Taxol-induced rose microtubule polymerization in vitro and its inhibition by colchicine

Abstract

Tubulin was isolated from cultured cells of rose (Rosa, sp.cv. Paul's scarlet) by DEAE-Sephadex A50 chromatography, and the taxol-induced polymerization of microtubules in vitro was characterized at 24 degrees C by turbidity development, sedimentation analysis, and electron microscopy. Numerous, short microtubules were formed in the presence of taxol, and maximum levels of turbidity and polymer yield were obtained at approximately 2:1 molar ratios of taxol to tubulin. The critical concentration of rose tubulin for polymerization in saturating taxol was estimated to be 0.21 mg/ml. Colchicine inhibited the taxol-induced polymerization of tubulin as shown by sedimentation assays; however, much higher concentrations of colchicine were required for the inhibition of taxol-induced rose tubulin assembly than for inhibition of taxol-induced mammalian brain tubulin assembly. On the basis of the relative sensitivity of rose tubulin assembly to taxol and its insensitivity to colchicine, we propose that the taxol-binding site(s) on plant and animal tubulins have been more conserved over evolution than the colchicine-binding site(s).