The roles of telomerase and non-coding RNA in cancer

端粒酶和非编码RNA在癌症中的作用

• 批准号: 10218067
• 负责人: Linnea Jansson-Fritzberg
• 金额: $ 10.51万
• 依托单位: UNIVERSITY OF COLORADO
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-01 至 2023-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10218067
• 关键词: Academia; Active Sites; Address; Amino Acids; Animal Model; Area; Biochemical; Biology; Boundary Elements; C-terminal; Catalysis; Cell Cycle Arrest; Cell division; Cells; Chromosomes; Collaborations; Communication; Computer Models; DNA Binding; DNA Structure; Data; Development; Disease; Drug Targeting; Enzymes; Event; Faculty; Fluorescence Resonance Energy Transfer; Functional disorder; Goals; Human; Hybrids; Individual; Journals; Knowledge; Label; Length; Link; Malignant Neoplasms; Measurement; Mediating; Mentors; Methods; Molecular Biology; Molecular Conformation; Monitor; N-terminal; Oral; Paper; Phase; Play; Proteins; Publications; RNA; RNA Recognition Motif; Refractory; Research; Research Project Grants; Resolution; Ribonucleoproteins; Role; Scheme; Structural Models; Structure; Techniques; Telomerase; Telomerase RNA Component; Telomere Maintenance; Telomere Shortening; Testing; Tetrahymena thermophila; Therapeutic; Time; Tissues; Untranslated RNA; Work; Writing; X-Ray Crystallography; biochemical tools; biophysical analysis; biophysical tools; cancer cell; career; human model; insight; interest; meetings; novel; pre-doctoral; senescence; single molecule; skills; symposium; targeted cancer therapy; telomere; therapeutic target; tool

Project Summary The predoctoral phase of this proposal will focus on the biochemical and biophysical studies of the enzyme telomerase. Telomerase is a ribonucleoprotein that maintains telomere lengths in rapidly dividing cells, thereby counteracting the chromosome shortening that is inherent to each round of cell division. Without telomerase, cells undergo senescence when telomere lengths become critically short. Telomerase is upregulated in about 90% of cancers due to the need for telomere maintenance in rapidly dividing cells making it an attractive drug target. Cancer therapeutics that target active telomerase directly have remained elusive however. The structure and function of human telomerase is still poorly understood, and any new insight would greatly benefit the development of novel cancer therapeutics. The dissertation project focuses on using single molecule techniques to study the conformation and dynamics of human telomerase. The minimal human telomerase is composed of two subunits, a protein component (TERT) that contains four evolutionarily conserved domains and an RNA component (TER) which contains the integral non-coding RNA template from which telomeres are reverse transcribed. Telomerase is unique in its ability to reverse transcribe multiple telomere repeats during a single DNA binding event, but the details of how telomerase maintain this processive action remains poorly understood. Additionally, details regarding how TERT and TER interact during catalysis, and how individual TERT domains regulate telomerase dynamics, remain unknown. Specifically, knowledge about the function and dynamics of two TERT domains, the Telomerase Essential N-terminal (TEN) domain and the C-terminal extension (CTE) remain especially poorly understood. This project will test the hypothesis that the TEN and CTE domains drive the processive action of telomerase via coordinated dynamic rearrangement of the telomere/template hybrid into its initial bound state at the end of each telomere repeat addition event. Single molecule Förster Resonance Energy Transfer (smFRET), in combination with a novel protein labeling scheme, will be used to probe the conformation and dynamics of these domains during different functional states. In addition, computational modeling in collaboration with the Das lab at Stanford will enable the construction of a refined working model of human telomerase during catalysis. Completion of this proposal will aid in the development of telomerase targeted cancer therapeutics. In addition, this proposal will facilitate the smooth transition from studying RNP structure and function during the predoctoral phase, to the study of long non-coding RNA (lncRNA) structure and its roles in the development of cancer during the postdoctoral phase. The techniques and approaches learned during the dissertation research will lend themselves perfectly to the study of RNA structure, which to date has remained a difficult area to study. The timing of the proposed postdoctoral work focused on lncRNA structure and cancer will be ideal considering the current stage of lncRNA biology and overall, this proposal is well suited to provide valuable insight into how RNA structure contributes to cancer development. !

项目概要 该提案的博士前阶段将重点关注酶的生物化学和生物物理研究 端粒酶。端粒酶是一种核糖核蛋白，可在快速分裂的细胞中维持端粒长度，从而 抵消每轮细胞分裂所固有的染色体缩短。如果没有端粒酶， 当端粒长度变得非常短时，细胞就会经历衰老​​。端粒酶上调约 90% 的癌症是由于快速分裂的细胞需要端粒维持而导致的，这使其成为一种有吸引力的药物 目标。然而，直接靶向活性端粒酶的癌症疗法仍然难以捉摸。结构 人类端粒酶的功能仍然知之甚少，任何新的见解都将极大地有益于人类端粒酶的功能。 开发新型癌症疗法。该论文项目侧重于使用单分子技术 研究人类端粒酶的构象和动力学。最小的人类端粒酶由以下组成 两个亚基，一个包含四个进化保守结构域的蛋白质成分 (TERT) 和一个 RNA 成分 (TER)，包含完整的非编码 RNA 模板，端粒与之相反 转录。端粒酶的独特之处在于它能够在一次逆转录多个端粒重复序列的过程中逆转录多个端粒重复序列。 DNA 结合事件，但端粒酶如何维持这种持续作用的细节仍然知之甚少。 此外，有关 TERT 和 TER 在催化过程中如何相互作用以及各个 TERT 域如何相互作用的详细信息 调节端粒酶动力学，仍然未知。具体来说，关于功能和动力学的知识 两个 TERT 结构域，端粒酶必需 N 端 (TEN) 结构域和 C 端延伸 (CTE) 人们对这一问题的了解仍然特别少。该项目将测试 TEN 和 CTE 域驱动的假设 端粒酶通过端粒/模板杂合体的协调动态重排的持续作用 每个端粒重复添加事件结束时的初始结合状态。单分子福斯特共振 能量转移（smFRET）与新型蛋白质标记方案相结合，将用于探测 这些结构域在不同功能状态下的构象和动力学。此外，计算 与斯坦福大学 Das 实验室合作进行建模将能够构建一个精细的工作模型 催化过程中的人类端粒酶。该提案的完成将有助于端粒酶的发展 靶向癌症治疗。此外，该提案将有助于从研究 RNP 的平稳过渡 博士前阶段的结构和功能，以研究长链非编码RNA（lncRNA）结构和 它在博士后阶段的癌症发展中的作用。学到的技术和方法 论文研究期间的研究将非常适合 RNA 结构的研究，迄今为止，RNA 结构的研究已 仍然是一个难以研究的领域。拟议博士后工作的时间重点是lncRNA结构 考虑到 lncRNA 生物学的当前阶段，癌症将是理想的选择，总体而言，该提案非常适合 提供有关 RNA 结构如何促进癌症发展的宝贵见解。 ！