Regulation of Microtubule Dynamics and Organization During Cell Division

精确的细胞分裂对生命至关重要。在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重要作用。纺锤体的一个关键特征是它的双极性，这是一种确保染色体双向运动的几何形状。非双极几何形状，如单极或多极，导致染色体分离错误，与生命不相容。因此，非双极结构必须在有丝分裂后期开始之前重组为双极。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）。纺锤体组装的机制在不同的生物体中是不同的，甚至在同一个生物体中也表现出显著的可塑性。纺锤体组装的延展性来自于MAP和马达蛋白如何通过动态MT网络直接或间接地相互接合。尽管经过了数十年的研究，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。我们独特的方法一直是分离和鉴定人类细胞系，这些细胞系在真核生物中由驱动蛋白Eg5驱动的主要纺锤体组装途径不存在的情况下存活。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，我们确定了驱动蛋白（Kif15），这是必不可少的替代机制。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纺锤体马达在纺锤体内系统级的功能。在这次更新申请中，我们将解决两个关键问题：1）Kif15如何驱动主轴组件？以及2）Kif15活性在细胞分裂期间是如何调节的？这项工作将推进我们的纺锤体力学的理解，并直接相关的发展，抗有丝分裂化疗策略。

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.