ES cells depleted of regulators of essential processes accumulate DNA damage but exhibit distinct cell cycle phenotypes. (A, left) Western blots for p53, γ-H2AX, and actin in ES cells (ESCs) and primary MEFs depleted of the indicated proteins. (right) p53 protein levels with (+) or without (−) UV exposure. For all blots, actin levels are shown as a loading control. Asterisks denote background bands of unknown identity. (B) Western blots for p53, p53 phosphorylated on serine 15, p53 phosphorylated on serine 20, p21, and actin in ES cells and primary MEFs depleted of the indicated proteins. (A and B) The positions of molecular mass standards (in kilodaltons) are shown. (C) Cell cycle profiles of ES cells knocked down for the proteins indicated. Cell number is plotted on the vertical axis, and DNA content is plotted on the horizontal axis. The arrowhead denotes a small 8N population observed in Smc2 KD ES cells, and the dashed lines center on the 2N and 4N peaks in control cells. The percentages of cells in each phase of the cell cycle or exhibiting tetraploidy (8N) are shown to the right.