Figure 2.

Msh2/Msh6 acts independently of NER to suppress the mutagenicity of photolesions. (A) Frequencies of mutants at the genomic Hprt gene in isogenic WT, Msh6^−/− (M), Xpa^−/− (X), and Msh6^−/−Xpa^−/− (MX, line 4) ES cells after mock or low-dose UVC (0.75 J/m²), treatment. Bars represent averages of at least three independent experiments. See Fig. S1 for results using an independently derived Msh6^−/−Xpa^−/− ES cell line (line 30). (B) UVC-induced mutant frequencies in isogenic ES cell lines, derived from data in A. (C) Msh2/Msh6-dependent suppression of UVC-induced mutagenesis in NER-proficient (WT) and in NER-deficient (X) backgrounds. Frequencies were derived by subtraction of induced mutant frequencies in Msh6-proficient cells from those in Msh6-deficient cells (B) in each individual experiment, followed by averaging. (D) Relative frequencies of UVC (0.75 J/m²)-induced nucleotide substitutions at dipyrimidines in Xpa^−/− (X) and Msh6^−/−Xpa^−/− (MX) ES cells, derived from Table S1. TS, dipyrimidine (photolesion site) located at the transcribed DNA strand; NTS, dipyrimidine at the nontranscribed DNA strand. (E) Slot blot illustrating photoreversal of the majority of CPD in UVC-treated (0.75 J/m²) ES cell lines. (F) UVC (0.75 J/m²)-induced mutant frequencies in Xpa^−/− (X) and in Msh6^−/−Xpa^−/− (MX) ES cells, with or without CPD photoreversal. All frequencies were corrected for spontaneous mutant frequencies. Bars represent averages of three independent experiments. (G) Effect of CPD photoreversal on UVC-induced mutant frequencies in Xpa^−/− (X) and in Msh6^−/−Xpa^−/− (MX) cells. Frequencies were calculated by subtraction of induced mutant frequencies in Msh6-proficient cells from those in Msh6-deficient cells (F) in each individual experiment, followed by averaging.