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. 2023 Jun 26;120(27):e2300761120. doi: 10.1073/pnas.2300761120

Fig. 4.

Fig. 4.

Three polymerases are required for TCR mutagenesis. (A and B) Mutation rates of B. subtilis strains. N = 54 (wt), 40 (ΔpolA), 36 (ΔuvrA ΔpolA), 44 (Δmfd ΔpolA), 57 (ΔpolY1), 56 (ΔpolY2), 35 (ΔpolY1 ΔpolY2), 43 (ΔpolA ΔpolY1 ΔpolY2) biological replicates. Error bars are 95% CI. (C) Model for the molecular mechanism of NER-dependent mutagenesis. (D) Primer extension assay with purified B. subtilis PolA (middle lane) or PolA-ΔSID (right lane) using a gap substrate. Due to DNA being single stranded in the transcription bubble and/or during NER, the nontranscribed strand is prone to damage that stalls PolA and leads to the recruitment of Y-family polymerases, further increasing the possibility of acquiring a mutation. (E) Primer extension assay with purified B. subtilis PolA using a substrate that has either a thymidine (middle lane) or an abasic site (right lane) in its ninth nucleotide.