Precise in vivo RNA base editing with a wobble-enhanced circular CLUSTER guide RNA.

Autor: Reautschnig P; Interfaculty Institute of Biochemistry, Faculty of Science, University of Tübingen, Tübingen, Germany., Fruhner C; Interfaculty Institute of Biochemistry, Faculty of Science, University of Tübingen, Tübingen, Germany., Wahn N; Interfaculty Institute of Biochemistry, Faculty of Science, University of Tübingen, Tübingen, Germany., Wiegand CP; Interfaculty Institute of Biochemistry, Faculty of Science, University of Tübingen, Tübingen, Germany., Kragness S; Vollum Institute, Oregon Health and Science University, Portland, OR, USA., Yung JF; Vollum Institute, Oregon Health and Science University, Portland, OR, USA., Hofacker DT; Interfaculty Institute of Biochemistry, Faculty of Science, University of Tübingen, Tübingen, Germany., Fisk J; Vollum Institute, Oregon Health and Science University, Portland, OR, USA., Eidelman M; Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat Gan, Israel.; The Institute of Nanotechnology and Advanced Materials, Bar-Ilan University, Ramat Gan, Israel., Waffenschmidt N; Interfaculty Institute of Biochemistry, Faculty of Science, University of Tübingen, Tübingen, Germany., Feige M; Interfaculty Institute of Biochemistry, Faculty of Science, University of Tübingen, Tübingen, Germany., Pfeiffer LS; Interfaculty Institute of Biochemistry, Faculty of Science, University of Tübingen, Tübingen, Germany., Schulz AE; Interfaculty Institute of Biochemistry, Faculty of Science, University of Tübingen, Tübingen, Germany., Füll Y; Interfaculty Institute of Biochemistry, Faculty of Science, University of Tübingen, Tübingen, Germany., Levanon EY; Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat Gan, Israel.; The Institute of Nanotechnology and Advanced Materials, Bar-Ilan University, Ramat Gan, Israel., Mandel G; Vollum Institute, Oregon Health and Science University, Portland, OR, USA., Stafforst T; Interfaculty Institute of Biochemistry, Faculty of Science, University of Tübingen, Tübingen, Germany. thorsten.stafforst@uni-tuebingen.de.; Gene and RNA Therapy Center (GRTC), Faculty of Medicine, University of Tübingen, Tübingen, Germany. thorsten.stafforst@uni-tuebingen.de.; iFIT Cluster of Excellence (EXC2180) Image-Guided and Functionally Instructed Tumor Therapies, University of Tübingen, Tübingen, Germany. thorsten.stafforst@uni-tuebingen.de.
Jazyk: angličtina
Zdroj: Nature biotechnology [Nat Biotechnol] 2024 Jul 12. Date of Electronic Publication: 2024 Jul 12.
DOI: 10.1038/s41587-024-02313-0
Abstrakt: Recruiting the endogenous editing enzyme adenosine deaminase acting on RNA (ADAR) with tailored guide RNAs for adenosine-to-inosine (A-to-I) RNA base editing is promising for safely manipulating genetic information at the RNA level. However, the precision and efficiency of editing are often compromised by bystander off-target editing. Here, we find that in 5'-UAN triplets, which dominate bystander editing, G•U wobble base pairs effectively mitigate off-target events while maintaining high on-target efficiency. This strategy is universally applicable to existing A-to-I RNA base-editing systems and complements other suppression methods such as G•A mismatches and uridine (U) depletion. Combining wobble base pairing with a circularized format of the CLUSTER approach achieves highly precise and efficient editing (up to 87%) of a disease-relevant mutation in the Mecp2 transcript in cell culture. Virus-mediated delivery of the guide RNA alone realizes functional MeCP2 protein restoration in the central nervous system of a murine Rett syndrome model with editing yields of up to 19% and excellent bystander control in vivo.
(© 2024. The Author(s).)
Databáze: MEDLINE
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