Whitehead C M, Winkfein R J, Rattner J B
Department of Anatomy, University of Calgary, Alberta, Canada.
Cell Motil Cytoskeleton. 1996;35(4):298-308. doi: 10.1002/(SICI)1097-0169(1996)35:4<298::AID-CM3>3.0.CO;2-3.
Although centrosome separation is essential to the formation of a bipolar spindle, it can proceed along several different pathways. This raises questions as to the similarity between the mechanism(s) underlying these various forms of separation. To address this question we reinvestigated centrosome separation in HeLa cells using a variety of techniques. We present a refined description of the two major pathways of centrosome separation found in HeLa cells and demonstrate that each of these pathways has its own timing, protein requirements, morphological characteristics, and relationship to spindle assembly. The first pathway, which occurs in prophase cells, is dependent on an intact actin cytoskeleton, and when this pathway is completed prior to nuclear envelope breakdown, the microtubules associated with this process do not become part of the spindle. Thus, centrosome separation and spindle pole organization can occur as two separate events. The second centrosome separation pathway is found in cells in which separation occurs concurrent with prometaphase. In this case, centrosome separation and the formation of the mitotic spindle are integrated together and an intact actin cytoskeleton is not required. The relationship between these multiple pathways of centrosome separation and the distribution of the human kinesin-like protein HsEg5 was also investigated. This protein was found associated with all centrosomal microtubules present during both prophase and prometaphase centrosome separation, as well as with prophase centrosomes displaying independent movement in Cytochalasin-D treated cells. In addition, we demonstrate that this protein is associated with post-mitotic centrosome movement which involves a single centrosome. Thus, HsEg5 is a feature of individual centrosome function and does not require anti-parallel microtubule arrays.
尽管中心体分离对于双极纺锤体的形成至关重要，但它可以通过几种不同的途径进行。这就引发了关于这些不同形式分离背后机制之间相似性的问题。为了解决这个问题，我们使用多种技术对HeLa细胞中的中心体分离进行了重新研究。我们对HeLa细胞中发现的中心体分离的两种主要途径进行了精确描述，并证明这些途径中的每一种都有其自身的时间安排、蛋白质需求、形态特征以及与纺锤体组装的关系。第一种途径发生在前期细胞中，依赖于完整的肌动蛋白细胞骨架，并且当该途径在核膜破裂之前完成时，与该过程相关的微管不会成为纺锤体的一部分。因此，中心体分离和纺锤体极组织可以作为两个独立的事件发生。第二种中心体分离途径存在于分离与前中期同时发生的细胞中。在这种情况下，中心体分离和有丝分裂纺锤体的形成是整合在一起的，并且不需要完整的肌动蛋白细胞骨架。我们还研究了中心体分离的这些多种途径与人类驱动蛋白样蛋白HsEg5分布之间的关系。发现该蛋白与前期和前中期中心体分离期间存在的所有中心体微管相关，以及与在细胞松弛素-D处理的细胞中显示独立运动的前期中心体相关。此外，我们证明该蛋白与涉及单个中心体的有丝分裂后中心体运动相关。因此，HsEg5是单个中心体功能的一个特征，并且不需要反平行微管阵列。
