Figure 4.
Disrupting the H3pT3–Survivin interaction delays metaphase–anaphase transition in a Bub1-dependent way.(A and B) HeLa and the indicated Survivin cell lines stably expressing H2B-GFP were released from STB, and then mitosis progression was analyzed by time-lapse live imaging. The time from NEB to metaphase chromosome alignment and from metaphase to anaphase onset was determined in ≥66 cells (A). Selected frames of the videos are shown (B). See also Videos 1, 2, and 3. (C) DMSO or 0.3–0.6 µM 5-ITu was added to HeLa cells stably expressing H2B-GFP 7 h after release from STB, and then mitosis progression was analyzed in ≥64 cells. See also Videos 4, 5, and 6. (D and E) MG132 was added to HeLa cells stably expressing H2B-GFP 10.5 h after release from STB. After 2-h incubation in MG132, cells were released into DMSO or 0.3–1.0 µM 5-ITu, and then mitosis progression was analyzed by time-lapse live imaging. The time from MG132 release to anaphase onset was determined in ≥200 cells (D). Selected frames of the videos are shown (E). See also Videos 7 and 8. (F and G) DMSO or BAY 1816032 was added to the indicated Survivin cell lines stably expressing H2B-GFP 7 h after release from STB, and then mitosis progression was analyzed in ≥66 cells (F). Selected frames of the videos are shown (G). (H and I) DMSO or the indicated inhibitors was added to HeLa cells stably expressing H2B-GFP at 7 h after release from STB, and then mitosis progression was analyzed in ≥45 cells (H). Selected frames of the videos are shown (I). Means and SDs are shown (A, C, D, F, and H; unpaired t test). N.S., not significant. Time stated in h:min. Scale bars, 10 μm. See also Fig. S1.

Disrupting the H3pT3–Survivin interaction delays metaphase–anaphase transition in a Bub1-dependent way. (A and B) HeLa and the indicated Survivin cell lines stably expressing H2B-GFP were released from STB, and then mitosis progression was analyzed by time-lapse live imaging. The time from NEB to metaphase chromosome alignment and from metaphase to anaphase onset was determined in ≥66 cells (A). Selected frames of the videos are shown (B). See also Videos 1, 2, and 3. (C) DMSO or 0.3–0.6 µM 5-ITu was added to HeLa cells stably expressing H2B-GFP 7 h after release from STB, and then mitosis progression was analyzed in ≥64 cells. See also Videos 4, 5, and 6. (D and E) MG132 was added to HeLa cells stably expressing H2B-GFP 10.5 h after release from STB. After 2-h incubation in MG132, cells were released into DMSO or 0.3–1.0 µM 5-ITu, and then mitosis progression was analyzed by time-lapse live imaging. The time from MG132 release to anaphase onset was determined in ≥200 cells (D). Selected frames of the videos are shown (E). See also Videos 7 and 8. (F and G) DMSO or BAY 1816032 was added to the indicated Survivin cell lines stably expressing H2B-GFP 7 h after release from STB, and then mitosis progression was analyzed in ≥66 cells (F). Selected frames of the videos are shown (G). (H and I) DMSO or the indicated inhibitors was added to HeLa cells stably expressing H2B-GFP at 7 h after release from STB, and then mitosis progression was analyzed in ≥45 cells (H). Selected frames of the videos are shown (I). Means and SDs are shown (A, C, D, F, and H; unpaired t test). N.S., not significant. Time stated in h:min. Scale bars, 10 μm. See also Fig. S1.

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