Kinetochore alignment within the metaphase plate is regulated by centromere stiffness and microtubule depolymerases
Video 1:Time-lapse sequence of an unperturbed (WT) EGFP–CENP-A HeLa cell in late prometaphase. (2.229Mb)
Video 3: Plane fit through aligned kinetochores and identification of unaligned kinetochores in the time-lapse sequence of Video 1. (1.563Mb)
Video 5: Identification of sister kinetochore pairs in the time-lapse sequence of Video 1. (2.241Mb)
King, Emma M.
Amaro, Ana C.
Winter, Jennifer R.
Dorn, Jonas F.
Elliott, Hunter L.
McClelland, Sarah E.
Porter, Iain M.
McAinsh, Andrew D.
Swedlow, Jason R.
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During mitosis in most eukaryotic cells, chromosomes align and form a metaphase plate halfway between the spindle poles, about which they exhibit oscillatory movement. These movements are accompanied by changes in the distance between sister kinetochores, commonly referred to as breathing. We developed a live cell imaging assay combined with computational image analysis to quantify the properties and dynamics of sister kinetochores in three dimensions. We show that baseline oscillation and breathing speeds in late prometaphase and metaphase are set by microtubule depolymerases, whereas oscillation and breathing periods depend on the stiffness of the mechanical linkage between sisters. Metaphase plates become thinner as cells progress toward anaphase as a result of reduced oscillation speed at a relatively constant oscillation period. The progressive slowdown of oscillation speed and its coupling to plate thickness depend nonlinearly on the stiffness of the mechanical linkage between sisters. We propose that metaphase plate formation and thinning require tight control of the state of the mechanical linkage between sisters mediated by centromeric chromatin and cohesion.
© The Authors, 2010. This article is distributed under the terms of the Creative Commons Attribution-Noncommercial-Share Alike 3.0 Unported License. The definitive version was published in Journal of Cell Biology 188 (2010): 665-679, doi:10.1083/jcb.200909005.
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