Molecular Analysis of Microtubule Function in Yeast

描述(申请人提供)：微管是一种动态聚合物 在细胞器等各种细胞过程中发挥重要作用 运输和染色体分离。这项建议的长期目标是更好地了解微管的生物学作用。 动力学以及这些动态特性在 手机。 细胞内微管的动力学可能由复合体决定 与微管结合并调节其组装的蛋白质的相互作用。 酿酒酵母中的三种微管结合蛋白：Stu2p、Bik1p和 Bim1p，在所有成对组合中相互作用。这个 这些蛋白质的组合效应及其功能意义 它们在体内调节微管动力学中的蛋白质-蛋白质相互作用 将通过制造和分析特定的突变体来确定。此外， 这些蛋白质单独和组合对动力学性质的影响 将测定体外培养的微管的数量。第四种微管结合蛋白， 在纺锤体微管上发现了Stu1p，最近的发现表明它在 对着丝点的研究。建议进行实验以确定Stu1p是否 定位于动点，即细胞中动点微管的动力学 缺少Stu1p，以及Stu1p对动力学特性的内在影响 体外培养的微管。 尽管微管的动力学特性被认为是 它们的功能，细胞对微管动力学的依赖程度并没有 经过了实验测试。改变动态的β-微管蛋白突变 将构建酿酒酵母中微管的性质。的影响 这些突变对体内和微管上的微管动力学和功能的影响 体外动力学将被测量。这项工作的目的是了解如何 微管动力学性质的内在变化影响微管 细胞内的动力学和功能。

DESCRIPTION (provided by applicant): Microtubules are dynamic polymers that play essential roles in a variety of cellular processes such as organelle transport and chromosome segregation. The long-term objective of this proposal is to gain a better understanding of the biological role of microtubule dynamics and the ways in which these dynamic properties are regulated in the cell. Microtubule dynamics in cells is likely to be determined by the complex interplay of proteins that bind to microtubules and regulate their assembly. Three microtubule-binding proteins in the yeast S. cerevisiae, Stu2p, Bik1p and Bim1p, interact with each other in all pairwise combinations. The combinatorial effects of these proteins and the functional significance of their protein-protein interactions in regulating microtubule dynamics in vivo will be determined by making and analyzing specific mutants. In addition, the effects of these proteins, alone and in combination, on the dynamic properties of microtubules in vitro will be assayed. A fourth microtubule-binding protein, Stu1p, is found on spindle microtubules and recent findings indicate a role for Stu1p at the kinetochore. Experiments are proposed to determine if Stu1p localizes to kinetochores, the dynamics of kinetochore microtubules in cells lacking Stu1p, and the intrinsic effects of Stu1p on the dynamic properties of microtubules in vitro. Although the dynamic properties of microtubules are believed to be central to their function, the extent to which cells rely on microtubule dynamics has not been tested experimentally. Mutations in beta-tubulin that alter the dynamic properties of microtubules in S. cerevisiae will be constructed. The effect of these mutations on microtubule dynamics and function in vivo and on microtubule dynamics in vitro will be measured. The aim of this work is to understand how intrinsic changes in the dynamic properties of microtubules affect microtubule dynamics and function in the cell.