Precise epigenomic editing with a SunTag-based modular epigenetic toolkit

Abstract

Epigenetic regulation is a crucial factor controlling gene expression. Here, we report our CRISPR/dCas9-based modular epigenetic toolkit that enables gene-specific modulation of epigenetic architecture. By modifying the SunTag framework of dCas9 tagged with five GCN4 moieties, each epigenetic writer is bound to scFv and target-specific sgRNA, and this system is able to modify multiple epigenetic marks in a target-specific manner. We successfully demonstrated that this system is efficient in modifying individual histone post-translational modifications. We display its utility as a tool to understand the contributions of specific histone marks on gene expression by screening a large promoter region and identifying differential outcomes with high base-pair resolution. This epigenetic toolkit can be easily altered with a large variety of epigenetic effectors and is a useful tool for researchers to use in understanding gene-specific epigenetic changes and their relation to gene expression.

Microbial Synthetic Epigenetic Tools Design and Applications

“…First, there is a need to broaden the spectrum of epigenetic toolkits to construct more complex regulatory systems. This tools expansion could involve: (i) exploring novel transferases and characterizing their stability and sensitivity to make toolkits for the desired applications; (ii) exploring the potential of the microbial R-M system to make programmable and powerful cleavage-based epigenetic tools; (iii) incorporating synthetic epigenetic circuits with autofeedback designs to compensate system noise and phenotypic variability; (iv) developing digital converter circuits or small-molecule-responsive switches with intermolecular sequestration could allow the design of orthogonal epigenetic tools; and (v) advancing the performance of microbial synthetic epigenetic tools by introducing an enhanced affinity inhibitory-binding domain to timely separated homodimer or heterodimer proteins (Figure A).…”