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bioRxiv · 10.1101/2023.02.17.528953

Precise and efficient C-to-U RNA Base Editing with SNAP-CDAR-S

Latifi, N.·Mack, A. M.·Tellioglu, I.·Di Giorgio, S.·Stafforst, T.
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Abstract

Site-directed RNA base editing enables the transient and dosable change of genetic information and represents a recent strategy to manipulate cellular processes, paving ways to novel therapeutic modalities. While tools to introduce adenosine-to-inosine changes have been explored quite intensively, the engineering of precise and programmable tools for cytidine-to-uridine editing is somewhat lacking behind. Here we demonstrate that the cytidine deaminase domain evolved from the ADAR2 adenosine deaminase, taken from the RESCUE-S tool, provides very efficient and highly programmable editing when changing the RNA targeting mechanism from Cas13-based to SNAP-tag-based. Optimization of the guide RNA chemistry further allowed to dramatically improve editing yields in the difficult-to-edit 5-CCN sequence context thus improving the substrate scope of the tool. Regarding editing efficiency, SNAP-CDAR-S outcompeted the RESCUE-S tool clearly on all tested targets, and was highly superior in perturbing the {beta}-catenin pathway. NGS analysis showed similar, moderate global off-target A-to-I and C-to-U editing for both tools.

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Latifi, N., Mack, A. M., Tellioglu, I., Di Giorgio, S., Stafforst, T.. 2023-02-19. Precise and efficient C-to-U RNA Base Editing with SNAP-CDAR-S. https://doi.org/10.1101/2023.02.17.528953

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