RNA targeting with CRISPR-Cas13a

By Omar Abudayyeh, Jonathan Gootenberg, Patrick Essletzbichler, Shuo Han, J. Keith Joung, Joseph J. Belanto, V. Verdine, David B. T. Cox, M. Kellner, Aviv Regev and 4 more. Nature, 2017

1,762 citations recorded by the source corpus.

RNA has important and diverse roles in biology, but molecular tools to manipulate and measure it are limited. For example, RNA interference can efficiently knockdown RNAs, but it is prone to off-target effects, and visualizing RNAs typically relies on the introduction of exogenous tags. Here we demonstrate that the class 2 type VI RNA-guided RNA-targeting CRISPR–Cas effector Cas13a (previously known as C2c2) can be engineered for mammalian cell RNA knockdown and binding. After initial screening of 15 orthologues, we identified Cas13a from Leptotrichia wadei (LwaCas13a) as the most effective in an interference assay in Escherichia coli. LwaCas13a can be heterologously expressed in mammalian and plant cells for targeted knockdown of either reporter or endogenous transcripts with comparable levels of knockdown as RNA interference and improved specificity. Catalytically inactive LwaCas13a maintains targeted RNA binding activity, which we leveraged for programmable tracking of transcripts in live cells. Our results establish CRISPR–Cas13a as a flexible platform for studying RNA in mammalian cells and therapeutic development.

Subfields: Molecular Biology

View Paper: https://www.semanticscholar.org/paper/0dd1750976aec4db08006e0b86add40caa263b02

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In Molecular Biology

1. Ling-Ling Chen
2. Li Yang
3. Arnab Rudra
4. Daniel G. Anderson
5. Lei Miao
6. Min Wu
7. Mengyuan Huang
8. Hongyi Li

Citations in the corpus

• Applications of genome editing technology in the targeted therapy of human diseases: mechanisms, advances and prospects (2020, methodology): “This approach appeared to be a highly sensitive detection method when used to detect two cancer mutants, BRAF V600E and EGFR L858R.57 Another system called DETECTR (DNA endonuclease-targeted CRISPR trans-reporter) has also been developed.261 Cas12a acts like Cas13a in this system, and another enzyme”
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• Delivering the Messenger: Advances in Technologies for Therapeutic mRNA Delivery. (2019, positive): “126 ,127 The barriers to broad clinical translation of CRISPR-Cas systems include (1) imperfect DNA-targeting speci fi city leading to off-target effects, 128 (2) low ef fi ciency of genome editing using HDR, 129 and (3) challenging delivery of CRISPR-Cas components.”
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