Time-lapse video microscopy analysis reveals astral microtubule detachment in the yeast spindle pole mutant cnm67
Time-lapse video microscopy analysis reveals astral microtubule detachment in the yeast spindle pole mutant cnm67
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DOI：
10.1091/mbc.11.4.1197
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发表时间：
2000-04-01
影响因子：
3.3
通讯作者：
Philippsen, P
中科院分区：
文献类型：
--
作者：
Hoepfner, D;Brachat, A;Philippsen, P
酿酒酵母cnm 67 δ细胞缺乏纺锤体极体（SPB）外斑，星状（细胞质）微管的主要附着位点，导致频繁的核分离失败。我们监测动态的绿色荧光蛋白标记的细胞核和微管在几个细胞周期。早期的核迁移步骤，如核定位和纺锤体的方向略有影响，但后期阶段，如快速振荡和插入的后期核到芽颈大多缺席。微管动力学分析显示，正常的行为，核纺锤体，但频繁的脱离后SPB分离的星形微管。同时，Spc 72蛋白，γ-微管蛋白复合物的细胞质锚，从SPB区域部分丢失，其动力学与微管观察到的相似。我们假设，在cnm 67三角洲细胞Spc 72-γ-微管蛋白复合帽的星形微管被释放的半桥SPB分离后，但未能被锚定到SPB的细胞质侧，因为没有一个外部斑块。然而，cnm 67 δ细胞中成功的核分离仍然可以通过Kip 3/Kar 3马达的作用在星形微管脱离之前正确定向的纺锤体的伸长力来实现。有趣的是，新生二倍体细胞的第一次核分离从未失败，即使发生星状微管脱离。
Saccharomyces cerevisiae cnm67 Delta cells lack the spindle pole body (SPB) outer plaque, the main attachment site for astral (cytoplasmic) microtubules, leading to frequent nuclear segregation failure. We monitored dynamics of green fluorescent protein-labeled nuclei and microtubules over several cell cycles. Early nuclear migration steps such as nuclear positioning and spindle orientation were slightly affected, but late phases such as rapid oscillations and insertion of the anaphase nucleus into the bud neck were mostly absent. Analyzes of microtubule dynamics revealed normal behavior of the nuclear spindle but frequent detachment of astral microtubules after SPB separation. Concomitantly, Spc72 protein, the cytoplasmic anchor for the gamma-tubulin complex, was partially lost from the SPB region with dynamics similar to those observed for microtubules. We postulate that in cnm67 Delta cells Spc72-gamma-tubulin complex-capped astral microtubules are released from the half-bridge upon SPB separation but fail to be anchored to the cytoplasmic side of the SPB because of the absence of an outer plaque. However, successful nuclear segregation in cnm67 Delta cells can still be achieved by elongation forces of spindles that were correctly oriented before astral microtubule detachment by action of Kip3/Kar3 motors. Interestingly, the first nuclear segregation in newborn diploid cells never fails, even though astral microtubule detachment occurs.