Repurposing the CRISPR-Cas9 system for targeted DNA methylation

By Aleksandar Vojta, P. Dobrinić, V. Tadić, L. Bočkor, P. Korać, B. Julg, Marija Klasić, V. Zoldoš. Nucleic Acids Research, 2016

720 citations recorded by the source corpus.

Epigenetic studies relied so far on correlations between epigenetic marks and gene expression pattern. Technologies developed for epigenome editing now enable direct study of functional relevance of precise epigenetic modifications and gene regulation. The reversible nature of epigenetic modifications, including DNA methylation, has been already exploited in cancer therapy for remodeling the aberrant epigenetic landscape. However, this was achieved non-selectively using epigenetic inhibitors. Epigenetic editing at specific loci represents a novel approach that might selectively and heritably alter gene expression. Here, we developed a CRISPR-Cas9-based tool for specific DNA methylation consisting of deactivated Cas9 (dCas9) nuclease and catalytic domain of the DNA methyltransferase DNMT3A targeted by co–expression of a guide RNA to any 20 bp DNA sequence followed by the NGG trinucleotide. We demonstrated targeted CpG methylation in a ∼35 bp wide region by the fusion protein. We also showed that multiple guide RNAs could target the dCas9-DNMT3A construct to multiple adjacent sites, which enabled methylation of a larger part of the promoter. DNA methylation activity was specific for the targeted region and heritable across mitotic divisions. Finally, we demonstrated that directed DNA methylation of a wider promoter region of the target loci IL6ST and BACH2 decreased their expression.

Subfields: Aging, Molecular Biology

View Paper: https://www.semanticscholar.org/paper/6ba714c69f5cb2f728907f20d6ce52fa5d6d3716

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In Molecular Biology

Citations in the corpus

Compact and highly active next-generation libraries for CRISPR-mediated gene repression and activation (2016, background): “…will greatly enhance the expanding set CRISPR-mediated technologies for controlling gene expression (Hilton et al., 2015; Perez-Pinera et al., 2013; Vojta et al., 2016), imaging targeted loci (Chen et al., 2013; Shao et al., 2016), or precisely editing the genome (Komor et al., 2016; Tsai et al.,…”
Editing DNA methylation in the mammalian genome (2016, positive): “also reported that dCas9 fused with Dnmt3a can be used to methylate two human gene promoters (Vojta et al., 2016), and Xu et al.”