Microtubule polarity in taxol-treated isolated spindles

Spindles were isolated from Spisula oocytes in the presence or absence of taxol to determine if cell lysis procedures previously used to analyze spindle microtubule polarity may have resulted in the loss of any microtubules of a particular polarity. Electrophoretic analysis indicated that spindle preparations isolated in the presence of taxol contained considerably more tubulin than did those isolated in the absence of taxol. Although there was no corresponding increase in the birefringence of the taxol-treated isolated spindles, thereby suggesting that they did not possess a greater concentration of aligned microtubules between the poles and the equatorial plate, electron microscopy revealed that the taxol-treated isolated spindles did, in fact, contain more microtubules than did the nontaxol-treated isolated spindles. These microtubules were predominantly localized immediately around the centrioles, suggesting that assembly may have occurred during the lysis into taxol. The spindles were decorated with Chlamydomonas dynein to display microtubule polarity. The majority of microtubules, located between the chromosomes and poles, possessed a uniform polarity in both taxol- and nontaxol-treated isolated spindles. In the region of the chromosomes, however, up to 45% of the microtubules were oriented with a polarity opposite to the remainder and were intermingled among them. It is probable that microtubules with opposite polarities arise from the opposing poles and overlap in the equatorial region. The extent of overlap appears to be quite substantial and thus may be of significance during mitotic movements.