Genomic instability, susceptibility to oxidative stress and cellular senescence

项目总结/摘要 所有生物体都不断地暴露于各种环境压力，无论是人为的还是 自然的起源。许多压力源都有一个共同的毒性机制，可以对细胞产生有害影响 (e.g.,高活性化学物质的产生）。这些化学物质的一部分，逃避细胞的 防御和破坏细胞成分，包括DNA，导致氧化。全球基因组测量 氧化损伤DNA的水平，暗示了主要人类健康问题中的环境应激源（例如， 癌症、衰老、心血管和神经变性疾病）。然而，目前DNA之间的关联 损害和疾病是基于对全球基因组损害的粗略评估， 关于损伤如何导致疾病的机械信息。此外，DNA损伤不是均匀分布的， 在整个基因组中;在基因组区域中对功能至关重要的损伤的积累或持续 则小区将具有下游后果。我们认为，DNA损伤在疾病中的作用只能是 通过在损害所在地的背景下审查损害来理解。我们目前缺乏关于 细胞如何维持氧化损伤DNA的基线水平，以及其在不同时间的时空分布。 基因组这对于我们理解细胞如何对损伤做出反应和靶向区域至关重要 优先修复。本研究的目的是确定基因组中易感性增加的区域 氧化损伤DNA的形成或缓慢修复，在衰老中发挥功能作用。本研究 将让学生独立，有价值的研究，加强机构的研究环境。 目标1。确定基因组中易受氧化应激损伤影响的区域。 A.表征基础水平8-oxodG的分布及其修复监测。DNA损伤的模式 并且在衰老干细胞模型中使用DDIP-seq针对8-oxodG和ChIP-seq针对hOGG 1鉴定修复。 B。开发一个模型来询问影响DNA损伤和修复分布的因素，并预测 下游影响。DNA损伤和修复将被映射到特定的基因序列，内含子/外显子， 调控序列和染色体位置。识别可能具有功能性的基因组区域 细胞功能障碍和ROS诱导的衰老的后果。 C.评估核组织在损伤和修复中的作用。FISH将用于形成时空 图1A和1B中鉴定的基因组区域的DNA损伤和修复的拓扑图。 目标2.确定氧化应激敏感性增加与衰老的关系。 检查增加的内源性ROS对DNA损伤和修复的靶向作用。我们假设 增加的活性氧改变了损伤的分布和驱动疾病进程的修复。使用上述 方法，在我们的衰老干细胞模型中增加内源性ROS产生。我们假设任何 确定的新区域将代表在触发衰老中起作用的候选者。

Project Summary / Abstract All living organisms are continually exposed to a variety of environmental stressors, be they anthropogenic or natural in origin. Many stressors share a common toxic mechanism that can have detrimental effects on the cell (e.g., generation of highly reactive chemical species). A proportion of these chemical species, evade the cell's defenses and damage cellular components including DNA, causing oxidation. Measurement of global genome levels of oxidatively damaged DNA, implicates environmental stressors in major human health issues (e.g., cancer, aging, cardiovascular, and neurodegenerative diseases). However, current associations between DNA damage and disease are based upon crude assessments of global genome damage, which provide limited mechanistic information on how damage leads to disease. Furthermore, DNA damage is not uniformly distributed across the genome; accumulation, or persistence, of damage in regions of the genome vital to the functioning of the cell will have downstream consequences. We propose, that the role of DNA damage in disease can only be understood by examination of damage in the context of its location. We currently lack information concerning how the cell maintains baseline levels of oxidatively damaged DNA, and its spatio-temporal distribution across the genome. This is fundamental to our understanding of how the cell responds to damage and targets regions for prioritized repair. The objective of this study is to identify regions of the genome with increased susceptibility to the formation, or slow repair, of oxidatively damaged DNA that play a functional role in senescence. This study will engage students in independent, meritorious research, strengthening the institutional research environment. Aim 1. To identify susceptible regions of the genome for oxidative stress-induced damage. A. Characterize the distribution of basal levels 8-oxodG, and its repair surveillance. Patterns of DNA damage and repair will be identified using DDIP-seq for 8-oxodG, and ChIP-seq for hOGG1 in an aging stem cell model. B. Develop a model to interrogate the factors influencing the distribution of DNA damage and repair and predict downstream effects. DNA damage and repair will be mapped to specific gene sequences, introns/exons, regulatory sequences, and chromosomal locations. Identifying regions of the genome that may have functional consequences in cellular dysfunction and ROS-induced senescence. C. Assess the role of nuclear organization on damage and repair. FISH will be utilized to form spatio-temporal topological maps of DNA damage and repair across the genomic regions identified in 1A and 1B. Aim 2. To determine the mechanisms linking increased susceptibility to oxidative stress to senescence. Examine the effect of increased endogenous ROS on the targeting of DNA damage and repair. We hypothesize that increased ROS alters the distribution of damage and repair driving the disease process. Using the above approaches, in our aging stem cell model with increased endogenous ROS production. We hypothesize that any new regions identified will represent candidates for having a role in triggering senescence.