Aging Mitochondrial Interactome

衰老线粒体相互作用组

基本信息

批准号：
10658412
负责人：
James Edward Bruce
金额：
$ 54.51万
依托单位：
UNIVERSITY OF WASHINGTON
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-05-01 至 2028-02-29
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10658412
关键词：
ATP Synthesis Pathway Activity Cycles Acute Address Adult Age Aging Biological Assay Biopsy Calcium Cardiovascular system Chemicals Citric Acid Cycle Complex Data Elderly Electron Transport Functional disorder Genes Glutamate Dehydrogenase Glutamate Metabolism Pathway Glutamates Heart Homeostasis Human Impairment In Vitro Intervention Lead Link Maps Mass Spectrum Analysis Measures Mediating Metabolism Mitochondria Mitochondrial Proteins Modeling Molecular Mus Muscle Muscle Mitochondria Myocardium Nature Nucleic Acid Regulatory Sequences Organ Organelles Oxidation-Reduction Pathology Peptides Permeability Phosphorylation Play Positioning Attribute Production Protein Conformation Proteins Quality of life Regulation Research Respiration Role SOD2 gene Site Skeletal Muscle Stress System Testing Tissues Translating Uncertainty age related aged alpha ketoglutarate catalase crosslink experimental study functional decline human subject improved in vivo induced pluripotent stem cell derived cardiomyocytes innovation insight knock-down mitochondrial dysfunction mitochondrial metabolism mouse model neuromuscular novel protein complex resilience tool young adult

项目摘要

Mitochondria play a central role in age-related pathologies, loss of resilience, and the decline in quality of life in older adults. As we age changes in mitochondrial function lead to disruption of redox and energy homeostasis, altered metabolite levels, impaired calcium regulation, and increased sensitivity to permeability transition, all of which contribute to tissue dysfunction. Mitochondria are dynamic organelles that continuously adapt to changing cellular demands by altering protein assembly and interactions to modify their function. Despite the obvious importance little is known about how age-related changes in mitochondrial protein interactions underlie changes in function. To address this fundamental question we propose to apply a state of the art novel quantitative chemical cross-linking with mass spectrometry approach (XL-MS) to quantify the changes in the mitochondrial interactome in heart and skeletal muscles with age. By combining this innovative XL-MS approach with targeted functional assays and interventions developed over the last several years to manipulate mitochondria in vivo and in vitro we are uniquely positioned to identify the changes in mitochondrial protein interactions that underlie age- related mitochondrial dysfunction. Our preliminary data indicate specific disruption of multiple protein interaction networks involved in ADP transport, ATP synthesis, glutamate metabolism, and complexes of the electron transport system with age. Many of these protein complexes that XL-MS indicates are disrupted in aging directly interact with the mitochondrial targeted peptide SS-31 that we have shown reverses mitochondrial dysfunction in heart and skeletal muscle. Our overall hypothesis is that changes in the mitochondrial interactome driven by elevated mitochondrial redox stress underlie decreased ATP production and altered metabolite levels in mitochondria from aged heart and skeletal muscle. In aim 1 we will apply XL-MS and site-specific assays to measure the link between the interactome and function in mitochondria from aged mouse heart and skeletal muscle. As a specific test of new mechanistic insights available from this approach we will use gene-edited iPSC derived cardiomyocytes to test the causal relationship between altered interactions in the glutamate dehydrogenase regulatory region, the largest change identified in the aged mitochondrial interactome, and impaired glutamate and α-ketoglutarate metabolism. In aim 2 we will use mitochondrial targeted interventions to reverse mitochondrial dysfunction, SS-31 and AAV-mitochondrial targeted catalase, and induce mitochondrial redox stress to identify the most functionally important changes in the mitochondrial interactome from aim 1. In aim 3 we use human muscle biopsies to test whether changes in the mitochondrial interactome identified in aims 1 and 2 translate into aged human skeletal muscle. We will separate older adults into low and high performing groups to compare the mitochondrial interactomes and function with those of young adults. Results from these experiments will have a transformative impact on the field by providing the first window into the links between the mitochondrial protein interaction network and age-related mitochondrial dysfunction. 1

线粒体在与年龄相关的病理、弹性丧失和生活质量下降中发挥着核心作用随着年龄的增长，线粒体功能的变化会导致氧化还原和能量稳态的破坏，代谢水平改变、钙调节受损以及对通透性转变的敏感性增加，所有这些导致组织功能障碍的线粒体是不断适应变化的动态细胞器。尽管显而易见，但通过改变蛋白质组装和相互作用来改变其功能的细胞需求。关于线粒体蛋白质相互作用中与年龄相关的变化如何引起变化，人们知之甚少为了解决这个基本问题，我们建议应用最先进的定量小说。化学交联与质谱法 (XL-MS) 来量化线粒体的变化通过将这种创新的 XL-MS 方法与有针对性的方法相结合，研究心脏和骨骼肌随年龄的相互作用。过去几年开发的功能测定和干预措施可在体内操纵线粒体在体外，我们具有独特的优势，可以识别年龄相关的线粒体蛋白质相互作用的变化我们的初步数据表明多种蛋白质相互作用的特定破坏。参与 ADP 运输、ATP 合成、谷氨酸代谢和电子复合物的网络 XL-MS 表明许多蛋白质复合物在衰老过程中直接受到破坏。与线粒体靶向肽 SS-31 相互作用，我们已证明该肽可逆转线粒体功能障碍我们的总体假设是线粒体相互作用组的变化是由线粒体相互作用驱动的。线粒体氧化还原应激升高是 ATP 产生和代谢水平改变的基础在目标 1 中，我们将应用 XL-MS 和位点特异性检测来检测来自衰老心脏和骨骼肌的线粒体。测量老年小鼠心脏和骨骼线粒体中相互作用组和功能之间的联系作为对这种方法新机制见解的具体测试，我们将使用基因编辑的 iPSC。衍生的心肌细胞来测试谷氨酸相互作用之间的因果关系脱氢酶调节区，在衰老的线粒体相互作用组中发现的最大变化，以及谷氨酸和 α-酮戊二酸代谢受损在目标 2 中，我们将使用线粒体靶向干预措施。逆转线粒体功能障碍，SS-31 和 AAV 线粒体靶向过氧化氢酶，并诱导线粒体氧化还原应激，以确定目标 1 中线粒体相互作用组中功能最重要的变化。目标 3 我们使用人体肌肉活检来测试目标中确定的线粒体相互作用组是否发生变化 1 和 2 转化为老年人的骨骼肌，我们将老年人分为低表现和高表现。小组将线粒体相互作用组和功能与年轻人的结果进行比较。实验将通过提供第一个窗口来了解之间的联系，从而对该领域产生变革性影响线粒体蛋白质相互作用网络和与年龄相关的线粒体功能障碍。 1