Microtubule dynamics and kinetochore function

Chromosome alignment during mitosis is frequently accompanied by a dynamic switching between elongation and shortening of kinetochore fibers (K-fibers) that connect kinetochores and spindle poles [1, 2]. In higher eukaryotes, mature K-fibers consist of 10–30 kinetochore microtubules (kMTs) whose plus-ends are embedded in the kinetochore [1-3]. A critical and long-standing question is how the dynamics of individual kMTs within the K-fiber are coordinated [1-5]. We have addressed this question by using electron tomography to determine the polymerization/depolymerization status of individual kMTs in the K-fibers of PtK1 and Drosophila S2 cells. Surprisingly, we find that the plus-ends of two-thirds of kMTs are in a depolymerizing state, even when the K-fiber exhibits net tubulin incorporation at the plus end [6-8]. Furthermore, almost all individual K-fibers examined had a mixture of kMTs in the polymerizing and depolymerizing states. Therefore, although K-fibers elongate and shrink as a unit, the dynamics of individual kMTs within a K-fiber are not coordinated at any given moment. Our results suggest a novel control mechanism through which attachment to the kinetochore outer plate prevents shrinkage of kMTs. We discuss the ramifications of this new model on the regulation of chromosome movement and the stability of K-fibers

The Drosophila kinetochore

Drosophila melanogaster is a widely used model organism for the molecular dissection of mitosis in animals. However, despite the popularity of this system, no studies have been published on the ultrastructure of Drosophila kinetochores and kinetochore fibers (K-fibers) in somatic cells. To amend this situation, we used correlative light (LM) and electron microscopy (EM) to study kinetochores in cultured Drosophila S2 cells during metaphase, and after colchicine treatment to depolymerize all microtubules (MTs). We find that the structure of attached kinetochores in S2 cells is indistinct, consisting of an amorphous inner zone associated with a more electron-dense peripheral surface layer that is approximately 40-50 nm thick. On average, each S2 kinetochore binds 11+/-2 MTs, in contrast to the 4-6 MTs per kinetochore reported for Drosophila spermatocytes. Importantly, nearly all of the kinetochore MT plus ends terminate in the peripheral surface layer, which we argue is analogous to the outer plate in vertebrate kinetochores. Our structural observations provide important data for assessing the results of RNAi studies of mitosis, as well as for the development of mathematical modelling and computer simulation studies in Drosophila and related organisms

The coupling between sister kinetochore directional instability and oscillations in centromere stretch in metaphase PtK1 cells

High-resolution kinetochore tracking reveals that oscillations in centromere stretch occur at twice the frequency of kinetochore oscillations because of the nonlinear kinetics of kinetochore directional instability, which produce poleward and away-from-the-pole kinetochore oscillations

Finding the middle ground: how kinetochores power chromosome congression

ISSN:1420-682XISSN:1420-907