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. 2016 Oct 15;27(20):3065–3084. doi: 10.1091/mbc.E16-05-0268

FIGURE 5:

FIGURE 5:

p53-dependent transcriptional and cell cycle effects in proliferating tetraploid cells. (A) Real-time qPCR analysis of p21 and p53 genes in parental RPE1 cells and proliferating tetraploid and isogenic diploid single-cell clones. p21 mRNA levels were increased threefold to fourfold in tetraploid cell lines, whereas p53 mRNA levels were unchanged. All values were normalized to the parental RPE1. Bar heights represent average relative fold change (n = 3); error bars represent SEM. (B) Effect of p53 knockdown on proliferation of tetraploid single-cell clones. Cells were treated with siRNA for 72 h and allowed to incorporate EdU for 8 h. Percentages of EdU-positive cells were normalized to parental cell line not transfected with p53 siRNA (the number of EdU-positive cells in the parental untransfected sample was assigned 100% value). Bar heights represent the mean (n = 4); error bars represent SD. (C) Flow cytometric profiles of diploid and tetraploid isogenic cell lines treated with ZM447439 for 72 h. Like the diploid cell lines, tetraploid cells showed doubled or quadrupled DNA content. Cells were fixed and stained with propidium iodide. (D) Western blot analysis of p53 and p21 protein levels in proliferating tetraploid and isogenic diploid single-cell clones treated with Aurora inhibitor ZM447439 for 72 h.