Computational Analysis Concerning the Impact of DNA Accessibility on CRISPR-Cas9 Cleavage Efficiency.
Base Sequence
/ genetics
CRISPR-Associated Protein 9
/ genetics
CRISPR-Cas Systems
/ genetics
Cell Line, Tumor
Chromatin
/ genetics
Clustered Regularly Interspaced Short Palindromic Repeats
/ genetics
Computational Biology
/ methods
DNA
/ genetics
Databases, Genetic
Deoxyribonuclease I
/ genetics
Gene Editing
/ methods
Genome, Human
HEK293 Cells
High-Throughput Nucleotide Sequencing
Humans
RNA, Guide, Kinetoplastida
/ genetics
RNA-Seq
Transcription, Genetic
Transcriptome
CIRCLE-seq
CRISPR
DNase-seq
GUIDE-seq
RNA-seq
bioinformatics
chromatin
Journal
Molecular therapy : the journal of the American Society of Gene Therapy
ISSN: 1525-0024
Titre abrégé: Mol Ther
Pays: United States
ID NLM: 100890581
Informations de publication
Date de publication:
08 01 2020
08 01 2020
Historique:
received:
23
03
2019
revised:
26
09
2019
accepted:
10
10
2019
pubmed:
2
11
2019
medline:
22
12
2020
entrez:
2
11
2019
Statut:
ppublish
Résumé
Defining the variables that impact the specificity of CRISPR/Cas9 has been a major research focus. Whereas sequence complementarity between guide RNA and target DNA substantially dictates cleavage efficiency, DNA accessibility of the targeted loci has also been hypothesized to be an important factor. In this study, functional data from two genome-wide assays, genome-wide, unbiased identification of DSBs enabled by sequencing (GUIDE-seq) and circularization for in vitro reporting of cleavage effects by sequencing (CIRCLE-seq), have been computationally analyzed in conjunction with DNA accessibility determined via DNase I-hypersensitive sequencing from the Encyclopedia of DNA Elements (ENCODE) Database and transcriptome from the Sequence Read Archive to determine whether cellular factors influence CRISPR-induced cleavage efficiency. CIRCLE-seq and GUIDE-seq datasets were selected to represent the absence and presence of cellular factors, respectively. Data analysis revealed that correlations between sequence similarity and CRISPR-induced cleavage frequency were altered by the presence of cellular factors that modulated the level of DNA accessibility. The above-mentioned correlation was abolished when cleavage sites were located in less accessible regions. Furthermore, CRISPR-mediated edits were permissive even at regions that were insufficient for most endogenous genes to be expressed. These results provide a strong basis to dissect the contribution of local chromatin modulation markers on CRISPR-induced cleavage efficiency.
Identifiants
pubmed: 31672284
pii: S1525-0016(19)30462-9
doi: 10.1016/j.ymthe.2019.10.008
pmc: PMC6953893
pii:
doi:
Substances chimiques
Chromatin
0
RNA, Guide
0
DNA
9007-49-2
CRISPR-Associated Protein 9
EC 3.1.-
Deoxyribonuclease I
EC 3.1.21.1
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
19-28Subventions
Organisme : NIMH NIH HHS
ID : P30 MH092177
Pays : United States
Organisme : NIMH NIH HHS
ID : R01 MH110360
Pays : United States
Organisme : NIMH NIH HHS
ID : T32 MH079785
Pays : United States
Informations de copyright
Copyright © 2019 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.
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