Regulation of Microtubule Dynamics and Organization During Cell Division

 描述(申请人提供)：有丝分裂是复制的一组染色体在两个子细胞中平均分布的过程。因为细胞需要完整的基因蓝图才能正常运作，所以有丝分裂必须没有错误--即使是一条染色体的错误分离也可能导致遗传性疾病、癌症和死亡。染色体由一种叫做纺锤体的基于微管的细胞机分离。纺锤体的一个关键特征是它的两极几何结构，这种结构自然允许染色体在两个方向上分离。其他组织状态，如独占或多极，在很大程度上与生命不相容，并通过细胞凋亡导致细胞死亡。这项续订申请的重点是如何建立和维持纺锤体两极。大多数真核细胞中的纺锤体两极是由运动蛋白-5马达EG5建立的。然而，我们在之前的资助周期中发现，人类癌细胞中的双极性可以通过一种新的EG5依赖的机制来建立。这一发现具有重要的临床意义，因为在早期临床试验中，Kinesin-5抑制剂(K5I)作为抗癌药物表现不佳。我们的工作表明，K5I的无效可能源于替代的代偿性纺锤体组装途径。一旦纺锤体形成，它必须保持双极状态，尽管存在与EG5相反的力量。EG5反/辅力的来源、大小和时间波动尚未得到很好的描述，但我们的数据暗示着动粒附着型微管(K-MT)的参与。在这项资助中，我们将：1)确定K-MT何时以及如何有助于维持双极性；2)研究第二Kinesin(Kif15)对EG5独立的主轴组装的关键因素的特征；以及3)进一步表征非规范的主轴组装机制及其生理缺陷。这项工作将促进我们对纺锤体机制的理解，并对开发抗有丝分裂化疗策略具有直接的意义。

 DESCRIPTION (provided by applicant): Mitosis is the process by which a replicated set of chromosomes is equally distributed between two daughter cells. Because cells require a complete genetic blueprint to function properly, it is essential for mitosis to occur without error mis-segregation of even a single chromosome can be the cause of genetic disease, cancer, and death. Chromosomes are segregated by a microtubule-based cellular machine termed the spindle. A key feature of the spindle is its bipolar geometry, an organization that naturally allow chromosomes to be segregated in two directions. Other organizational states, such as monopolarity or multipolarity, are largely incompatible with life and cause cell death via apoptosis. This renewal application is focused on how spindle bipolarity is both established and maintained. Spindle bipolarity in most eukaryotic cells is established by Eg5, a kinesin-5 motor. However, we discovered in the previous grant cycle that bipolarity in human cancer cells can be established through a novel Eg5-indpendent mechanism. This finding has important clinical implications, as kinesin-5 inhibitors (K5Is) have not performed well as anti-cancer agents in early stage clinical trials. Our work suggests that the inefficacy of K5Is may stem from alternative, compensatory spindle assembly pathways. Once the spindle has formed, it must remain bipolar despite the presence of forces that act in opposition to Eg5. The source, magnitude and temporal fluctuations of Eg5-opposing/assisting forces are not well-characterized, but our data implicate the involvement of kinetochore-attached microtubules (K- MTs). In this grant, we will: 1) Determine when and how K-MTs contribute to bipolarity maintenance; 2) Study features of a second kinesin (Kif15) key for Eg5-independent spindle assembly; and 3) Further characterize non-canonical spindle assembly mechanisms and their physiological shortcomings. This work will advance our understanding of spindle mechanics and have immediate relevance to the development of anti-mitotic chemotherapeutic strategies.