REMOVAL OF BETA(III) ISOTYPE ENHANCES TAXOL-INDUCED MICROTUBULE ASSEMBLY

The interaction of beta(III)-depleted tubulin with taxol was investiga ted. A monoclonal antibody against the beta(III) tubulin isotype was i mmobilized on a sepharose 4B column and used to remove the beta(III) t ubulin isotype from unfractionated tubulin. The assembly of beta(III)- depleted tubulin in the presence of taxol was enhanced compared to unf ractionated tubulin. The critical concentration of unfractionated tubu lin in the presence of 10 muM taxol is 0.4 mg/ml, while the critical c oncentration of beta(III)-depleted tubulin is 0.16 mg/ml. At different concentrations of taxol, the assembly of beta(III)-depleted tubulin i s increased relative to that of unfractionated tubulin and reaches the maximum at about a 1:1 ratio of tubulin and taxol. The assembly of un fractionated tubulin and beta(III)-depleted tubulin has also been stud ied by electron microscopy. After 2 minutes at 37-degrees-C, unfractio nated tubulin assembly in the presence of 10 muM taxol results only in ribbon-like and ring structures; there are no visible microtubules. B y 5 minutes, microtubules appear and increase in length. The assembly of beta(III)-depleted tubulin in the presence of 10 muM taxol occurs m ore quickly. In contrast to the case with unfractionated tubulin, beta (III)-depleted tubulin assembles within 2 minutes into microtubules wh ich increase in length with time. At 30 minutes, microtubules assemble d from beta(III)-depleted tubulin are shorter than the microtubules as sembled from unfractionated tubulin. There is no visible difference be tween the microtubules assembled from unfractionated tubulin and beta( III)-depleted tubulin. Taxol-induced beta(III)-depleted tubulin assemb ly is more resistant to the inhibiting effect of podophyllotoxin and c olchicine. It is also less sensitive to the inhibiting effect of cold temperature.