Taxol suppresses dynamics of individual microtubules in living human tumor cells - PubMed (original) (raw)

Taxol suppresses dynamics of individual microtubules in living human tumor cells

A M Yvon et al. Mol Biol Cell. 1999 Apr.

Free PMC article

Abstract

Microtubules are intrinsically dynamic polymers, and their dynamics play a crucial role in mitotic spindle assembly, the mitotic checkpoint, and chromosome movement. We hypothesized that, in living cells, suppression of microtubule dynamics is responsible for the ability of taxol to inhibit mitotic progression and cell proliferation. Using quantitative fluorescence video microscopy, we examined the effects of taxol (30-100 nM) on the dynamics of individual microtubules in two living human tumor cell lines: Caov-3 ovarian adenocarcinoma cells and A-498 kidney carcinoma cells. Taxol accumulated more in Caov-3 cells than in A-498 cells. At equivalent intracellular taxol concentrations, dynamic instability was inhibited similarly in the two cell lines. Microtubule shortening rates were inhibited in Caov-3 cells and in A-498 cells by 32 and 26%, growing rates were inhibited by 24 and 18%, and dynamicity was inhibited by 31 and 63%, respectively. All mitotic spindles were abnormal, and many interphase cells became multinucleate (Caov-3, 30%; A-498, 58%). Taxol blocked cell cycle progress at the metaphase/anaphase transition and inhibited cell proliferation. The results indicate that suppression of microtubule dynamics by taxol deleteriously affects the ability of cancer cells to properly assemble a mitotic spindle, pass the metaphase/anaphase checkpoint, and produce progeny.

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Figures

Figure 1

Inhibition of proliferation of Caov-3 ovarian carcinoma cells (○) and A-498 kidney carcinoma cells (●) by taxol (24 h). Cell proliferation was determined by counting live cells at the time of taxol addition and 24 h later. Values >100% inhibition of proliferation indicate net loss of cells over the 24-h duration of taxol incubation. Values are means and SEs of 11 independent experiments for A-498 cells and 4 experiments for Caov-3 cells.

Figure 2

Dynamic behavior of microtubules in living Caov-3 cells in the absence of drug (A–C) or incubated with 30 nM taxol for 4 h (D–F). Images were collected at 2-s intervals; panels shown are 10 s apart. Arrowheads in A indicate microtubules that undergo growing and shortening events. Arrowheads in D indicate stable microtubules. Bar, 2 μm.

Figure 3

Life history plots of microtubules in control (A) and taxol-treated (B) Caov-3 cells. The graphs represent the excursions made by individual microtubules during an 86- to 94-s observation period. Microtubules in the presence of taxol are characterized by longer pause events and less extensive growing and shortening events than those in control cells. The plots are offset on the _y_-axis for clarity.

Figure 4

Arrangement of microtubules in A-498 kidney cells (A, B) and Caov-3 ovary cells (C, D) in interphase in the absence (A, C) or presence (B, 100 nM; D, 30 nM) of taxol (4-h incubation). Cells were stained with an antibody to α-tubulin and imaged using confocal microscopy. A single optical section is shown. Microtubules retracted slightly from the plasma membrane and formed occasional bundles (arrows in B and D) after taxol incubation. Bar, 20 μm.

Figure 5

Microtubules in mitotic A-498 kidney cells (A, B) and Caov-3 ovary cells (C, D) in the absence (A, C) or presence (B, D) of taxol (100 nM and 30 nM, respectively). Cells were incubated with taxol and stained for microtubules (left panels) as described for Figure 4. They were subsequently stained with propidium iodide for DNA (right panels of each pair), and an extended focus series using confocal microscopy was collected. (A, B) Control spindles are bipolar, with congressed chromosomes forming a compact metaphase plate midway between the poles. After 4-h incubation with taxol, spindle morphology is dramatically altered. Many spindles are multipolar (D) with uncongressed chromosomes (B and D). Bar, 10 μm.

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Taxol suppresses dynamics of individual microtubules in living human tumor cells - PubMed (original) (raw)