Chk2-deficient cells exit mitosis when the majority of kinetochores are unattached. (A and B) Mitotic index analysis. Cells transfected with negative siRNA (control), Chk2 siRNA (siChk2), Mps1 siRNA (siMps1), or treated with Chk2 inhibitor II were incubated with 3.32 µM nocodazole (A) or taxol (B). ***, P < 0.001. (C) Wild-type (WT) or Chk2^−/− DT40 cells were treated with 3.32 µM nocodazole (nocod) or taxol. Error bars show the SD from the means from three independent experiments. ***, P < 0.001 compared with WT plus nocodazole. (D) An example of mitotic exit. Cells expressing H2B-GFP were treated with 3.32 µM nocodazole in the absence or presence of Chk2 inhibitor II (inh II) and analyzed by time-lapse microscopy. Phase-contrast (right) and fluorescence images (left) of a cell arrested with condensed chromatin (top row) or a cell that exited mitosis and formed micronuclei (bottom row). Time is from the start of chromatin condensation. See also Video 1 and Video 2. (E) Microtubule–kinetochore attachments in CHO cells. Values in parentheses indicate the frequency of unattached kinetochores from n cells. A 4× magnification of kinetochores is shown. Arrows indicate unattached kinetochores. (F) Cdk1-associated histone H1 kinase activity and Western blot analysis of immunoprecipitated (IP) Cdk1. Cells were treated with 3.32 µM nocodazole in the absence (control) or presence of Chk2 inhibitor II or AZ3146 (AZ). Phosphorylated H1 (pH1) values at 0 h were taken as 1. (G) Western blot analysis of total Cyclin B and actin in cells treated as in F. Values at 0 h were taken as 1. (H) Cyclin B fluorescence in cells treated as in F. Values show mean Cyclin B fluorescence intensity from n cells ± SDs. Bars, 5 µm.