Molecular Mechanisms of Telomere Length Homeostasis

描述（由申请人提供）：本提案的长期目标是通过对端粒蛋白-DNA复合物的全面表征，详细了解端粒稳态的分子机制。端粒通过端粒酶促进染色体末端复制来确保基因组的完整性。此外，端粒还保护染色体末端免受DNA修复和降解活动的影响。端粒酶亚单位或端粒成分的突变与过早衰老和癌症有关。维持适当的端粒结构和长度是完成其染色体末端复制和末端保护任务所必需的。该复合物的结构-功能关系是理解端粒稳态机制的核心问题。我们将利用遗传上可分离的S。pombe模型系统，并采用生物化学，结构和遗传工具的组合，以实现以下具体目标：1.绘制Tpz 1与其相互作用伙伴之间的蛋白质-蛋白质相互作用界面，并研究体外以Tpz 1为中心的复合物形成的动力学。2.揭示Tpz 1介导的相互作用在端粒长度稳态和染色体末端保护中的功能作用。3.使用单粒子低温电子显微镜（cryo-EM）和X射线晶体学对Tpz 1中心复合物进行全面的结构分析。这些目标的实现将为端粒复合物的结构-功能关系和动力学提供新的和重要的机制见解，也可能为开发针对端粒功能障碍引起的疾病的新治疗方法提供基础。

DESCRIPTION (provided by applicant): The long-term goal of this proposal is to understand, in detail, the molecular mechanisms of telomere homeostasis through comprehensive characterization of the telomere protein-DNA complex. Telomeres ensure genome integrity by facilitating chromosome end replication through telomerase. In addition, telomeres also protect the chromosome ends from DNA repair and degradation activities. Mutations in telomerase subunits or telomere components have been linked to premature aging and cancer. Maintaining proper telomere structure and length is required for accomplishing both its chromosome end replication and end protection tasks. The structure-function relationship of this complex is a central question in understanding the mechanisms of telomere homeostasis. We will exploit the genetically amendable S. pombe model system and employ a combination of biochemical, structural and genetic tools to achieve the following specific aims: 1. Map the respective protein-protein interaction interfaces between Tpz1 and its interaction partners and examine the dynamics of the Tpz1-centered complex formation in vitro. 2. Reveal the functional roles of Tpz1 mediated interactions in telomere length homeostasis and chromosome end protection. 3. Achieve a comprehensive structural analysis of the Tpz1-centered complex using single particle cryo-electron microscopy (cryo-EM) and X-ray crystallography. Accomplishment of the proposed aims will provide new and significant mechanistic insights into the structure-function relationship and the dynamics of the telomere complex, and may also provide the foundation for development of new therapeutic approaches against diseases caused by telomere dysfunction.