Figure 4.

ML111 induced prometaphase arrest without altering microtubule dynamics. (A) Cell cycle analyses by flow cytometry in asynchronous SK-N-MC cells treated with 0.1% DMSO (upper panel) or 100 nM ML111 (lower panel). (B) Immunoblot analysis for cyclin B1, p-H3^Ser10, and CDC20 in lysates from asynchronized SK-N-MC cells treated with 100 nM ML111 or 0.1% DMSO. Total histone H3 (H3) antibody was used as a control. (C) Cell-free tubulin polymerization assay. ML111 neither stimulated nor inhibited tubulin polymerization in vitro. Paclitaxel and colchicine were used as tubulin polymerization inducer and inhibitor agents, respectively. General tubulin buffer (Cytoskeleton, Inc.) was used as a control. AU: absorbance units. (D) ML111 induced prometaphase arrest. Asynchronized SK-N-MC cells were treated with either DMSO (0.1%) or ML111 (25 nM) for 12 h. The nuclei were stained with DAPI (blue), microtubules (α-tubulin, red), and mitotic chromosomes (p-H3^Ser10, green). One representative result was shown from three independent experiments. Magnification: 63×. Scale bar = 10 μm.