Open AccessEfficient in vitro and in vivo RNA editing via recruitment of endogenous ADARs using circular guide RNAs
Author(s) -
Dhruva Katrekar,
James Yen,
Yichen Xiang,
Anushka Saha,
Dario Meluzzi,
Yiannis A. Savva,
Prashant Mali
Publication year - 2022
Publication title -
nature biotechnology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 15.358
H-Index - 445
eISSN - 1546-1696
pISSN - 1087-0156
DOI - 10.1038/s41587-021-01171-4
Subject(s) - rna editing , rna , biology , competing endogenous rna , adar , guide rna , rna silencing , computational biology , nucleic acid secondary structure , inosine , non coding rna , microrna , rna binding protein , long non coding rna , genetics , microbiology and biotechnology , rna interference , genome editing , crispr , gene , adenosine , biochemistry
Recruiting endogenous adenosine deaminases using exogenous guide RNAs to edit cellular RNAs is a promising therapeutic strategy, but editing efficiency and durability remain low using current guide RNA designs. In this study, we engineered circular ADAR-recruiting guide RNAs (cadRNAs) to enable more efficient programmable adenosine-to-inosine RNA editing without requiring co-delivery of any exogenous proteins. Using these cadRNAs, we observed robust and durable RNA editing across multiple sites and cell lines, in both untranslated and coding regions of RNAs, and high transcriptome-wide specificity. Additionally, we increased transcript-level specificity for the target adenosine by incorporating interspersed loops in the antisense domains, reducing bystander editing. In vivo delivery of cadRNAs via adeno-associated viruses enabled 53% RNA editing of the mPCSK9 transcript in C57BL/6J mice livers and 12% UAG-to-UGG RNA correction of the amber nonsense mutation in the IDUA-W392X mouse model of mucopolysaccharidosis type I-Hurler syndrome. cadRNAs enable efficient programmable RNA editing in vivo with diverse protein modulation and gene therapeutic applications.