FIGURE 4:

Importazole causes astral MT-dependent spindle movement. (A) Selected frames from 60-min movies of HeLa cells stably expressing GFP-tubulin and mCherry-H2B treated with DMSO, 40 μM importazole, 20 nM nocodazole, or 20 nM nocodazole and 40 μM importazole. Spindles in DMSO-treated cells move to the cell center before anaphase, whereas spindles in importazole-treated cells move about the cytoplasm. Spindles in nocodazole- and nocodazole/importazole-treated cells remained close to the center of the cell. (B) Mean square displacement as a function of time measured for mitotic spindles from GFP-tubulin– and mCherry-H2B–expressing HeLa cells under the indicated treatment conditions. (C) Asynchronously growing HeLa cells were treated with DMSO, 40 μM importazole, 20 nM nocodazole, or 20 nM nocodazole and 40 μM importazole for 1 h, and cells displaying spindle-centering defects were quantified as a percentage of total mitotic cells. Dashed white lines indicate the cell cortex, white Xs indicate the cell center, and white Os indicate the spindle center. Scale bars, 10 μm. Spindles from six cells were analyzed for each condition in B. For C, n = 3, and 100 cells were counted per condition. Bars, SE. Asterisk denotes statistical significance from all other conditions (p < 0.001).