Deformations Within Moving Kinetochores Reveal Different Sites of Active and Passive Force Generation

Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA.
Science (Impact Factor: 33.61). 06/2012; 337(6092):355-8. DOI: 10.1126/science.1221886
Source: PubMed


Kinetochores mediate chromosome segregation at mitosis. They are thought to contain both active, force-producing and passive,
frictional interfaces with microtubules whose relative locations have been unclear. We inferred mechanical deformation within
single kinetochores during metaphase oscillations by measuring average separations between fluorescently labeled kinetochore
subunits in living cells undergoing mitosis. Inter-subunit distances were shorter in kinetochores moving toward poles than
in those moving away. Inter-subunit separation decreased abruptly when kinetochores switched to poleward movement and decreased
further when pulling force increased, suggesting that active force generation during poleward movement compresses kinetochores.
The data revealed an active force-generating interface within kinetochores and a separate passive frictional interface located
at least 20 nanometers away poleward. Together, these interfaces allow persistent attachment with intermittent active force

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    • "Both the model and the experimental data also show that the AP-moving KT reaches its maximum speed earlier than the P-moving KT (Fig. 5, I and J; and Fig. 1, E and F, insets; Wan et al., 2012). In fact, these differences in acquisition of the maximal speed during P and AP movement are responsible for the observed period doubling in the inter-KT distance (Dumont et al., 2012; Wan et al., 2012; Fig. 5 A). Our model suggests that this is due to the biphasic kinetics of the Ndc80 bonds attached to depolymerizing MTs, resulting in a slower turnover of the kMT bonds of the P-moving sister (under moderate tension), hindering the increase in P velocity until a critical tension level is reached. "
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    ABSTRACT: Duplicated mitotic chromosomes aligned at the metaphase plate maintain dynamic attachments to spindle microtubules via their kinetochores, and multiple motor and nonmotor proteins cooperate to regulate their behavior. Depending on the system, sister chromatids may display either of two distinct behaviors, namely (1) the presence or (2) the absence of oscillations about the metaphase plate. Significantly, in PtK1 cells, in which chromosome behavior appears to be dependent on the position along the metaphase plate, both types of behavior are observed within the same spindle, but how and why these distinct behaviors are manifested is unclear. Here, we developed a new quantitative model to describe metaphase chromosome dynamics via kinetochore-microtubule interactions mediated by nonmotor viscoelastic linkages. Our model reproduces all the key features of metaphase sister kinetochore dynamics in PtK1 cells and suggests that differences in the distribution of polar ejection forces at the periphery and in the middle of PtK1 cell spindles underlie the observed dichotomy of chromosome behavior.
    The Journal of Cell Biology 05/2013; 201(4):577-93. DOI:10.1083/jcb.201301022 · 9.83 Impact Factor
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    • "The observed structural changes at kinetochores have been assumed to correlate with the presence of tension, but thus far, such studies have not made direct measurements of force or tension. Nevertheless, careful quantitative analysis of the dynamic changes in the distances between CENP-C and Hec1 or Cdc20 during sister chromatid oscillations has supported the model that changes in intrakinetochore distance are force dependent (Dumont et al., 2012). However, these structural alterations may also be the result of changes in the conformation, organization, or localization of proteins within the kinetochore. "
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    ABSTRACT: Chromosome segregation requires the generation of force at the kinetochore-the multiprotein structure that facilitates attachment of chromosomes to spindle microtubules. This force is required both to move chromosomes and to signal the formation of proper bioriented attachments. To understand the role of force in these processes, it is critical to define how force is generated at kinetochores, the contributions of this force to chromosome movement, and how the kinetochore is structured and organized to withstand and respond to force. Classical studies and recent work provide a framework to dissect the mechanisms, functions, and consequences of force at kinetochores.
    The Journal of Cell Biology 03/2013; 200(5):557-65. DOI:10.1083/jcb.201211113 · 9.83 Impact Factor
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    ABSTRACT: A molecular motor is made of either a single macromolecule or a macromolecular complex. Just like their macroscopic counterparts, molecular motors "transduce" input energy into mechanical work. All the nano-motors considered here operate under isothermal conditions far from equilibrium. Moreover, one of the possible mechanisms of energy transduction, called Brownian ratchet, does not even have any macroscopic counterpart. But, molecular motor is not synonymous with Brownian ratchet; a large number of molecular motors execute a noisy power stroke, rather than operating as Brownian ratchet. We review not only the structural design and stochastic kinetics of individual single motors, but also their coordination, cooperation and competition as well as the assembly of multi-module motors in various intracellular kinetic processes. Although all the motors considered here execute mechanical movements, efficiency and power output are not necessarily good measures of performance of some motors. Among the intracellular nano-motors, we consider the porters, sliders and rowers, pistons and hooks, exporters, importers, packers and movers as well as those that also synthesize, manipulate and degrade "macromolecules of life". We review mostly the quantitative models for the kinetics of these motors. We also describe several of those motor-driven intracellular stochastic processes for which quantitative models are yet to be developed. In part I, we discuss mainly the methodology and the generic models of various important classes of molecular motors. In part II, we review many specific examples emphasizing the unity of the basic mechanisms as well as diversity of operations arising from the differences in their detailed structure and kinetics. Multi-disciplinary research is presented here from the perspective of physicists.
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