Evaluating Synthetic Activation and Repression of Neuropsychiatric-Related Genes in hiPSC-Derived NPCs, Neurons, and Astrocytes

Abstract

Summary: Modulation of transcription, either synthetic activation or repression, via dCas9-fusion proteins is a relatively new methodology with the potential to facilitate high-throughput up- or downregulation studies of gene function. Genetic studies of neurodevelopmental disorders have identified a growing list of risk variants, including both common single-nucleotide variants and rare copy-number variations, many of which are associated with genes having limited functional annotations. By applying a CRISPR-mediated gene-activation/repression platform to populations of human-induced pluripotent stem cell-derived neural progenitor cells, neurons, and astrocytes, we demonstrate that it is possible to manipulate endogenous expression levels of candidate neuropsychiatric risk genes across these three cell types. Although proof-of-concept studies using catalytically inactive Cas9-fusion proteins to modulate transcription have been reported, here we present a detailed survey of the reproducibility of gRNA positional effects across a variety of neurodevelopmental disorder-relevant risk genes, donors, neural cell types, and dCas9 effectors. : Brennand and colleagues report a survey of the reproducibility of CRISPR-mediated transcriptional modulation across varied neurodevelopmental disorder risk genes, donors, neural cell types, and dCas9 effectors. We report a number of practical limitations that must be considered when designing hiPSC-based studies using this promising new tool. Keywords: CRISPR, human-induced pluripotent stem cell, neural progenitor cell, transcriptional modulation, dCas9-VP64, dCas9-VPR, dCas9-KRAB