Fig. 5.

Sin3a is required for cell cycle progression and genome maintenance in ES cells. Sin3a^Flox/–:CreER^T2 ES cells were either untreated (No Drug) or treated with 400 nM 4-hydroxytamoxifen (+ 4-OHT) and harvested at the indicated times after tamoxifen addition. A. Cell cycle distribution of Sin3a^Flox/–:CreER^T2 ES cells with and without 4-OHT treatment. DNA content is indicated by propidium iodide staining (X-axis), where 2C is the diploid DNA content in G1 and 4C is the replicated diploid DNA content in G2. B. Mean % of tamoxifen-treated (grey) and untreated (black) Sin3a^Flox/–:CreER^T2 ES cells with a G2 (4 C) DNA content that are positive for phospho-histone H3 (S10), indicating mitotic entry. Error bars represent standard deviation from the mean among independent replicates, and p < 0.0001 according to Fisher's exact test. C. Mean % of live, unfixed Sin3a^Flox/–:CreER^T2 ES cells staining positive for Annexin V, indicating apoptosis, after 36 h of tamoxifen (+ 4-OHT) or mock (No Drug) treatment. D. Western blots of total lysates prepared from Sin3a^Flox/–:CreER^T2 ES cells at the indicated times after tamoxifen addition. Protein names beginning with ‘p’ indicate the antibody specifically recognises the phosphorylated form of the protein.