Mechanisms of microtubule-based kinetochore positioning in the yeast metaphase spindle
Mechanisms of microtubule-based kinetochore positioning in the yeast metaphase spindle
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DOI：
10.1016/s0006-3495(03)75087-5
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发表时间：
2003-06-01
影响因子：
3.4
通讯作者：
Odde, DJ
中科院分区：
文献类型：
--
作者：
Sprague, BL;Pearson, CG;Odde, DJ
据推测，kMT动态不稳定性的空间梯度促进有丝分裂纺锤体的形成和染色体的运动。为了验证这一假设，需要分析kMT动力学，这还没有解决在活细胞中的单个kMT水平。芽殖酵母纺锤体提供了一个有吸引力的系统，在其中研究kMT动力学，因为与动物细胞相比，每个动粒只有一个kMT。为了可视化酵母中的中期kMT加末端动力学，通过荧光显微镜对含有与动粒蛋白Cse 4融合的绿色荧光蛋白的菌株进行成像。虽然个别kinetochores是不可分辨的，我们发现，模型的kMT动力学可以通过模拟随机kMT动力学，然后模拟荧光成像的kMT加上端相关的kinetochores进行评估。模型预测的图像，实验观察到的图像的统计比较表明，一个纯粹的动态不稳定模型kMT动态在酵母中期纺锤体是不可接受的。然而，当灾难或救援频率的时间稳定的空间梯度被添加到模型中时，模型和实验之间存在合理的一致性。这些结果提供了第一个证据的时间稳定的空间梯度的kMT灾难和/或救援频率在活细胞。
It has been hypothesized that spatial gradients in kMT dynamic instability facilitate mitotic spindle formation and chromosome movement. To test this hypothesis requires the analysis of kMT dynamics, which have not been resolved at the single kMT level in living cells. The budding yeast spindle offers an attractive system in which to study kMT dynamics because, in contrast to animal cells, there is only one kMT per kinetochore. To visualize metaphase kMT plus-end dynamics in yeast, a strain containing a green fluorescent protein fusion to the kinetochore protein, Cse4, was imaged by fluorescence microscopy. Although individual kinetochores were not resolvable, we found that models of kMT dynamics could be evaluated by simulating the stochastic kMT dynamics and then simulating the fluorescence imaging of kMT plus-end-associated kinetochores. Statistical comparison of model-predicted images to experimentally observed images demonstrated that a pure dynamic instability model for kMT dynamics in the yeast metaphase spindle was unacceptable. However, when a temporally stable spatial gradient in the catastrophe or rescue frequency was added to the model, there was reasonable agreement between the model and the experiment. These results provide the first evidence of temporally stable spatial gradients of kMT catastrophe and/or rescue frequency in living cells.