Taxol-induced microtubule asters in mitotic extracts of Xenopus eggs: requirement for phosphorylated factors and cytoplasmic dynein.

Abstract

Abstract. Taxol, a microtubule stabilizing drug, induces the formation of numerous microtubule asters in the cytoplasm of mitotic cells (De Brabander, M., G. Geuens, R. Nuydens, R. Willebrords, J. DeMey. 1981. Proc. Natl. Acad. Sci. USA. 78:5608-5612). The center of these asters share with spindle poles some characteristics such as the presence of centrosomal material and calmodulin. We have recently reproduced the assembly of taxol asters in a cell-free system (Buendia, B., C. Antony, F. Verde, M. Bornens, and E. . J. Cell Sci. 97:259-271) using extracts of Xenopus eggs. In this paper, we show that taxol aster assembly requires phosphorylation, and that they do not grow from preformed centers, but rather by a reorganization of microtubules first crosslinked into bundles. This process seems to involve sliding of microtubules along each other and we show that cytoplasmic dynein is required for taxol aster assembly. This result provides a possible functional basis to the recent findings, that dynein is present in the spindle and enriched near spindle poles (Pfarr, C. M., M. Cove, P. M. Grissom, T. S. Hays, M. E. Porter, and

Low-dose laulimalide represents a novel molecular probe for investigating microtubule organization

“…Several studies suggest these asters arise from increased polymerization of MT states in the cytoplasm. 35,36 Alternatively, Hornick et al propose that multiple MT aster formation is driven by a redistribution of pre-assembled microtubules from the centrosomes to the cell cortex. 32 Our data suggest that both laulimalide and docetaxel drive MT assembly and lower the barrier for effective nucleation of minus-end MTs; this is supported by the formation of multiple acentrosomal poles without centrosomal proteins or pericentriolar material.…”

Protein 4.1R, a Microtubule-associated Protein Involved in Microtubule Aster Assembly in Mammalian Mitotic Extract

“…Dynein-dependent and centrosome-free spindle-pole formation was mimicked by the induction of microtubule asters in the presence of the drug taxol in both Xenopus (46) and mammalian systems (25). An in vitro aster assembly assay using HeLa mitotic extracts, an assay that recapitulates many of the structural events in mitosis, has been used widely to characterize the function of a number of proteins localized at the spindle poles (29 -33).…”

The dynein genes of Paramecium tetraurelia: the structure and expression of the ciliary beta and cytoplasmic heavy chains.

“…Each heavy chain has two structural domains that perform distinct functions: the catalytic head domain, which contains the MgATPase and ATP-sensitive microtubule-binding activities; and the flexible dynein specialization occurs in the motor domain: all axonemal dyneins carry an outer doublet microtubule as their cargo, but each arm is specialized to generate precise mechanical force so that the dynein arms working together initiate and propagate ciliary bends (Asai and Brokaw, 1993;Brokaw, 1994). In contrast to axonemal dynein, cytoplasmic dynein carries a diverse array of molecular cargoes (Schnapp and Reese, 1989;Verde et al, 1991;Corthesy-Theulaz et al, 1992;Lin and Collins, 1992;Aniento et al, 1993;Vaisberg et al, 1993;Saunders et al, 1995;Wang et al, 1995) and is probably less specialized in force production. The current view is that cytoplasmic dynein is a homodimer of the heavy chain MAPlC (Vallee et al, 1988;Neely et al, 1990).…”