CRISPR-Mediated Epigenome Editing

Abstract

Mounting evidence has called into question our understanding of the role that the central dogma of molecular biology plays in human pathology. The conventional view that elucidating the mechanisms for translating genes into proteins can account for a panoply of diseases has proven incomplete. Landmark studies point to epigenetics as a missing piece of the puzzle. However, technological limitations have hindered the study of specific roles for histone post-translational modifications, DNA modifications, and non-coding RNAs in regulation of the epigenome and chromatin structure. This feature highlights CRISPR systems, including CRISPR-Cas9, as novel tools for targeted epigenome editing. It summarizes recent developments in the field, including integration of optogenetic and functional genomic approaches to explore new therapeutic opportunities, and underscores the importance of mitigating current limitations in the field. This comprehensive, analytical assessment identifies current research gaps, forecasts future research opportunities, and argues that as epigenome editing technologies mature, overcoming critical challenges in delivery, specificity, and fidelity should clear the path to bring these technologies into the clinic.

Keywords: CRISPR; CRISPR systems for epigenome editing; CRISPR-Cas9; Epigenome editing; epigenetics; epigenome engineering; histone and DNA epigenetic modifications; optogenetics; transcription activation; transcription repression.

Figure 1

Structural representation of the nuclease-null Cas9 (dCas9) from S. pyogenes. A. Crystal structure of Cas9 in complex with a single guide RNA (red) and its target DNA (orange). The HNH (cyan) and RuvC (purple) catalytic domains are shown. The non-catalytic regions of Cas9 are colored in blue. Mutation of the catalytic residues (D10A and H840A) that render Cas9 inactive (dCas9) are colored in yellow. B. Close-up view of the active site and position of the catalytic residues shown in A. C. Schematic representation of nuclease-null Cas9 in complex with sgRNA and target DNA; colors as shown in A. [PDB 4OO8].

Figure 2

Schematic representation of epigenome editing using dCas9 fused to an epigenetic effector. A. dCas9 (colors are the same as in Figure 1) binds to target DNA without triggering double-stranded DNA cleavage. An epigenetic effector—p300 histone acetyltransferase (HAT) in this drawing (brown)—is fused to dCas9 by a linker region (gray). Once positioned near the target epigenetic loci, the effector catalyzes covalent modification of the target histone tail on nearby nucleosomes (the reaction is represented by the black arrow). B. Cartoon illustration of A. Covalent modification illustrated by a blue star on the substrate histone tail. [PDB 4OO8, 3BIY, 1KX5].

Figure 3

Repurposing CRISPR- and non-CRISPR-based tools for epigenome editing. A. Structure of a TALE bound to its target DNA. B. Zinc Finger proteins in complex with DNA. C. Structure of Cpf1 in complex with crRNA and target DNA. D. Cas9 bound to sgRNA and PAM-containing target DNA. E. Structure of an Argonaute protein in complex with guide and target DNA fragments. F. E. coli Cascade bound to crRNA and PAM-containing dsDNA. All structures feature DNA in red and RNA (where applicable) in teal. [PDB 3UGM, 3DFX, 5B43, 4UN3, 4NCB, 5H9F].