Genome-wide programmable transcriptional memory by CRISPR-based epigenome editing.

A general approach for heritably altering gene expression has the potential to enable many discovery and therapeutic efforts. Here, we present CRISPRoff-a programmable epigenetic memory writer consisting of a single dead Cas9 fusion protein that establishes DNA methylation and repressive histone modifications. Transient CRISPRoff expression initiates highly specific DNA methylation and gene repression that is maintained through cell division and differentiation of stem cells to neurons. Pairing CRISPRoff with genome-wide screens and analysis of chromatin marks establishes rules for heritable gene silencing. We identify single guide RNAs (sgRNAs) capable of silencing the large majority of genes including those lacking canonical CpG islands (CGIs) and reveal a wide targeting window extending beyond annotated CGIs. The broad ability of CRISPRoff to initiate heritable gene silencing even outside of CGIs expands the canonical model of methylation-based silencing and enables diverse applications including genome-wide screens, multiplexed cell engineering, enhancer silencing, and mechanistic exploration of epigenetic inheritance.
Competing Interests: Declaration of interests J.K.N., J.C., G.C.P., L.A.G., and J.S.W. have filed patent applications related to CRISPRoff, CRISPRon, and CRISPRi/a screening. J.M.R. consults for Maze Therapeutics. L.A.G., J.S.W., H.Y.C., and B.E.B. consult for and hold equity in Chroma Medicine. J.S.W. declares outside interest in KSQ Therapeutics, Maze Therapeutics, Amgen, and Tessera Therapeutics. M.K. serves on the Scientific Advisory Boards of Engine Biosciences, Casma Therapeutics, and Cajal Neuroscience. B.E.B. declares outside interests in Fulcrum Therapeutics, Arsenal Biosciences, HiFiBio, and Cell Signaling Technologies. H.Y.C. is a co-founder of Accent Therapeutics, Boundless Bio, and is an advisor for 10x Genomics, Arsenal Biosciences, and Spring Discovery.
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