Mechanisms of spindle formation

描述（由申请人提供）：有丝分裂是细胞生命中的关键阶段。这是一个双极纺锤体结构被组织起来，将复制的染色体分离到两个子细胞中的阶段。主轴的组织和功能要求极高的精度、坚固性和保真度。与纺锤体相关的缺陷可导致染色体分离缺陷或非整倍性，这与某些类型的癌症相关。纺锤体是由微管、微管相关蛋白、分子马达和其他调节蛋白组成的大分子机器。最令人感兴趣的是分子马达，它执行诸如交联和滑动微管以形成双极纺锤体，或将微管分离以保持适当的纺锤体长度，或将染色体携带到相反的纺锤体两极等工作。令人惊讶的是，虽然我们已经了解了很多关于有丝分裂的马达，但我们仍然对MAP和其他调节蛋白以及它们如何与马达协调以实现正确的纺锤体形成知之甚少。我的实验室使用相对简单的裂殖酵母裂殖酵母和人类培养细胞来解决纺锤体组织和功能的保守机制。这个特殊的项目集中在如何形成的初始双极纺锤体在有丝分裂开始，阶段称为前期。我们专注于有助于纺锤体形成的地图。使用裂变酵母作为基因发现工具，我们已经开始定义一个新基因的作用，我们称之为psr1+（两极分离调节因子1）。我们的工作表明，psr1p组织的初始双极主轴期间， 前期Psr1缺失导致高频率的单极纺锤体和随后的染色体分离缺陷。裂变酵母psr1+似乎有人类功能同源物。我们已经开始描述一种新的人类基因，我们称之为PSR1。在HeLa细胞中，PSR1的siRNA也导致高频率的单极纺锤体和随后的染色体分离缺陷。本项目旨在结合联合收割机现代细胞和分子生物学技术在分裂酵母和人类培养细胞、生物化学、高分辨率光学活细胞成像和创新的微流体技术来控制细胞微环境，以达到双极纺锤体形成的机制理解。 公共卫生相关性：细胞分裂是细胞生命的关键阶段，遗传信息（以染色体的形式）被复制并平均分配到子细胞中。细胞分裂的缺陷会导致癌症。染色体分离是由一种叫做纺锤体的结构完成的。纺锤体是由微管、马达、微管相关蛋白（MAP）和其他调节蛋白组成的大分子机器。我们建议研究保守的纺锤体组织和功能的遗传学上易于处理的模型系统裂殖酵母和人类培养细胞的机制。

DESCRIPTION (provided by applicant): Mitosis is a key stage during the life of a cell. It is the stage where a bipolar spindle structure is organized to segregate duplicated chromosomes into the two daughter cells. Spindle organization and function require exquisite precision, robustness and fidelity. Defects associated with the spindle can lead to defects in chromosomal segregation, or aneuploidy, which has been correlated with some types of cancer. The spindle is a macromolecular machine made of microtubules, microtubule-associated proteins (MAPs), molecular motors and other regulatory proteins. Of intense interest have been molecular motors, which perform work such as cross-linking and sliding microtubules apart to form the bipolar spindle, or to depolymerize microtubules to maintain proper spindle lengths, or to carry chromosomes to opposite spindle poles. Surprisingly, while we have learned much about motors involved in mitosis, we still know very little about the MAPs and other regulatory proteins and how they coordinate with motors to bring about proper spindle formation. My laboratory uses the relatively simple fission yeast Schizosaccharomyces pombe and human cultured cells to address conserved mechanisms of spindle organization and function. This particular project focuses on how the initial bipolar spindle is formed at the start of mitosis, the stage termed prophase. We focus on the MAPs that contribute to spindle formation. Using fission yeast as a gene discovery tool, we have begun to define the roles of a new gene we called psr1+ (poles separation regulator 1). Our work indicates that psr1p organizes the initial bipolar spindle during prophase. Psr1-deletion leads to high frequency of monopolar spindles and subsequent chromosome segregation defects. Fission yeast psr1+ appears to have a human functional homolog. We have begun to characterize a novel human gene we called PSR1. In HeLa cells, siRNA of PSR1 also leads to high frequency of monopolar spindles and subsequent chromosome segregation defects. This proposal aims to combine modern cell and molecular biology techniques in fission yeast and human cultured cells, biochemistry, high-resolution optical live-cell imaging, and innovative microfluidic techniques to control cellular microenvironment, to reach a mechanistic understanding of bipolar spindle formation. PUBLIC HEALTH RELEVANCE: Cell division is a key stage in the life of a cell, where genetic information (in the form of chromosomes) is duplicated and partitioned equally into the daughter cells. Defects in cell division can lead to cancer. Chromosome segregation is accomplished by a structure call the mitotic spindle. The spindle is a macromolecular machine composed of microtubules, motors, microtubule-associated proteins (MAPs), and other regulatory proteins. We propose to study conserved mechanisms of spindle organization and function in the genetically-tractable model system fission yeast and in human cultured cells.