STRUCTURE & FUNCTION OF BASE MODIFICATIONS IN SUBTELOMERIC & TELOMERIC DNA

结构

• 批准号: 7355192
• 负责人: Woodring Erik Wright
• 金额: $ 0.42万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-02-01 至 2007-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7355192
• 关键词: STRUCTURE; FUNCTION; BASE; MODIFICATIONS; SUBTELOMERIC

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Normal human cells have a limited capacity to proliferate, a process termed replicative aging. Increasing evidence has implicated telomeres, the structures that cap the ends of the chromosomes, as the molecular clock that counts the number of times the cell has divided. The mechanism of lagging-strand DNA synthesis prevents DNA polymerase from replicating the DNA all the way to the 5'''' end of a linear chromosome, leaving a 3'''' overhang and causing the chromosomes to shorten every time a cell divides. Human telomeres are composed of many kilobases of the repetitive sequence TTAGGG that, together with telomere-binding proteins, prevent the cell from recognizing the end of the chromosome as a DNA break needing repair. Cellular senescence may occur when some of the telomeres have shortened sufficiently to induce a DNA damage signal. Cancer cells escape the proliferative limits of replicative aging by up-regulating the expression of telomerase, an enzyme capable of adding telomere repeats to the ends of the chreomsomes and maintaining their length. Using methods for identifying the presence of modified nucleotides in subtelomeric DNA, for purifying telomeres (based on the presence of the 3'''' G-rich overhang) that yields a greater than 1000-fold enrichment in a single step, for determining the size of the overhangs, and for measuring telomere sizes in interphase nuclei. These advances will permit us to pursue the following goals: (1) To understand the structure and function of base modifications in subtelomeric/telomeric DNA; (2) to determine what regulates the rate of telomere shortening; and (3) To define when and where chells with short telomeres accumulate in vivo in humans. The overall goal is to manipulate rates of telomere shortening, with obvious applications in developing therapeutics for aging and cancer. LC/MS and LC/MS/MS methods will be implemented and applied to analyzing samples for modified nucleobases.

该子项目是利用 NIH/NCRR 资助的中心拨款提供的资源的众多研究子项目之一。子项目和研究者 (PI) 可能已从另一个 NIH 来源获得主要资金，因此可以在其他 CRISP 条目中出现。列出的机构是中心的机构，不一定是研究者的机构。正常人体细胞的增殖能力有限，这一过程称为复制衰老。越来越多的证据表明，端粒（覆盖染色体末端的结构）是计算细胞分裂次数的分子钟。滞后链 DNA 合成机制阻止 DNA 聚合酶将 DNA 一直复制到线性染色体的 5'''' 末端，留下 3'''' 突出端，导致染色体在每次细胞分裂时缩短。人类端粒由数千个碱基的重复序列 TTAGGG 组成，与端粒结合蛋白一起，防止细胞将染色体末端识别为需要修复的 DNA 断裂。当一些端粒缩短到足以诱导 DNA 损伤信号时，细胞可能会发生衰老。癌细胞通过上调端粒酶的表达来逃避复制衰老的增殖限制，端粒酶是一种能够在染色体末端添加端粒重复序列并保持其长度的酶。使用方法鉴定亚端粒 DNA 中是否存在修饰核苷酸，纯化端粒（基于富含 3'''' G 的突出端），一步即可获得超过 1000 倍的富集，用于确定突出端的大小，并用于测量间期核中的端粒大小。这些进展将使我们能够实现以下目标：（1）了解亚端粒/端粒DNA中碱基修饰的结构和功能； (2)确定端粒缩短率的调节因素； (3) 确定短端粒细胞在人类体内积累的时间和地点。总体目标是控制端粒缩短率，在开发衰老和癌症疗法方面具有明显的应用。将实施 LC/MS 和 LC/MS/MS 方法并应用于分析样品中的修饰核碱基。