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Direct observation of microtubule dynamics at kinetochores in Xenopus extract spindles: implications for spindle mechanics.

Maddox P, Straight A, Coughlin P, Mitchison TJ, Salmon ED - J. Cell Biol. (2003)

Bottom Line: At anaphase onset, kinetochores switched to depolymerization of microtubule plus ends, resulting in chromosome-to-pole rates transiently greater than flux.Kinetochores switched from persistent depolymerization to persistent polymerization and back again during anaphase, bistability exhibited by kinetochores in vertebrate tissue cells.These results provide the most complete description of spindle microtubule poleward flux to date, with important implications for the microtubule-kinetochore interface and for how flux regulates kinetochore function.

View Article: PubMed Central - PubMed

Affiliation: Cell Division Group, Marine Biological Laboratory, Woods Hole, MA 02543, USA.

ABSTRACT
Microtubule plus ends dynamically attach to kinetochores on mitotic chromosomes. We directly imaged this dynamic interface using high resolution fluorescent speckle microscopy and direct labeling of kinetochores in Xenopus extract spindles. During metaphase, kinetochores were stationary and under tension while plus end polymerization and poleward microtubule flux (flux) occurred at velocities varying from 1.5-2.5 micro m/min. Because kinetochore microtubules polymerize at metaphase kinetochores, the primary source of kinetochore tension must be the spindle forces that produce flux and not a kinetochore-based mechanism. We infer that the kinetochore resists translocation of kinetochore microtubules through their attachment sites, and that the polymerization state of the kinetochore acts a "slip-clutch" mechanism that prevents detachment at high tension. At anaphase onset, kinetochores switched to depolymerization of microtubule plus ends, resulting in chromosome-to-pole rates transiently greater than flux. Kinetochores switched from persistent depolymerization to persistent polymerization and back again during anaphase, bistability exhibited by kinetochores in vertebrate tissue cells. These results provide the most complete description of spindle microtubule poleward flux to date, with important implications for the microtubule-kinetochore interface and for how flux regulates kinetochore function.

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Related in: MedlinePlus

Three models based on different contributions of kinetochore motility and kinetochore microtubule flux to metaphase kinetochore tension and anaphase A. For each model, only a single kinetochore microtubule (KMT) is shown with the polar minus end (−) at the left and the plus end (+) attached to the kinetochore on the right. Arrows indicate sites of polymerization or depolymerization. Metaphase includes sequential times, t1 and t2; anaphase onset occurs at t2 and continues for sequential times t2, t3, and t4. At metaphase, the centromeric linkage between sister kinetochores and polar ejection forces on the arms support tension at kinetochores (large blue arrow). Tension is lost at anaphase onset when sisters separate and the polar ejection forces on the arms are inactivated (Funabiki and Murray, 2000). See text for details.
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fig1: Three models based on different contributions of kinetochore motility and kinetochore microtubule flux to metaphase kinetochore tension and anaphase A. For each model, only a single kinetochore microtubule (KMT) is shown with the polar minus end (−) at the left and the plus end (+) attached to the kinetochore on the right. Arrows indicate sites of polymerization or depolymerization. Metaphase includes sequential times, t1 and t2; anaphase onset occurs at t2 and continues for sequential times t2, t3, and t4. At metaphase, the centromeric linkage between sister kinetochores and polar ejection forces on the arms support tension at kinetochores (large blue arrow). Tension is lost at anaphase onset when sisters separate and the polar ejection forces on the arms are inactivated (Funabiki and Murray, 2000). See text for details.

Mentions: In Xenopus and Drosophila spindles, where flux is fast, metaphase kinetochores do not exhibit directional instability (Desai et al., 1998; Maddox et al., 2002). Chromosomes in spermatocytes, oocytes, early embryos, and higher plants also do not exhibit kinetochore oscillations. Fig. 1 shows three possible models that could explain both the lack of metaphase chromosome oscillations and the mechanism of anaphase A seen in the systems just mentioned.


Direct observation of microtubule dynamics at kinetochores in Xenopus extract spindles: implications for spindle mechanics.

Maddox P, Straight A, Coughlin P, Mitchison TJ, Salmon ED - J. Cell Biol. (2003)

Three models based on different contributions of kinetochore motility and kinetochore microtubule flux to metaphase kinetochore tension and anaphase A. For each model, only a single kinetochore microtubule (KMT) is shown with the polar minus end (−) at the left and the plus end (+) attached to the kinetochore on the right. Arrows indicate sites of polymerization or depolymerization. Metaphase includes sequential times, t1 and t2; anaphase onset occurs at t2 and continues for sequential times t2, t3, and t4. At metaphase, the centromeric linkage between sister kinetochores and polar ejection forces on the arms support tension at kinetochores (large blue arrow). Tension is lost at anaphase onset when sisters separate and the polar ejection forces on the arms are inactivated (Funabiki and Murray, 2000). See text for details.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172681&req=5

fig1: Three models based on different contributions of kinetochore motility and kinetochore microtubule flux to metaphase kinetochore tension and anaphase A. For each model, only a single kinetochore microtubule (KMT) is shown with the polar minus end (−) at the left and the plus end (+) attached to the kinetochore on the right. Arrows indicate sites of polymerization or depolymerization. Metaphase includes sequential times, t1 and t2; anaphase onset occurs at t2 and continues for sequential times t2, t3, and t4. At metaphase, the centromeric linkage between sister kinetochores and polar ejection forces on the arms support tension at kinetochores (large blue arrow). Tension is lost at anaphase onset when sisters separate and the polar ejection forces on the arms are inactivated (Funabiki and Murray, 2000). See text for details.
Mentions: In Xenopus and Drosophila spindles, where flux is fast, metaphase kinetochores do not exhibit directional instability (Desai et al., 1998; Maddox et al., 2002). Chromosomes in spermatocytes, oocytes, early embryos, and higher plants also do not exhibit kinetochore oscillations. Fig. 1 shows three possible models that could explain both the lack of metaphase chromosome oscillations and the mechanism of anaphase A seen in the systems just mentioned.

Bottom Line: At anaphase onset, kinetochores switched to depolymerization of microtubule plus ends, resulting in chromosome-to-pole rates transiently greater than flux.Kinetochores switched from persistent depolymerization to persistent polymerization and back again during anaphase, bistability exhibited by kinetochores in vertebrate tissue cells.These results provide the most complete description of spindle microtubule poleward flux to date, with important implications for the microtubule-kinetochore interface and for how flux regulates kinetochore function.

View Article: PubMed Central - PubMed

Affiliation: Cell Division Group, Marine Biological Laboratory, Woods Hole, MA 02543, USA.

ABSTRACT
Microtubule plus ends dynamically attach to kinetochores on mitotic chromosomes. We directly imaged this dynamic interface using high resolution fluorescent speckle microscopy and direct labeling of kinetochores in Xenopus extract spindles. During metaphase, kinetochores were stationary and under tension while plus end polymerization and poleward microtubule flux (flux) occurred at velocities varying from 1.5-2.5 micro m/min. Because kinetochore microtubules polymerize at metaphase kinetochores, the primary source of kinetochore tension must be the spindle forces that produce flux and not a kinetochore-based mechanism. We infer that the kinetochore resists translocation of kinetochore microtubules through their attachment sites, and that the polymerization state of the kinetochore acts a "slip-clutch" mechanism that prevents detachment at high tension. At anaphase onset, kinetochores switched to depolymerization of microtubule plus ends, resulting in chromosome-to-pole rates transiently greater than flux. Kinetochores switched from persistent depolymerization to persistent polymerization and back again during anaphase, bistability exhibited by kinetochores in vertebrate tissue cells. These results provide the most complete description of spindle microtubule poleward flux to date, with important implications for the microtubule-kinetochore interface and for how flux regulates kinetochore function.

Show MeSH
Related in: MedlinePlus