The double life of CRISPR–Cas13

• Published in: Current opinion in biotechnology Vol. 78; p. 102789
• Main Authors: Bot, Jorik F, van der Oost, John, Geijsen, Niels
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01-12-2022
• Subjects: Animals; CRISPR-Cas Systems - genetics; Gene Editing - methods; Mammals - genetics; RNA - genetics
• ISSN: 0958-1669; 1879-0429
• DOI: 10.1016/j.copbio.2022.102789

Since the discovery of RNA-programmable nucleases from the prokaryotic adaptive immune system CRISPR–Cas, these proteins have seen rapid and widespread adoption for biotechnological and clinical research. A recently discovered system, CRISPR–Cas13, uses CRISPR RNA guides to target RNA. Interestingly, RNA targeting by Cas13 results in cleavage of both target RNA and bystander RNA. This feature has been used to develop innovative diagnostic tools for the detection of specific RNAs. Unlike in vitro detection of RNA using collateral RNA cleavage, however, initial studies of mammalian cells only revealed highly specific target RNA-knockdown activity. Although these findings have been confirmed subsequently, several recent publications do report Cas13-mediated toxicity and collateral RNA cleavage when using Cas13 in eukaryotes. Here, we review these conflicting observations and discuss its potential molecular basis.
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•CRISPR–Cas13 is potentially capable of collateral RNA cleavage in eukaryotic cells.•Target RNA levels, cell size, and cell-type-specific properties may influence the occurrence of collateral cleavage.•Commonly used methodologies are inadequate for assessing collateral RNA cleavage.•Collateral cleavage activity might limit the use of Cas13 for transcriptome-wide screens.•Clinical applications of Cas13 might be based on its collateral cleavage potential.