Fig. 7
Increased levels of translesion synthesis DNA Polymerase zeta in melanoma cells improves their response to cisplatin. a and b Increased levels of DNA Polymerase zeta, but not DNA Polymerase eta after cisplatin treatment. Melanoma cell lines (A375, C32, G361), ovarian cancer cell line, PEO4, and immortalised fibroblast line, MRC5v1, were untreated, or treated with 3 μM cisplatin for 24, 48 or 72 h and western blotted for translesion synthesis DNA Polymerases. a DNA Pol ζ. b DNA Pol η. Graphs shows the expression relative to β-actin and normalised to the level in untreated cells. Results are the means from two separate determinations. Representative western blots themselves are shown in Additional file 1: Figure S6. c Reduced growth in cisplatin-treated cultures following siRNA knockdown of DNA Polymerase zeta. The table shows the mean cell growth (±SEM, n = 4) of wells after 72 h of 1 μM cisplatin treatment for non-transfected A375 cells and cells transfected with DNA Polymerase zeta siRNA or scrambled control siRNA, expressed as a percentage of the untreated control. The P value for Student’s paired t test, comparing the effect of DNA Polymerase zeta siRNA or scrambled control siRNA transfection on cell growth of cisplatin-treated wells is also shown. d Increased apoptosis in cisplatin-treated DNA Pol ζ siRNA-transfected A375 cells. Cultures treated as in (c) after 72 h of 1 μM cisplatin. A representative flow cytometry profile for each culture condition is shown, with the percentage of cells with subG1 (apoptotic), G1, S and G2/M DNA contents indicated across the top of each profile. Note the increased level of subG1 (apoptotic) material in the cisplatin-treated DNA Pol ζ siRNA-transfected cells. The table below shows the mean level of apoptosis (±SEM, n = 4) in wells after 72 h of cisplatin treatment. The P values for Student’s paired t test, comparing the level of apoptosis between cisplatin-treated and untreated control wells, and between cisplatin-treated DNA Pol ζ siRNA-transfected wells and cisplatin-treated scrambled control siRNA-transfected wells are also shown