Role of skeletal muscle mitophagy in healthy aging

项目摘要 骨骼肌线粒体质量随着年龄的增长而逐渐下降，导致组织功能障碍 和疾病几条证据表明老年动物骨骼肌中线粒体质量较差， 在很大程度上是由于受损或不足的能力，以降低受损/功能障碍 线粒体自噬。运动促进线粒体质量，导致健康衰老，但潜在的 机制以及它们如何随年龄而不同还没有很好地定义，特别是在线粒体自噬，抑制 制定有效的干预措施。在我之前的工作基础上，我证明了锻炼确实 促进骨骼肌中的线粒体自噬，但只占总线粒体网的一小部分，我在这里展示 Ampk依赖性机制的证据，可以区分受损与健康的区域， 随着年龄的增长而丧失的线粒体网状结构。此外，我发现关键的下游线粒体自噬相关因子 在运动中被招募到骨骼肌线粒体中的蛋白质是线粒体质量所必需的， 健康老龄化在D.黑腹菌拟议的研究测试的假设，识别受损 老年小鼠骨骼肌中对运动作出反应的线粒体网状区域受损， 关键线粒体自噬蛋白的局部募集，导致线粒体质量差。这些研究将提供 深入了解骨骼肌线粒体自噬对运动反应的新调节，并为研究运动对骨骼肌线粒体自噬的影响奠定基础。 开发有针对性的干预措施，以提高骨骼肌中的线粒体质量，改善组织 功能和健康老化。在指导阶段，我将采用最先进的双光子显微镜， 进行骨骼肌线粒体上Ampk活性的活体和离体荧光寿命显微镜检查 年轻和老年小鼠，以确定年龄依赖性，局部反应的Ampk持续收缩 和线粒体损伤此外，我将继续我的专业和科学发展，为 在我的指导委员会的持续指导下，独立阶段。在独立阶段， 我将采用共体基因转移在骨骼肌的年轻和老年野生型小鼠以及骨骼肌， 肌肉特异性、条件性Ulk 1敲除小鼠，以确定募集的年龄依赖性调节 下游线粒体自噬相关因子对运动中骨骼肌线粒体的反应。我必 开发新的磷酸模拟构建体以组成性激活或抑制线粒体自噬相关因子Atg 9， atg2在青年和老年小鼠骨骼肌中的作用，并探讨其对骨骼肌破坏的必要性和充分性 通过最先进的高分辨率蛋白质组学技术， 和共焦成像方法。总的来说，这些研究和职业发展活动将促进我的 持续的科学和专业培训，导致一个成功的独立，学术研究计划。

Project Summary Mitochondrial quality in skeletal muscle progressively declines with advancing age, leading to tissue dysfunction and disease. Several lines of evidence suggest poor mitochondrial quality in skeletal muscle of old animals and humans is due in large part to an impaired or insufficient capacity to degrade damaged/dysfunctional mitochondria via mitophagy. Exercise promotes mitochondrial quality leading to healthy aging but the underlying mechanisms and how they differ with age is not well defined, particularly in regards to mitophagy, restraining development of effective interventions. Building upon my previous work demonstrating that exercise does indeed promote mitophagy in skeletal muscle but only of a small fraction of the total mitochondrial reticulum, I show here evidence of an Ampk-dependent mechanism that may distinguish damaged vs. healthy regions of the mitochondrial reticulum that is lost with age. Additionally, I show that key downstream mitophagy-related factors that are recruited to mitochondria in skeletal muscle with exercise are required for mitochondrial quality and healthy aging in d. melanogaster. The proposed research tests the hypothesis that recognition of damaged regions of the mitochondrial reticulum in response to exercise is impaired in skeletal muscle of old mice, blunting local recruitment of key mitophagy proteins, leading to poor mitochondrial quality. These studies will provide insight into novel regulation of skeletal muscle mitophagy in response to exercise and lay a foundation for the development of targeted interventions to promote mitochondrial quality in skeletal muscle for improved tissue function and healthy aging. During the mentored phase, I will employ state-of-the-art two-photon microscopy to perform intravital and ex vivo fluorescent lifetime microscopy of Ampk activity on mitochondria in skeletal muscle of young and old mice to determine the age-dependent, localized response of Ampk to sustained contraction and mitochondrial damage. Also, I will continue my professional and scientific development in preparation for the independent phase with continuous guidance from my mentoring committee. During the independent phase, I will employ co-somatic gene transfer in skeletal muscle of young and old wild-type mice as well as skeletal muscle-specific, conditional Ulk1 knock-out mice to determine the age-dependent regulation for the recruitment of downstream mitophagy-related factors to mitochondria in skeletal muscle in response to exercise. Also, I will develop novel phospho-mimetic constructs to constitutively activate or inhibit mitophagy-related factors Atg9 and Atg2 in young and old mouse skeletal muscle and investigate their necessity and sufficiency for the breakdown of mitochondrial proteins and maintenance of mitochondrial quality, via state-of-the-art high resolution proteomic and confocal imaging approaches. Collectively, these studies and career development activities will foster my continued scientific and professional training, leading to a successful independent, academic research program.