Mechanism and Consequences of Telomere Dysfunction

端粒功能障碍的机制和后果

• 批准号: 7559976
• 负责人: Zhou Songyang
• 金额: $ 31.85万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-04-01 至 2013-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7559976
• 关键词: Affect; Age; Binding; Binding Proteins; Biological Assay; Cell Cycle; Cell Cycle Checkpoint; Cell Survival; Cell physiology; Cells; Complex; Confusion; DNA Damage; Development; Disease; Dominant-Negative Mutation; Dysplasia; Fluorescence; Functional disorder; Genes; Genome Stability; Genomic Instability; Goals; Hematologic Neoplasms; Hematopoiesis; Hematopoietic; Hematopoietic System; Hematopoietic stem cells; Human; Knock-out; Lead; Length; Link; Malignant Neoplasms; Mammalian Cell; Mammals; Mass Spectrum Analysis; Mediating; Molecular; Mus; Mutation; Normal Cell; Physiological; Point Mutation; Post-Translational Protein Processing; Premature aging syndrome; Proteins; RNA Interference; RNA Splicing; Regulation; Regulatory Pathway; Role; Signal Transduction; Stem cells; TERF1 gene; TINF2 gene; Telomerase; Telomere Maintenance; Testing; Therapeutic; Tissues; Ubiquitination; Work; design; in vivo; insight; loss of function; mouse model; mutant; overexpression; protein complex; protein function; protein purification; reconstitution; response; self-renewal; small hairpin RNA; telomere; therapy development; tumorigenesis

DESCRIPTION (provided by applicant): Telomere integrity is important for mammalian cell survival and proliferation. Telomere attrition or exposed chromosomal ends could result in genome instability, DNA damage response, and ultimately cancer. Mutations in telomerase subunits or telomere regulators have been linked to premature aging and cancer. Understanding the cellular mechanisms of telomere maintenance may therefore help in developing therapies for diseases including cancer. The human 3' overhang binding protein POT1 helps to protect the telomere ssDNA and regulate telomerase access. Recently, the lab identified a new telomere associated protein TPP1/PTOP (also known as PIP1/TINT1). We found it to directly interact with POT1 and the telomerase. Furthermore, TPP1 forms the human telosome, along with POT1, TRF1, TRF2, RAP1, and TIN2. Our findings suggest that TPP1 controls telomere length by regulating telomere recruitment of POT1 and telomerase. The long-term goal is to study how dysregulation of telosome components may lead to telomere dysfunction and cancer development. The overall objective of this proposal is to understand the function of TPP1 in telomere protection and hematopoietic cancer. We hypothesize that TPP1 may heterodimerize with POT1 to protect telomere ends, and control telomerase access and activity. Specific Aim 1. To determine the molecular mechanism of TPP1-mediated telomere end protection and length control. (1) The dynamic assembly, protein stability, and localization of the POT1-TPP1 complex in human cells will be investigated. We will determine protein modifications such as ubiquitination, and the cell cycle regulated localization and organization of the POT1-TPP1 complex. (2) The role of TPP1 in telomere end protection, cell survival, and cell cycle checkpoint activation will be determined through knockout, RNAi, and dominant negative expression. Specific Aim 2: To elucidate the role of TPP1 in controlling telomerase activity. How TPP1 interacts with the telomerase and controls telomerase activity, regulates cell cycle-dependent telomerase access to telomeres will be studied. Specific Aim 3: To investigate the consequences of telomere dysfunction in development and cancer. Chimeric mice whose hematopoietic systems are reconstituted with cells expressing dominant-negative TPP1 and TPP1 shRNA will be generated, and used to determine how TPP1 affects hematopoietic survival and development of hematopoietic cancers. The studies described in this proposal should help to delineate the signaling mechanisms by which TPP1 regulates telomere maintenance in vivo, and provide insight into the regulation and mechanisms of TPP1 in development and telomere protection, especially during hematopoiesis. This work may provide a more refined picture about the complex signaling networks that are employed by the cells to control genome stability. Furthermore, valuable targets for mechanism-driven design of cancer and aging therapeutics may be identified through this project.

描述（由申请人提供）：端粒完整性对于哺乳动物细胞的存活和增殖很重要。端粒损耗或裸露的染色体末端可能导致基因组不稳定性，DNA损伤反应和最终导致癌症。端粒酶亚基或端粒调节剂的突变与过早的衰老和癌症有关。因此，了解端粒维持的细胞机制可能有助于开发包括癌症在内的疾病的疗法。人3'悬垂结合蛋白POT1有助于保护端粒ssDNA并调节端粒酶的访问。最近，该实验室确定了一种新的端粒蛋白TPP1/PTOP（也称为PIP1/TINT1）。我们发现它可以直接与POT1和端粒酶相互作用。此外，TPP1与POT1，TRF1，TRF2，RAP1和TIN2形成人端体。我们的发现表明，TPP1通过调节POT1和端粒酶的端粒募集来控制端粒长度。长期目标是研究端体组成分的失调可能导致端粒功能障碍和癌症发展。该提案的总体目的是了解TPP1在端粒保护和造血癌中的功能。我们假设TPP1可能与POT1异二聚二聚体以保护端粒末端，并控制端粒酶的访问和活动。具体目的1。确定TPP1介导的端粒保护和长度控制的分子机制。 （1）将研究人类细胞中POT1-TPP1复合物的动态组装，蛋白质稳定性和定位。我们将确定诸如泛素化之类的蛋白质修饰，以及细胞周期调节POT1-TPP1复合物的定位和组织。 （2）TPP1在端粒端保护，细胞存活和细胞周期检查点激活中的作用将通过敲除，RNAi和显性负表达确定。具体目标2：阐明TPP1在控制端粒酶活性中的作用。 TPP1如何与端粒酶相互作用并控制端粒酶活性，将研究调节细胞周期依赖性端粒酶对端粒的访问。特定目的3：研究端粒功能障碍在发育和癌症中的后果。将与表达显性阴性TPP1和TPP1 shRNA的细胞重构的造血系统的嵌合小鼠将产生，并用于确定TPP1如何影响造血生存和造血癌的发育。该提案中描述的研究应有助于描述TPP1调节体内端粒维持的信号传导机制，并洞悉TPP1在发育和端粒保护中的调节和机制，尤其是在造血期间。这项工作可能会提供有关细胞控制基因组稳定性的复杂信号网络的更精致图片。此外，可以通过该项目确定以机制驱动的癌症和衰老治疗剂设计的宝贵目标。