Taxol, a potent promoter of microtubule polymerization in vitro, induces massive assembly offree microtubules in cultured cells as visualized by immunocytochemistry and electron microscopy. The centrosomes and kinetochores largely lost their capacity to organize microtubule assembly, as became evident by the disappearance ofthe cytoplasmic microtubule complex and the mitotic spindle. The taxol-induced microtubules were partially resistant to nocodazole, an inhibitor of tubulin polymerization. Moreover, taxol induced microtubule assembly in cells pretreated with nocodazole. Increasing the ratio ofnocodazole to taxol restored the ability of the centrosomes and kdnetochores to specifically induce microtubule assembly in their immediate vicinity. The data suggest that taxol lowers the critical tubulin concentration in vivo as well as in vitro and that the organizing capacity of the microtubule-organizing centers depends on the cytoplasmic polymerization threshold.Taxol, an experimental antitumor drug (1) isolated from Taxus brevifolia, was recently shown by Schiff et aL (2, 3) to affect microtubule assembly in vitro and in living cells. It essentially eliminated the initial lag phase and decreased the critical tubulin concentration to less than 0.01 mg/ml. The rate and extent of the polymerization was increased and the microtubules were relatively resistant to depolymerization by cold and CaCl2. In living cells (3), taxol was shown to be a potent inhibitor of HeLa and mouse. fibroblast replication. The cells were blocked in the G2 and M phases of the cell cycle. It was inferred that this was due to the stabilization of cytoplasmic microtubules. Indirect immunofluorescence was used to show that taxoltreated cells displayed bundles of microtubules radiating from a common site in addition to their cytoplasmic microtubules. These microtubules were resistant to treatment with steganacin or incubation of the cells at low temperature, both of which disintegrated microtubules in control cells. Ultrastructural observations showed that the mitotic cells contained microtubule bundles but no normal spindle. It was concluded that the inability of the cells to form a mitotic spindle in the presence of taxol could be due to the fact that the cells were unable to depolymerize their microtubule cytoskeletons (3).We have investigated the effects of taxol on the cytoplasmic microtubule complex and the mitotic spindle in cultured cells. Our observations and conclusions differ partly from those published previously (3). Taxol apparently induces the assembly of free microtubules in the cytoplasm, not attached to the centrosomes or kinetochores. The preexisting microtubules, attached to the organizing centers, are not stabilized and disappear gradually.The publication costs ofthis article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact. MATERIAL AND METHODSPt K2 potoroo cells, C3H mouse 3T3 cells, and human...

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Abstract. Immunofluorescence with specific peptide antibodies has previously established that tyrosinated (Tyr) and detyrosinated (Glu) tubulin, the two species generated by posttranslational modification of the COOH-terminus of a-tubulin, are present in distinct, but overlapping, subsets of microtubules in cultured cells

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