RNA Editing with CRISPR-Cas13

By David B. T. Cox, Jonathan Gootenberg, Omar Abudayyeh, Brian Franklin, M. Kellner, J. Keith Joung, Feng Zhang. Science, 2017

1,587 citations recorded by the source corpus.

Precise transcriptome engineering Efficient and precise RNA editing to correct disease-relevant transcripts holds great promise for treating genetic disease. Cox et al. took advantage of the ability of Cas13b, an effector from a type VI CRISPR-Cas system, to target specific RNAs directly (see the Perspective by Yang and Chen). They fused Cas13b with the ADAR2 adenosine deaminase domain and used rational protein engineering to improve the resultant enzyme. These approaches yielded an RNA knockdown and editing platform that allowed efficient and specific RNA depletion and correction in mammalian cells. Science, this issue p. 1019; see also p. 996 An RNA-editing technology is developed to efficiently and specifically deplete and correct RNA directly in mammalian cells. Nucleic acid editing holds promise for treating genetic disease, particularly at the RNA level, where disease-relevant sequences can be rescued to yield functional protein products. Type VI CRISPR-Cas systems contain the programmable single-effector RNA-guided ribonuclease Cas13. We profiled type VI systems in order to engineer a Cas13 ortholog capable of robust knockdown and demonstrated RNA editing by using catalytically inactive Cas13 (dCas13) to direct adenosine-to-inosine deaminase activity by ADAR2 (adenosine deaminase acting on RNA type 2) to transcripts in mammalian cells. This system, referred to as RNA Editing for Programmable A to I Replacement (REPAIR), which has no strict sequence constraints, can be used to edit full-length transcripts containing pathogenic mutations. We further engineered this system to create a high-specificity variant and minimized the system to facilitate viral delivery. REPAIR presents a promising RNA-editing platform with broad applicability for research, therapeutics, and biotechnology.

Subfields: Cardiology and Cardiovascular Medicine, Molecular Biology

View Paper: https://www.semanticscholar.org/paper/4bbcf976eb0155f881fe1e09c4980c8985795c03

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Citations in the corpus

Multiplexed and portable nucleic acid detection platform with Cas13, Cas12a, and Csm6 (2018, neutral): “(C) Sequences of target and REPAIR guide design used for correction of APC alleles.”
Transcriptome engineering with RNA-targeting Type VI-D CRISPR effectors (2018, neutral): “G Laboratory of Didier Trono Addgene #12259 EF1a-PspCas13b-NES(HIV)-2A-EGFP This paper Adapted from Addgene #103862 from Feng Zhang pC0043-PspCas13b crRNA backbone Cox et al., 2017 Addgene #103854 pC016-LwCas13a guide expression backbone with U6 promoter Abudayyeh et al., 2017 Addgene #91906 EF1a-Lw”
Deciphering miRNAs’ Action through miRNA Editing (2019, methodology): “More recently, the CRISPR-Cas13b RNA editing for programmable A-to-I replacement (REPAIR) system has been established by fusing a Cas13b protein and wild-type or mutated ADAR1/2 DD, and it acts similarly to the CRISPR-Cas9 system [93].”
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.”
CRISPR/Cas Genome Editing and Precision Plant Breeding in Agriculture. (2019, background): “However, class 2 type VI systems, which are characterized by the presence of the single effector protein C2c2 (now known as Cas13), such as LwaCas13a (1) and PspCas13b (22), have been engineered to target RNA or to perform RNA editing (Figure 2d ).”