TRYPROSTATIN-A, A SPECIFIC AND NOVEL INHIBITOR OF MICROTUBULE ASSEMBLY

We have investigated the cell cycle inhibition mechanism and primary t arget of tryprostatin A (TPS-A) purified from Aspergillus fumigatus. T PS-A inhibited cell cycle progression of asynchronously cultured 3Y1 c ells in the M phase in a dose- and time-dependent manner. In contrast, TPS-B (the demethoxy analogue of TPS-A) showed cell-cycle non-specifi c inhibition on cell growth even though it inhibited cell growth at lo wer concentrations than TPS-A. TPS-A treatment induced the reversible disruption of the cytoplasmic microtubules of 3Y1 cells as observed by indirect immunofluorescence microscopy in the range of concentrations that specifically inhibited M-phase progression. TPS-A inhibited the assembly in vitro of microtubules purified from bovine brains (40 % in hibition at 250 mu M); however, there was little or no effect on the s elf-assembly of purified tubulin when polymerization was induced by gl utamate even at 250 mu M TPS-A. TPS-A did not inhibit assembly promote d by taxol or by digestion of the C-terminal domain of tubulin. Howeve r, TPS-A blocked the tubulin assembly induced by inducers interacting with the C-terminal domain, microtubule-associated protein 2 (MAP2), t au and poly-(L-lysine). These results indicate that TPS-A is a novel i nhibitor of MAP-dependent microtubule assembly and, through the disrup tion of the microtubule spindle, specifically inhibits cell cycle prog ression at the M phase.