Molecular Analysis of the Kinetochore-Microtubule Interface

描述(申请人提供)：人体中的每个细胞包含46条不同的染色体，这是一种大的DNA单位，编码了细胞生长、分裂和执行其特殊功能的指令。在有丝分裂期间，当细胞分裂时，这些染色体中的每一条都必须准确地分配给两个新的子细胞。如果这一过程发生错误，即使只有一条染色体，由此产生的子细胞将失去或获得数千个基因和它们包含的指令。这种类型的染色体分离错误可能导致细胞死亡，并被认为是导致肿瘤发生的原因。事实上，多达70%的肿瘤被观察到具有异常数量的染色体。为了促进有丝分裂过程中DNA的分离，染色体必须与被称为微管的杆状聚合物产生物理连接，微管提供移动染色体的结构和力量。破坏这些微管与染色体连接能力的抗有丝分裂药物通常用于癌症化疗。然而，由于神经系统中微管的额外作用，这些药物中的许多都有有害的次要影响。染色体分离的一个关键因素是一个巨大的蛋白质结构，称为着丝粒，它形成了染色体和微管之间的界面。动粒活性的抑制被预测为靶向癌细胞，同时避免了与微管结合的化疗药物相关的剂量限制的神经元毒性。事实上，针对几种动粒蛋白的抑制剂目前正在进行临床试验。确定每个人动粒蛋白的特定活性对于提供它们在染色体分离中的功能、评估疾病诊断和治疗的最佳靶点以及产生适合于分离小分子抑制物的方法至关重要。这项拟议的工作将分析与微管相互作用所需的人类动粒蛋白的功能和调节。这项工作将集中在两组关键的，最近发现的动粒相关蛋白，直接结合到微管聚合物。这些研究将定义这些蛋白质的性质，确定这些蛋白质与微管相互作用的机制，剖析控制动粒-微管连接的上游激酶对它们的调节，并检查它们在人类细胞中的功能。总而言之，这些研究将确定动粒-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。公共卫生相关性：有丝分裂中导致染色体数目错误的项目叙事缺陷可能导致细胞死亡，并被认为有助于肿瘤的进展。了解这些DNA单位及其包含的遗传信息均匀分布到新细胞的方式对于癌症的诊断和治疗至关重要。这项拟议的工作将确定指导和控制人类细胞中染色体分离的机制。

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.