ROS driven mitochondrial-telomere dysfunction during environmental stress

 描述(由申请人提供)：维持线粒体和端粒功能对健康衰老至关重要，这些不同的间隔之间会发生显著的串扰。许多环境因素导致线粒体功能障碍和随后的活性氧物种(ROS)的产生，特别是对端粒的损伤。该项目将直接测试这一假说，即端粒的氧化损伤会导致线粒体功能障碍，反过来，线粒体DNA的氧化损伤会导致端粒功能障碍，这种相互作用的损伤会导致几种环境诱导的人类疾病，包括帕金森氏病(PD)中的神经变性。我们将使用一种高度创新的系统来监测不同细胞隔间的ROS通量，该系统包括快速诱导ROS的荧光蛋白标记和可见光，以及带有独特化学传感器的荧光素激活肽(FAP)来检测ROS。这些FAP还将与不同的化学部分一起使用，以产生不同类型的RO。我们将使用一种与PD相关的环境杀虫剂作为线粒体毒剂来检测ROS通量和随后的端粒损伤。R21阶段将首先在人类细胞中开发和验证这种方法，并将产生将该系统应用于斑马鱼的转基因动物。Aim 1将使用KillerRed ROS生成系统来研究线粒体中ROS的生成如何影响端粒功能，以及端粒上ROS的生成如何改变线粒体功能。AIM 2将开发FAP系统，在线粒体或端粒内感应和产生ROS，并将使用这项技术来检测从线粒体到端粒的ROS通量。我们将创建转基因斑马鱼驱动系，用于在线粒体或端粒中定位FAP介导的ROS传感和生成。R33阶段将应用靶向ROS传感/产生系统来研究人类神经细胞(目标3)、转基因斑马鱼胚胎(目标4)和帕金森病特定斑马鱼模型(目标5)中功能失调的线粒体和端粒串扰的潜在机制。这些创新研究将测量活细胞中ROS的时间和空间生成，并提供机制洞察功能失调的端粒或线粒体在环境诱导的人类疾病(包括帕金森病)的过程中如何相互影响。这个项目建立了检测ROS介导的通量和线粒体对环境应激源的串扰的工具和能力。该项目的完成将为开发新的干预措施奠定基础，以更好地减轻环境暴露对端粒和线粒体功能的负面影响，有助于改善或延缓与衰老有关的疾病和病理。

 DESCRIPTION (provided by applicant): Maintenance of mitochondrial and telomere function are critical for healthy aging, and significant cross-talk occurs between these distinct compartments. Many environmental factors cause mitochondrial dysfunction and subsequent reactive oxygen species (ROS) generation that particularly damage the telomeres. This project will directly test the hypothesis that oxidative damage at telomeres cause mitochondrial dysfunction, and conversely that oxidative damage to mitochondrial DNA cause telomere dysfunction, and that this reciprocal damage contributes to several environmentally-induced human disease, including neurodegeneration in Parkinson's disease (PD). We will monitor ROS flux in distinct cellular compartments using a highly innovative system consisting of fluorescent protein tagging and visible light to rapidly induce ROS, and fluorogen-activating peptides (FAPs) with unique chemical sensors to detect ROS. These FAPs will also be used with different chemical moieties to generate different types of ROS. We will use an environmental pesticide associated with PD as a mitochondrial toxicant to examine ROS flux and subsequent telomere damage. The R21 phase will develop and validate this approach first in human cells and will generate transgenic animals for applying this system to zebrafish. Aim 1 will use the KillerRed ROS-generating system to examine how ROS generation in mitochondria impacts telomere function, and reciprocally how ROS generation at telomeres alters mitochondrial function. Aim 2 will develop the FAP system for sensing and producing ROS within the mitochondria or telomeres, and will use this technology to examine ROS flux from the mitochondria to the telomeres. We will create transgenic zebrafish driver lines for localized FAP-mediated ROS sensing and generation in the mitochondria or telomeres. The R33 phase will apply the targeted ROS sensing/producing system toward investigating the underlying mechanisms of dysfunctional mitochondria and telomere cross-talk in human neuronal cells (Aim 3), in transgenic zebrafish embryos (Aim 4) and in a specific zebrafish model of PD (Aim 5). These innovative studies will measure the temporal and spatial generation of ROS in living cells and provide mechanistic insight into how dysfunctional telomeres or mitochondria influence each other in the process of environmentally-induced human diseases, including PD. This project builds tools and capacity for examining ROS-mediated flux and mitochondrial cross-talk in response to environmental stressors. Completion of this project will lay the foundation for developing new interventions to better mitigate the negative effects of environmental exposures on telomere and mitochondria function, serving to ameliorate or delay aging-related diseases and pathologies.