Treatment with colchicine or vinblastine, both inhibitors of microtubule assembly, renders quiescent 3T3 cells in an "activated state" as evidenced by induction of DNA synthesis and other criteria. Microtubule disassembly caused by colchicine or vinblastine brings about a dramatic expansion of total cellular UTP pools with a concomitant diminution in total cellular ATP pools, thus resulting in a marked imbalance in total cellular nucleotide pools. Colchicine and vinblastine also stimulate total cellular RNA synthesis without enhancing uridine phosphorylation, suggesting that these drugs affect the G, phase of the cell cycle at a point beyond the enhancement of uridine phosphorylation that usually accompanies mitogenic stimulation of quiescent mammalian cells.The markedly expanded cellular UTP pools appear to be necessary for initiation of the colchicine-stimulated DNA synthesis because decreasing cellular UTP pools by addition of D-glucosamine results in a selective inhibition of DNA synthesis in the colchicine-stimulated, but not control, cells. Furthermore, Dglucosamine exerts its inhibitory effect only when it is present in the cultures within the first 14 hr after colchicine treatment. When added at 21 hr, D-glucosamine still decreases cellular UTP pools, but it is no longer inhibitory for DNA synthesis, which commences 14-16 hr after colchicine stimulation. Taxol, an antitumor drug, prevents microtubule disassembly and also blocks such events as expansion of total cellular UTP pools and stimulation of RNA and DNA synthesis, indicating that microtubule depolymerization acts as a primary event initiating the process of cell activation induced by colchicine.suggest that MT depolymerization is involved in modulating the initiation of DNA replication after mitogenic stimulation, although the underlying mechanism is not known.Since MT assembly in vitro is accompanied by GTP hydrolysis (12, 13), we have studied the effect of the colchicine-stimulated GO/G1 to S phase transition of 3T3 cells on the levels of cellular acid-soluble nucleotide pools, which have been shown to possess a regulatory role in the progression of mammalian cells along their cycle (14-17). We report here that MT disassembly induced by colchicine or vinblastine results in a sharp expansion of total cellular UTP pools with a concomitant diminution in total ATP pools and thus creates an imbalance in the cellular nucleotide pools as evidenced by large increases in the UTP/ATP ratio. In addition, we report that total cellular RNA synthesis, but not uridine phosphorylation, is stimulated following colchicine treatment. Furthermore, we also report that D-glucosamine effectively decreases the expanded cellular UTP pools and causes a selective inhibition of DNA synthesis in colchicinetreated, but not control, cells. Finally, we also report that taxol treatment effectively blocks events that normally occur following MT disassembly induced by colchicine or vinblastine. Taken together, our results suggest that the markedly increased cellular UTP p...