Dataset Information


CRISPRδ: dCas13-mediated translational repression for accurate gene silencing in mammalian cells

ABSTRACT: Current gene silencing tools based on RNA interference (RNAi) or more recently the CRISPR-Cas13 systems have critical drawbacks in such as off-target effects (RNAi) or collateral mRNA cleavage (CRISPR-Cas13). Thus, a more specific method of gene knockdown has been demanded. Here we developed “CRISPRδ”, an approach for translational silencing, harnessing the catalytically inactive Cas13 proteins (dCas13). Owing to the tight association with mRNA, dCas13 serves as a physical roadblock for scanning ribosomes in translation initiation, but not for elongating ribosomes, with no effect on mRNA stability. Guide RNAs covering the start codon lead to the highest efficacy, irrespective of translation initiation mechanisms: cap-dependent, IRES-dependent, or RAN translations. Strikingly, genome-wide ribosome profiling revealed extremely high specificity of CRISPRδ in a gene knockdown. Moreover, the fusion of a translation repressor to dCas13 ensured further improvement of the knockdown efficacy. Our method provides a framework for translational repression-based gene silencing in eukaryotes.

ORGANISM(S): Homo sapiens

PROVIDER: GSE232383 | GEO | 2024/02/10


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