Investigating the Impact of Telomere Specific Oxidative Base Damage in Cellular Aging

项目概要/摘要 端粒在细胞衰老和癌发生之间的界面上起着至关重要的作用，因为它们 对氧化应激高度敏感。氧化应激，或自由基过量，是DNA的普遍来源 人类因正常新陈代谢、身体压力、污染等外源性来源而遭受的损害， 太阳辐射和免疫细胞信号的副产品。虽然端粒磨损和氧化损伤之间存在联系， 虽然存在应激，但还没有关于端粒DNA的直接氧化碱基损伤如何影响正常 细胞生物学我们的小组已经开发了第一个系统，以诱导常见的氧化损伤8-氧代鸟嘌呤 通过将荧光团激活肽（FAP）融合到端粒结合蛋白， TRF 1。我们已经产生了在端粒具有同源FAP-TRF 1表达的克隆细胞系， 我发现在端粒8 oxoG的单一诱导后，正常细胞表现出生长停滞，这是我们没有观察到的 在两个癌细胞系中。该研究的假设是端粒8 oxoG足以产生DDR。 在正常细胞中诱导生长停滞，不依赖于端粒缩短，这取决于p53/p21 信号轴此外，在诱导端粒8 oxoG后，我们观察到端粒数目的增加。 微核早在治疗后一天。该项目还将解决这些微核是如何形成的， 如果它们依赖于DNA复制，并且如果它们被cGAS/STING轴感测。竞争这个 该项目将首次描述端粒氧化应激对细胞结果的贡献 与衰老和癌症有关。这项工作将普遍适用于所有人类，但也将告知 与氧化应激水平增加有关的年龄相关疾病的生物学，并可能促进 这种奖学金还允许获得与本组织目标有关的新技能， 项目，并为每个项目列出了具体的导师。这包括端粒的单分子分析 复制、活细胞成像、RNA测序和检测与免疫相关的信号分子。 系统此外，职业发展也是该奖学金的重点。它包括参加讲习班的计划 与实验技术和科学写作有关，除了参加国家和国际 会议上介绍了这个研究项目的结果。

Project Summary/Abstract Telomeres play an essential role at the interface between cellular aging and carcinogenesis because they are highly sensitive to oxidative stress. Oxidative stress, or the excess of free radicals, is a ubiquitous source of DNA damage humans experience from normal metabolism, physical stress, exogenous sources such as pollution and solar radiation, and by-products of immune cell signaling. While a link between telomere attrition and oxidative stress exists, there have been no reports on how direct oxidative base damage to telomeric DNA impacts normal cell biology. Our group has developed the first system to induce the common oxidative lesion 8-oxo-guanine (8oxoG) specifically at telomeres by fusing a fluorogen activated peptide (FAP) to the telomere binding protein TRF1. We have generated clonal cell lines that have homogenous FAP-TRF1 expression at the telomere, and find that after a single induction of telomeric 8oxoG, normal cells exhibit a growth arrest, which we did not observe in two cancer cell lines. The hypothesis of this fellowship is that telomeric 8oxoG is sufficient to produce a DDR in normal cells that induces growth arrest, independent of telomere shortening, that depends on the p53/p21 signaling axis. Moreover, following induction of telomeric 8oxoG we observe an increase in the number of micronuclei as early as one day after treatment. This project will also address how these micronuclei are forming, if they are dependent on DNA replication, and if they are sensed by the cGAS/STING axis. Competition of this project will delineate for the first time the contribution of telomeric oxidative stress to cellular outcomes related to aging and cancer. This work will have general applications to all humans, but will also inform the biology of age-related diseases that are associated with increased levels of oxidative stress, and may promote health-span in either case.This fellowship also allows for the acquisition of new skills related to the aims of the project, and has outlines specific mentors for each. This includes single molecule analyses of telomere replication, live-cell imaging, RNA-sequencing, and detection of signaling molecules related to the immune system. Moreover, career development is also a focus of this fellowship. It includes plans to attend workshops related to experimental techniques and scientific writing, in addition to attending national and international conferences to present the findings of this research project.