Molecular Analysis of the Kinetochore-Microtubule Interface

描述（由申请人提供）：人体中的每个细胞包含46个不同的染色体，大单位的DNA编码指令细胞生长，分裂，并执行其特殊功能。在有丝分裂过程中，当细胞分裂时，这些染色体中的每一条都必须准确地分布到两个新的子细胞中。如果这个过程在单个染色体上发生错误，那么产生的子细胞将失去或获得数千个基因和它们所包含的指令。这种类型的染色体分离错误可导致细胞死亡，并被认为有助于肿瘤的发生。事实上，多达70%的肿瘤被观察到有异常数量的染色体。为了在有丝分裂过程中促进DNA的分离，染色体必须产生物理附着在称为微管的棒状聚合物上，微管提供了染色体移动的结构和力量。抗有丝分裂药物破坏这些微管与染色体连接的能力，通常用于癌症化疗。然而，由于微管在神经系统中的额外作用，许多这些药物具有有害的继发性影响。在染色体分离中起关键作用的是一种称为着丝点的大型蛋白质结构，它形成染色体和微管之间的界面。在避免与微管结合化疗药物相关的剂量限制性神经元毒性的同时，预测着丝粒活性的抑制作用可靶向癌细胞。事实上，针对几种着丝点蛋白的抑制剂目前正在临床试验中。确定每个人类着丝点蛋白的特定活性对于提供其在染色体分离中的功能背景，评估疾病诊断和治疗的最佳靶点以及生成适合分离小分子抑制剂的测定方法至关重要。拟议的工作将分析与微管相互作用所需的人类着丝点蛋白的功能和调控。这项工作将集中在最近发现的两个关键的与微管聚合物直接结合的着丝点相关蛋白组。这些研究将定义这些蛋白质的特性，确定这些蛋白质与微管相互作用的机制，剖析它们通过控制着丝点-微管附着的上游激酶的调节，并检查它们在人类细胞中的功能。总的来说，这些研究将定义着丝点-微管相互作用的基础，最终将为癌症诊断和治疗的实验提供基础。有丝分裂缺陷导致染色体数目错误，可导致细胞死亡，并被认为有助于肿瘤进展。了解这些DNA单元及其包含的遗传信息如何均匀地分布到新细胞中，对于癌症的诊断和治疗至关重要。这项工作将确定在人类细胞中指导和控制染色体分离的机制。

DESCRIPTION (provided by applicant): Each cell in the human body contains 46 different chromosomes, large units of DNA that encode instructions for that cell to grow, divide, and carry out its specialized functions. During mitosis, when a cell divides, each of these chromosomes must be accurately distributed to the two new daughter cells. If this process occurs incorrectly for even a single chromosome, the resulting daughter cells will lose or gain thousands of genes and the instructions that they contain. This type of error in chromosome segregation can result in the death of the cell and is thought to contribute to tumorigenesis. Indeed, as many as 70% of tumors are observed to have abnormal numbers of chromosomes. To facilitate the segregation of DNA during mitosis, chromosomes must generate physical attachments to rod-like polymers termed microtubules that provide the structure and forces to move the chromosomes. Anti-mitotic drugs that disrupt the ability of these microtubules to connect with the chromosomes are routinely used for cancer chemotherapy. However, many of these drugs have deleterious secondary affects due to additional roles for microtubules in the nervous system. A key player in chromosome segregation is a large proteinaceous structure termed the kinetochore that forms the interface between the chromosomes and the microtubules. Inhibition of kinetochore activities is predicted target cancer cells while avoiding the dose-limiting neuronal toxicity associated with microtubule-binding chemotherapy drugs. Indeed, inhibitors against several kinetochore proteins are currently in clinical trials. Determining the specific activities of each human kinetochore protein is crucial to provide a context for their functions in chromosome segregation, to evaluate the best targets for the diagnosis and treatment of disease, and to generate assays suitable for the isolation of small molecule inhibitors. The proposed work will analyze the function and regulation of the human kinetochore proteins that are required to generate interactions with microtubules. This work will focus on two key, recently identified groups of kinetochore-associated proteins that bind to microtubule polymers directly. These studies will define the properties of these proteins and determine the mechanisms by which these proteins interact with microtubules, dissect their regulation by upstream kinases that control kinetochore-microtubule attachments, and examine their functions in human cells. In total, these studies will define the basis for kinetochore-microtubule interactions that will ultimately provide the foundation for experiments on the diagnosis and treatment of cancer. PUBLIC HEALTH RELEVANCE: Project Narrative Defects in mitosis that result in errors in chromosome numbers can cause the death of a cell and are thought to contribute to tumor progression. Understanding the means by which these units of DNA, and the genetic information that they contain, are evenly distributed to new cells is critical for the diagnosis and treatment of cancer. This proposed work will determine the mechanisms that direct and control chromosome segregation in human cells.