Molecular analysis of microtubule function in yeast

微管在包括染色体在内的各种细胞过程中扮演着重要的角色。 分离和细胞器运输。它们是动态的聚合物，经常交替 在生长和收缩阶段之间，一种允许快速重新安排的属性 微管阵列对细胞信号的响应。微管动力学被调节，在 很大一部分是由一组被称为正端跟踪蛋白或+TIPS的蛋白质组成的。 目前，已经在真核生物中鉴定出大约12个保守的+TIP家族 酵母对人类的影响，但这些蛋白质中的大多数影响微管的机制 动力学还没有被很好地理解。除了结合微管的能力外，大多数+TIPS 与其他几个+TIP互动，创建+TIP互动的复杂网络。这些 互动很可能是+TIP活动的关键调节因素，也是这一活动的主要焦点 申请。我们建议研究在调节中发挥突出作用的四个+提示 酿酒酵母中微管的动态变化。这些蛋白是Bim1、Bik1、Stu1和Stu2 分别属于+TIP家族EB、CLIP、CLASP和XMAP215。我们的目标是 纯化这些蛋白质并确定它们如何影响体外培养的微管动力学 微管组装系统。然后我们将结合成对的蛋白质来研究 特殊的+末端相互作用影响它们调节微管动力学的能力。我们有 在Bim1和Bik1的初步研究中开发了所有必要的分析方法。我们现在 计划通过包括Stu1和Stu2来扩展这些研究。

Microtubules play essential roles in a variety of cellular processes, including chromosome segregation and organelle transport. They are dynamic polymers that alternate frequently between phases of growing and shrinking, a property that allows the rapid rearrangement of microtubule arrays in response to cellular signals. Microtubule dynamics are regulated, in large part, by a group of proteins referred to as plus-end tracking proteins or +TIPs. Currently, about a dozen conserved +TIP families have been identified in eukaryotes from yeast to human, but the mechanisms by which most of these proteins influence microtubule dynamics are not well understood. In addition to their ability to bind microtubules, most +TIPs interact with several other +TIPs, creating a complex web of +TIP interactions. These interactions are likely to be key regulators of +TIP activities and are the primary focus of this application. We propose to study four +TIPs that play prominent roles in regulating microtubule dynamics in the yeast, S. cerevisiae. These proteins, Bim1, Bik1, Stu1 and Stu2 belong to the +TIP families EB, CLIP, CLASP, and XMAP215, respectively. Our goal is to purify these proteins and determine how they influence microtubule dynamics in an in vitro microtubule assembly system. Then we will combine pairs of proteins to examine how particular +TIP interactions affect their abilities to regulate microtubule dynamics. We have developed all of the necessary assays in preliminary studies with Bim1 and Bik1. We now plan to expand these studies by including Stu1 and Stu2.