GSE160945
Harnessing Self-Labeling Enzymes for Selective and Concurrent A-to-I and C-to-U RNA Base Editing
The SNAP-ADAR tool enables precise and efficient A-to-I RNA editing in a guideRNA-dependent  manner by applying the self-labeling SNAP-tag enzyme to generate RNA-guided  editases in cell culture. Here, we extend this platform by combining the SNAP-tagged tool with  further effectors steered by the orthogonal HALO-tag. Due to their small size (ca. 2 kb), both  effectors are readily integrated into one genomic locus. We demonstrate selective and  concurrent recruitment of ADAR1 and ADAR2 deaminase activity for optimal editing with  extended substrate scope and moderate global off-target effects. Furthermore, we combine the  recruitment of ADAR1 and APOBEC1 deaminase activity to achieve selective and concurrent  A-to-I and C-to-U RNA base editing of endogenous transcripts inside living cells again with moderate global off-target effects. The platform  should be readily transferable to further epitranscriptomic writers and erasers to manipulate  epitranscriptomic marks in a programmable way with high molecular precision. Overall design: Identification of off-target editing events in a 293 Flp-In T-REx cell line stably expressing both HALO-ADAR1Q and mAPOBEC1-SNAP by RNA-Seq. Overall 4 samples × 2 independent experiments.