It is generally assumed that these bundles, termed kinetochore fibers (K-fibers), are formed via repetitive MT capture, however this mechanism has not been directly validated ( McEwen et al., 1997).įor years, the centrosomal and acentrosomal routes of spindle formation have been viewed as mutually exclusive pathways, in that any given cell is thought to use one or the other, but not both mechanisms. After the initial MT capture, kinetochores develop a bundle of 15–30 parallel MTs that connect them to spindle poles. The capture of a single astral MT by a kinetochore has been visualized directly in newt lung cultures ( Hayden et al., 1990 Rieder and Alexander, 1990), demonstrating the existence of this mechanism in vertebrate somatic cells. Selective stabilization of captured MTs results in the formation of the typical fusiform spindle in which the poles are focused on the centrosomes. When an astral MT encounters a specialized structure on the chromosome's primary constriction-the kinetochore-it is captured and stabilized ( Kirschner and Mitchison, 1986). In cells that contain centrosomes, these organelles generate arrays of highly dynamic astral microtubules (MTs), which explore the cytoplasm, searching for chromosomes. ![]() However, centrosomes facilitate the proper orientation of K-fibers toward spindle poles by integrating them into a common spindle.ĭepending on the organism, spindle assembly during mitosis can occur via two distinct pathways. Thus, even in the presence of centrosomes, the formation of some K-fibers is initiated by the kinetochores. Furthermore, when individual K-fibers are severed by laser microsurgery, they regrow from the kinetochore outward via MT plus-end polymerization at the kinetochore. This poleward transport results in chromosome bi-orientation and congression. Initially, kinetochore-formed K-fibers are not oriented toward a spindle pole but, as they grow, their minus ends are captured by astral microtubules (MTs) and transported poleward through a dynein-dependent mechanism. Here, we show that many of the kinetochore fibers (K-fibers) in centrosomal Drosophila S2 cells are formed by the kinetochores. The question remains, however, whether this pathway only activates when centrosome activity is compromised, or whether it contributes to spindle morphogenesis during a normal mitosis. It is now clear that a centrosome-independent pathway for mitotic spindle assembly exists even in cells that normally possess centrosomes.
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