Minimizing far-extending chromatin perturbation in genome editing preserves stem cell identity
Advancing CRISPR/Cas9 genome editing demands a thorough understanding of its associated risks.Yet, most of risk assessments have focused on the mutagenic effects stemming from off-target cleavage in gene-coding regions, largely neglecting the implications of cleavage within the extensive non-coding genome. We observed that CRISPR/Cas9-induced DNA cleavage, even at sites several kilobases away from the nearest regulatory elements, can trigger premature differentiation of somatic neural stem cell in vivo.Leveraging a refined multi-omics technology, we delved into the consequences of CRISPR/Cas9-induced DNA cleavage in these non-coding regions. Our findings reveal an impact, especially pronounced in cells with high stemness, where cleavage-induced disruptions can span long ranges, potentially culminating in the loss of stemness. These effects are tied to extensive chromatin perturbations. Building on this understanding, we used chromatin perturbation as a key marker to refine gene editing technologies that could minimize such disruptions.This key finding illuminates the path towards the development of safer and more broadly applicable genome editing technologies. Overall design: See individual sub-series.