dCas9: A Versatile Tool for Epigenome Editing

Abstract

The epigenome is a heritable layer of information not encoded in the DNA sequence of the genome, but in chemical modifications of DNA or histones. These chemical modifications, together with transcription factors, operate as spatiotemporal regulators of genome activity. Dissecting epigenome function requires controlled site-specific alteration of epigenetic information. Such control can be obtained using designed DNA-binding platforms associated with effector domains to function as targeted transcription factors or epigenetic modifiers. Here, we review the use of dCas9 as a novel and versatile tool for fundamental studies on epigenetic landscapes, chromatin structure and transcription regulation, and the potential of this approach in basic research in these fields.

Regulation of transposable elements by DNA modifications

“…These considerations seem particularly relevant for the more recently described 6mAmodifying enzymes, whose effects on TEs may be unrelated to their ability to modulate 6mA levels. The advent of epigenetic editing tools offers the opportunity to tackle these questions by altering the levels of DNA modifications at specific loci while also controlling for catalytic effects 161 .…”

Advances in epigenetics link genetics to the environment and disease

“…It is possible to mutate the genome precisely, reversibly and at multiple sites simultaneously 168 . Furthermore, one can tether proteins of interest to selected genomic positions in order to silence or activate genes, to induce reversible changes in 3D chromatin architecture and to visualize loci of interest in live imaging experiments 169,170 . This surge in approaches and technologies is revolutionizing the ways in which epigenetic processes can be studied, understood and harnessed to understand biological and pathological processes and to develop novel therapeutic strategies.…”

Unexpected binding behaviors of bacterial Argonautes in human cells cast doubts on their use as targetable gene regulators

“…While CRISPR/Cas9 is able to target near many genetic features, the necessity of a protospacer adjacent motif (PAM) in the target site often makes it impossible to design guide RNAs (gRNAs) to bind exactly at features such as SNPs, individual CpGs, intron/exon boundaries, or specific transcription factor binding sites ( Fig 1A ). Indeed this limitation has led to many efforts to expand the targeting lexicon using engineered or natural variant Cas9 proteins [ 7 – 10 ]. However, nearly all have been similarly or even more restricted than the 5’-NGG-3’ PAM site of the canonical Sp Cas9 [ 11 ].…”