Quantitative Analysis and Modeling of Microtubule Structure and Regulation

描述(申请人提供)：癌症的一个标志是不受控制的细胞分裂，许多癌症的治疗措施通过破坏微管动力学和/或有丝分裂纺锤体结构来阻止细胞分裂。显然，适当的微管组织和长度调节对于成功完成细胞分裂是重要的。在这项研究中，我们将重点研究微管相关蛋白如何在有丝分裂过程中调节微管长度和组织纺锤体微管的机制。为了为这些和未来的研究提供基础，我们首先将重点放在缺乏调节蛋白的情况下的微管动力学，以更好地了解微管尖端结构和微管动力学之间的关系。我们的初步模拟结果预测，微管尖端结构对于微管动力学的自我调节是重要的。我们将使用体外荧光和电子显微镜以及在萌芽酵母中的体内分析来研究这一假说。然后，我们将研究重要的微管+末端结合蛋白的末端跟踪机制，如Eb1，以确定微管末端结构是否可以提供一个平台，将重要的长度调节蛋白靶向微管末端。最后，我们将研究微管相关蛋白在细胞分裂过程中将微管组织成有丝分裂纺锤体的作用。具体地说，我们将使用发芽中的Yas细胞的体内实验、体外重建和计算机模拟来确定Kinesin-14负端定向马达在有丝分裂纺锤体中建立适当的微管束的作用和机制。

DESCRIPTION (provided by applicant): A hallmark of cancer is uncontrolled cell division, and many cancers therapeutic measures work by disrupting microtubule dynamics and/or mitotic spindle architecture to arrest cell division. It is clear that proper microtubule organization and length regulation are important for the successful completion of cell division. In this study, we will focus on mechanisms for how microtubule-associated proteins regulate microtubule lengths and organize spindle microtubules during mitosis. To provide a basis for these and future studies, we will first focus on microtubule dynamics in the absence of regulating proteins to better understand the relationship between microtubule tip structures and microtubule dynamics. Our preliminary modeling results predict that microtubule tip structures are important for self-regulating microtubule dynamics. We will investigate this hypothesis using in-vitro fluorescence and electron microscopy, as well as in-vivo analysis in budding yeast. Then, we will investigate the mechanism for tip-tracking of important microtubule plus end-binding proteins, such as Eb1, to determine whether the microtubule tip structures could provide a platform for targeting of important length-regulating proteins to the microtubule tip. Finally, we will investigate the role of microtubule- associated proteins in organizing microtubules into a mitotic spindle during cell division. Specifically, we will use in-vivo experiments in budding yeas cells, in-vitro reconstitution, and computer simulations to determine the role and mechanism of Kinesin-14 minus-end directed motors in establishing proper microtubule bundling in mitotic spindles.