Effects of the Tubulin-Colchicine Complex on Microtubule Dynamic Instability

Abstract

The effects of the tubulin-colchicine complex (Tu-Col) on the dynamic behavior of microtubules have been examined under steady-state conditions in vitro. The addition of Tu-Col to tubulin microtubules at steady state results in only partial microtubule disassembly. Nevertheless, both the rate and the extent of tubulin exchange into microtubules are markedly suppressed by concentrations of Tu-Col which are low relative to the total amount of free tubulin. In addition, the time-dependent changes in microtubule length distribution, characteristic of dynamic instability, are suppressed by the addition of Tu-Col. Examination by video-enhanced dark-field microscopy of individual microtubules in the presence of Tu-Col shows that the principal effect of this complex is to reduce the growth rate at both ends of the microtubule. We have used computer simulation to rationalize the action of Tu-Col in terms of its effects on the experimentally observable parameters, namely, the rates of growth and shortening and the mean lifetimes of growth and shortening, which provide an empirical description of the dynamic behavior of microtubules. The results have been interpreted within the framework of the lateral cap formulation for microtubule dynamic instability [Martin, S. R., Schilstra, M. J., & Bayley, P. M. (1993) Biophys. J. 65, 578-596]. The simplest model mechanism requires only that Tu-Col binds to the microtubule end and inhibits further addition reactions in either the 5-start or the 8-start direction of the microtubule lattice. Monte Carlo simulations show that Tu-Col can, in this way, cause major suppression of the dynamic transitions of microtubules without inducing bulk microtubule disassembly. This type of mechanism could be important for the regulation of microtubule dynamics in vivo.

Tubulin Folding Cofactors as GTPase-activating Proteins

“…31). These toxins can poison microtubules substoichiometrically; if poisoned subunits are added to the growing end of a microtubule, the growth and stability of the whole tubule is affected (32,37). By analogy, the quality control of tubulin by cofactors in vivo may be important for protecting the microtubules of the cell from poisoning by misfolded, non-GTP-hydrolyzing subunits.…”

Phosphatase inhibition in human neuroblastoma cells alters tau antigenicity and renders it incompetent to associate with exogenous microtubules

“…Accordingly, experimental alteration in kinase and phosphatase activities in neuronal cell culture systems represent a useful approach towards unraveling antecedent events. In previous studies, phosphatase inhibition by OA treatment has been shown to disrupt neuroblastoma MTs [50,56] and increase hyperphosphorylated tau immunoreactivity in cultured neurons, neuroblastoma [39][40][41]57], brain slices [39] and brain in situ [40,41]. While tau that has been dissociated from MTs is susceptible to hyper- Tubulin purified from bovine brain by two assembly/disassembly cycles was incubated for 30 min at 4°C in the absence of taxol and at 30°C in the presence of 10,uM taxol, then subjected to high-speed centrifugation and separated from endogenous MAPs as described in section 2.…”

Response of microtubules to the addition of colchicine and tubulin–colchicine: evaluation of models for the interaction of drugs with microtubules

“…These complexes bind rapidly and reversibly to the microtubule ends, with an affinity (K d " 0.3 µM at 35 mC) similar to that of unbound tubulin [9,11]. TuCol inhibits microtubule elongation [11][12][13][14][15], but does not seem to affect the nucleation process [11,16]. TuCol inhibits growth at both microtubule ends to a similar extent [13][14][15].…”

“…TuCol inhibits microtubule elongation [11][12][13][14][15], but does not seem to affect the nucleation process [11,16]. TuCol inhibits growth at both microtubule ends to a similar extent [13][14][15]. Under suitable conditions, relatively large amounts of TuCol are incorporated into the microtubule lattice [12,17,18], but the co-assembly of tubulin and TuCol into the Nevertheless, drug-induced disassembly can be extremely slow, because the frequency of addition reactions increases as the concentration of soluble dimers increases.…”