Highly Specific Epigenome Editing by CRISPR/Cas9 Repressors for Silencing of Distal Regulatory Elements

By Pratiksha I. Thakore, A. D'Ippolito, Lingyun Song, Alexias Safi, Nishkala Shivakumar, A. Kabadi, Timothy E. Reddy, Gregory E. Crawford, C. Gersbach. Nature Methods, 2015

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Epigenome editing with the CRISPR (clustered, regularly interspaced, short palindromic repeats)-Cas9 platform is a promising technology for modulating gene expression to direct cell phenotype and to dissect the causal epigenetic mechanisms of gene regulation. Fusions of nuclease-inactive dCas9 to the Krüppel-associated box (KRAB) repressor (dCas9-KRAB) can silence target gene expression, but the genome-wide specificity and the extent of heterochromatin formation catalyzed by dCas9-KRAB are not known. We targeted dCas9-KRAB to the HS2 enhancer, a distal regulatory element that orchestrates the expression of multiple globin genes, and observed highly specific induction of H3K9 trimethylation (H3K9me3) at the enhancer and decreased chromatin accessibility of both the enhancer and its promoter targets. Targeted epigenetic modification of HS2 silenced the expression of multiple globin genes, with minimal off-target changes in global gene expression. These results demonstrate that repression mediated by dCas9-KRAB is sufficiently specific to disrupt the activity of individual enhancers via local modification of the epigenome.

Subfields: Molecular Biology, Plant Science

View Paper: https://www.semanticscholar.org/paper/887bac75946344aba8e3ef1d1da31a4b1860e13f

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Citations in the corpus

CRISPRi-based genome-scale identification of functional long non-coding RNA loci in human cells (2016, neutral): “H3K9me3 is a chromatin modification that is a result of local dCas9-KRAB activity (31), and in both U87 andHeLa cells with LINC00263CRISPRi targeting, ChIP-seq analysis demonstrated equal enrichment of H3K9me3 specifically at the LINC00263 promoter for two independent sgRNAs (Fig.”
CRISPR/Cas9 in Genome Editing and Beyond. (2016, methodology): “The studies (38, 39, 203) described above have demonstrated that dCas9 fusion proteins can act as sequence-specific, synthetic epigenome modifiers, which not only change local epigenetic status but also change the gene expression of relevant genes.”
Activity-by-Contact model of enhancer-promoter regulation from thousands of CRISPR perturbations (2019, methodology): “(KRAB-dCas9 has previously been shown to repress many promoters and enhancers, and affects elements within ~200–500 bp of the gRNA; Supplementary Note 1) 7–9 .”
Non-viral delivery systems for CRISPR/Cas9-based genome editing: challenges and opportunities (2018, background): “dCas9-KRAB system, recruiting a diverse array of histone modifiers that reversibly suppresses gene expression through the formation of heterochromatin, significantly reduced the expression of a series of endogenous genes in the genome [125, 128].”