INVIVO AND INVITRO STUDIES ON THE ROLE OF HMW-MAPS IN TAXOL-INDUCED MICROTUBULE BUNDLING

  • 1 January 1984
    • journal article
    • research article
    • Vol. 33  (1) , 134-143
Abstract
The involvement of high MW microtubule-associated proteins (HMW-MAP) in the process of taxol-induced microtubule bundling was studied using immunofluorescence and EM. Immunofluorescence microscopy shows that HMW-MAP are released from microtubules in [rat] granulosa cells which were extracted in a Triton X-100 microtubule-stabilizing buffer (T-MTSB), unless the cells are pretreated with taxol. 1.0 .mu.M taxol treatment for 48 h results in microtubule bundle formation and the retention of HMW-MAP in these cells on extraction with T-MTSB. EM demonstrates that microtubules in control cytoskeletons are devoid of surface structures whereas the microtubules in taxol-treated cytoskeletons are decorated by globular particles of a mean diameter of 19.5 nm. The assembly of 3 .times. cycled whole microtubule protein (tubulin plus associated proteins) in vitro in the presence of 1.0 .mu.M taxol, results in the formation of closely packed microtubules decorated with irregularly spaced globular particles, similar in size to those observed in cytoskeletons of taxol-treated granulosa cells. Microtubules assembled in vitro in the absence of taxol display prominent filamentous extensions from the microtubule surface and center-to-center spacings greater than that observed for microtubules assembled in the presence of taxol. Brain microtubule protein was purified into 6 s and HMW-MAP-enriched fractions, and the effects of taxol on the assembly and morphology of these fractions, separately or in combination, were examined. Microtubules assembled from 6 s tubulin alone or 6 s tubulin plus taxol (without HMW-MAP) were short, free structures whereas those formed in the presence of taxol from 6 s tubulin and a HMW-MAP-enriched fraction were extensively crosslinked into aggregates. Taxol induces microtubule bundling by stabilizing the association of HMW-MAP with the microtubule surface which promotes lateral aggregation.