FIGURE 2.

STAT3 is phosphorylated on serine 727 during G₂ arrest. A, HT29 cells were treated or not with different concentration of sn38 for 10–14 days. Colony formation was then counted using an inverted microscope, and the growth of non-treated cells was set up at 100%. Clonogenic survival was then plotted as a fraction relative to these untreated cells (n = 5 ± S.D.). In parallel, growing HT29 cells were treated with sn38 (5 ng/ml) for 48 h, and DNA content and apoptosis were then evaluated by flow cytometry (n = 5). B, HT29 cells were treated with sn38 (5 ng/ml) or IL-6 (20 ng/ml) as indicated, and DNA content and serine phosphorylation were then analyzed by flow cytometry analysis using polyclonal antibodies directed against the serine 727-phosphorylated form of STAT3 (n = 3). C, growing HCT116 cells were treated or not with sn38 for different times as indicated. The percentage of senescent cells was evaluated as the number of cells expressing SA-β-gal activity and micronuclei (left part, n = 3). In parallel, DNA content was evaluated by flow cytometry after 48 h (right part), and the phosphorylation of STAT3 on its serine residue was analyzed by Western blot as described above (n = 3, bottom).