Microtubule dynamics in vitro are regulated by the tubulin isotype composition.

Abstract

The growing and shortening dynamics of individual bovine brain microtubules at their plus ends at steady state in vitro, assembled from isotypically pure a4u, abim, or acsv tubulin dimers, were determined by differential interference contrast video microscopy. Microtubules assembled from the purified afm isotype were considerably more dynamic than microtubules made from the apu or a4iv isotypes or from unfractionated phosphocellulose-purified tubulin. Furthermore, increasing the proportion of the aoil isotype in a mixture of the alI and atom isotypes suppressed microtubule dynamics, demonstrating that microtubule dynamics can be influenced by the tubulin isotype composition.The data support the hypothesis that cells might determine the dynamic properties and functions of its microtubules in part by altering the relative amounts of the different tubulin isotypes.Microtubules (1) are required for structural organization and for many kinds of movements within the eukaryotic cell cytoplasm. They are especially prominent in the central nervous system where they appear to be essential for the organization and function of axonal and dendritic processes of neurons. Although it is unclear how microtubule functions in cells are controlled, a large body of evidence indicates that the dynamic properties of the microtubules play an important role (for reviews, see refs. 2 and 3). For example, microtubule dynamics increase dramatically when cells progress from interphase to mitosis (4-6), and the rapid dynamics of spindle microtubules appear to be important for establishing spindle microtubule organization, for facilitating the linkage of chromosomes to the spindles, and for chromosome movement (2-7).Little is known about how microtubule dynamics are regulated in cells. In vitro and in cells, microtubule ends switch between states of growing and shortening, a process known as "dynamic instability" (8-13), apparently due to the gain and loss ofa stabilizing GTP-or GDP-P1-liganded tubulin cap at the microtubule ends. Also, in vitro and in cells, net growing of microtubules can occur at one microtubule end and net shortening can occur at the opposite end, a process termed "treadmilling" or "flux" (9, 14-16). Both dynamic instability and treadmilling are logical targets for control. Biochemical studies in vitro have indicated that microtubule dynamics in cells could be regulated at several levels. One possibility is that control of microtubule dynamics involves regulation of the gain and loss of the stabilizing cap. A second possibility is that control of microtubule dynamics occurs through interactions of microtubule-associated proteins (MAPs) with microtubule surfaces and ends.Another possible mechanism for control of microtubule polymerization dynamics could involve the isotypic composition of the tubulin itself. Tubulin is composed oftwo 50-kDa polypeptide subunits, a and (3, which exist in multiple forms called isotypes. For example, mammalian brain tubulin consists of at least five a-and five j-tubulin isotypes (17,...

Altered β-tubulin isotype expression in paclitaxel-resistant human prostate carcinoma cells

“…In this study, we observed an increase in P. (Jordan et al, 1992;Derry et al, 1995;Dhamodharan et al, 1995). Using purified 0-tubulin isotypes, Panda et al (1994) have shown that microtubule dynamics are regulated by the tubulin isotype composition.…”

Drug resistance associated with loss of p53 involves extensive alterations in microtubule composition and dynamics

“…We also observed that the relative proportion of Class IV was greatly increased (seven-fold), whereas the proportions of classes I and II were reduced in mut-p53 cells. The b-tubulin isotype composition of microtubules in vitro can alter their dynamic properties (Panda et al, 1994;Derry et al, 1997), suggesting that the differences we observed in isotype composition between wt-p53 and mut-p53 cells may contribute to the observed differences in dynamics. These alterations may also be involved in drug resistance to microtubule-targeted drugs (Burkhart et al, 2001).…”

Modulation of endogenous β -tubulin isotype expression as a result of human βIIIcDNA transfection into prostate carcinoma cells

“…Several in vitro studies have demonstrated that the β tubulin isotype composition of microtubules determines the differences in their functions including dynamics, assembly properties and sensitivity to antimicrotubule agents (Banerjee et al, 1990Benerjee and Luduena, 1992;Luduena, 1993, 1994;Panda et al, 1994Panda et al, , 1997Derry et al, 1997). Because antimicrotubule agents are widely used in cancer therapy, tubulin isotype composition of tumour cells may be an important factor in the success of therapy.…”

“…αβ III dimers assembled at a very slow rate and had the highest critical concentration. However, in vitro analysis by Panda et al (1994) has shown that microtubules assembled from αβ III dimers were more dynamic than the microtubules from αβ II and αβ IV isotypes. They further demonstrated that addition of αβ II dimers to αβ III dimers suppressed microtubule dynamics.…”