a, Cell viability assay comparing HEK293T cells transfected with plasmid expressing nCas9–UGI–NLS, wild-type (WT) BE3, BE3-R33A, BE3-R33A/K34A or BE3-E63Q (n = 3 biologically independent samples per condition). Each dot represents one biological replicate (and is the mean of three technical replicates). All data points were normalized to the mean luminescence of the nCas9–UGI–NLS controls (set to 100%, grey dotted line) that were performed for each biological replicate experiment. The assay was performed on days 1, 2, 3 and 4 after plating (following sorting for all GFP-positive cells). Data shown as mean ± s.e.m. RLU, relative light unit; n.s., not significantly decreased compared to matched nCas9–UGI–NLS control; *P < 0.05, ***P < 0.001 for a significant decrease compared to matched nCas9–UGI–NLS control. Statistical significance was determined as described in Supplementary Methods. b, Structural model of rAPOBEC1 with locations of catalytic residues and the R33 and K34 positions that were altered in SECURE variants. A predicted rAPOBEC1 structure is shown that was generated with Protein Homology/analogY Recognition Engine v 2.0 (Phyre2)37 and visualized in PyMOL (v 1.8.2.1). The R33 and K34 residues mutated in the SECURE variants are shown in orange and blue, respectively. Catalytic site residues (H61, E63, C93 and C96) have previously been described19 and are shown in green. c–f, Extended sequence logos for BE3- and ABEmax-induced RNA editing sites. Sequence logos derived with the nucleotides 100 base pairs upstream and downstream of the motifs edited in RNA by BE3 (ACW) or ABEmax (UA) are shown. Logos were derived from data for BE3 expression in HepG2 cells (c; Fig. 1c), BE3 expression in HEK293T cells (d; all GFP-sorted cells; Extended Data Fig. 2c), higher BE3 expression in HEK293T cells (e; top-5% GFP-sorted cells; Extended Data Fig. 4c), and ABEmax expression in HEK293T experiments (f; top 5% GFP-sorted cells; Fig. 3b). Analysis was done using RNA-seq data generated from cDNA; every T depicted should be considered a U in RNA.