution of microhomology-mediated end-joining (MMEJ) is higher in specific heterochromatin contexts. In H3K27me3-marked heterochromatin, inhibition of the H3K27 methyltransferase EZH2 reverts the balance toward NHEJ. Single-stranded template repair (SSTR), often used for precise CRISPR editing, competes with MMEJ and is moderately linked to chromatin context. These results provide insight into the impact of chromatin on DSB repair pathway balance and guidance for the design of Cas9-mediated genome editing experiments.
Identifiants
pubmed: 33848455
pii: S1097-2765(21)00227-6
doi: 10.1016/j.molcel.2021.03.032
pmc: PMC8153251
pii:
doi:
Substances chimiques
Chromatin
0
Euchromatin
0
Heterochromatin
0
CRISPR-Associated Protein 9
EC 3.1.-
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
2216-2230.e10Subventions
Organisme : NIDDK NIH HHS
ID : U54 DK107965
Pays : United States
Commentaires et corrections
Type : CommentIn
Type : CommentIn
Informations de copyright
Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.
Déclaration de conflit d'intérêts
Declaration of interests B.v.S. is a member of the Advisory Board of Molecular Cell.
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