Fig. 2: Mutational spectrum is tuned by engineered targeting of repair factors.
From: Polymerase-guided base editing enables in vivo mutagenesis and rapid protein engineering
a Schematic of DNA-repair factor targeting using SH3 domains (purple) fused to integrated PmCDA1-T7 RNAP and SHL domains (pink) fused to host DNA-repair proteins. Localization of repair proteins initiates different mutational outcomes compared to PmCDA1-T7 RNAP alone. b Mutation frequencies for G > A substitutions plotted by position along URA3 for YHM2.MSH6 (blue), YHM2 (gray), and YHM2.PT7KO (yellow) when induced with 100 nM β-estradiol. In order to match YHM2 and YHM2.PT7KO, data from YHM2.PT7KO is off-set to account for the gap created by the 23 bp insertion which disrupts PT7. Mutation frequencies for all other base substitutions are shown in Supplementary Fig. 8b. Mutation data generated by next-generation sequencing at the URA3 locus as described in “Methods”. c A/T targeted mutation rate in strain YHM2.TAA having Apn2p and Msh6p DNA-repair proteins tagged with an SHL peptide or short random peptide. A/T rate is measured by reversion of a TAA stop codon in URA3 and quantified using the Falcor algorithm. Error bars represent 95% confidence intervals measured from 24 independent cultures induced with 100 nM β-estradiol. Source data are available as a Source data file.
