Regulation of Microtubule Dynamics and Organization During Cell Division

精确的细胞分裂对于生命至关重要。这个过程中的错误会导致许多疾病，包括癌症。有丝分裂纺锤体是移动染色体的引擎，因此它在确保新生细胞继承完整的遗传蓝图方面发挥着至关重要的作用。纺锤体的一个关键特征是它的双极性，这是一种确保双向染色体运动的几何形状。非双极几何形状，例如单极或多极，会导致与生命不相容的染色体分离错误。因此，在有丝分裂后期阶段开始之前，必须将非双极结构重组为双极结构。纺锤体形成是一个复杂的过程，由许多微管相关蛋白（MAP）和分子马达（即驱动蛋白和动力蛋白）介导。纺锤体组装的机制在生物体之间可能有所不同，即使在单一生物体内也可以表现出显着的可塑性。纺锤体组装的可塑性源自 MAP 和运动蛋白如何直接或通过动态 MT 网络间接相互作用。尽管进行了数十年的研究，但这些系统级关系的复杂性使得人们很难完全理解驱动纺锤体形成的机制。我们独特的方法是分离和表征真核生物中在缺乏由驱动蛋白 Eg5 驱动的主要纺锤体组装途径的情况下存活的人类细胞系。我们的工作告诉我们，人类细胞可以使用辅助途径组装适当的有丝分裂纺锤体。此外，我们还发现了一种对于这种替代机制至关重要的驱动蛋白 (Kif15)。我们实验室正在进行的工作揭示了细胞中需要 Kif15 进行细胞分裂的系统级变化，这促使人们努力更好地了解主轴电机如何在主轴内的系统级发挥作用。在本次更新申请中，我们将解决两个关键问题：1）Kif15如何驱动主轴组件？ 2) 细胞分裂过程中 Kif15 活性如何受到调节？这项工作将增进我们对纺锤体力学的理解，并与抗有丝分裂化疗策略的发展直接相关。

Precise cell division is essential for life. Mistakes during this process lead to many diseases, including cancer. The mitotic spindle is the engine that moves chromosomes, and it therefore plays an essential role in ensuring that newly-born cells inherit a complete genetic blueprint. A key feature of the spindle is its bipolarity, a geometry that ensures bi-directional chromosome movements. Non-bipolar geometries, such as monopolarity or multipolarity, cause errors in chromosome segregation that are incompatible with life. Therefore, non-bipolar structures must be reorganized to be bipolar prior to the onset of the anaphase stage of mitosis. Spindle formation is a complex process that is mediated by many microtubule-associated proteins (MAPs) and molecular motors (i.e., kinesins and dynein). The mechanisms of spindle assembly can vary among organisms, and can show remarkable plasticity even within a single organism. The malleability of spindle assembly is derived from how MAPs and motor proteins engage each other, either directly or indirectly through a network of dynamic MTs. The sophisticated nature of these systems-level relationships has made it difficult to fully understand the mechanisms that drive spindle formation, despite decades of research. Our unique approach has been to isolate and characterize human cell lines that survive in the absence of the major spindle assembly pathway driven by the kinesin Eg5 in eukaryotes. Our work has taught us that human cells can assemble a proper mitotic spindle using an auxiliary pathway. Furthermore, we identified a kinesin (Kif15) that is essential for this alternate mechanism. Ongoing work in our lab has unveiled systems-level changes in cells that require Kif15 for cell division, motivating efforts to obtain a better understanding of how spindle motors function at a systems level within the spindle. In this renewal application, we will address two key questions: 1) How does Kif15 drive spindle assembly?; and 2) How is Kif15 activity regulated during cell division? This work will advance our understanding of spindle mechanics and have immediate relevance to the development of anti-mitotic chemotherapeutic strategies.