Fig. 1. Mitotic arrest deficient 2 like 2 (Mad2L2, also known as Mad2B) levels increase in HeLa cell after cisplatin-induced DNA damage.
Activation of the proliferating cell nuclear antigen (PCNA) and phosphorylation of H2A.X following cisplatin treatment (A, B). HeLa cells were treated with cisplatin (100 μM) for 12 h. PCNA (A) and γH2A.X (B) were stained with anti-PCNA and anti-pH2AX (S139) (red) antibodies, respectively. DNA was stained with Hoechst 33342 (blue). Cells were observed using a Nikon inverted fluorescence microscope (TE300). Approximately 400 cells were counted. White bar, control; Black bar, cisplatin. Scale bar, 10 μm. (C) Mad2B binding to chromatin was increased following cisplatin treatment. Exponentially growing HeLa cells were treated with cisplatin (50 μM) for 12 h and the cytoplasmic and chromatin-associated protein fractions were immunoblotted with antibodies against Mad2B, histone H3, and α-tubulin. (D) Cisplatin (50 μM) treatment increased the expression of Mad2B as determined via Western blotting of whole HeLa cell extracts (n = 3). Quantitation of the data shown in lower panel. Each bar represents the mean ± SD of three independent experiments (*p < 0.05, **p < 0.01 and ***p < 0.001 using a two-tailed t-test).
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