MITOCHONDRIAL IRON METABOLISM IN AGING

衰老过程中的线粒体铁代谢

• 批准号: 6207160
• 负责人: KATHLEEN S CROWLEY
• 金额: $ 3.92万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-04-01 至 2000-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6207160
• 关键词: adenosinetriphosphatase; aging; cell cycle; cell death; cell line; cell senescence; enzyme activity; flow cytometry; fluorimetry; free radical oxygen; immunologic assay /test; iron metabolism; laboratory rabbit; mitochondria; nutrition of aging; nutrition related tag; oxidative stress; protein localization; transfection; yeasts

The long-term goals of this work is to understand the role of mitochondrial iron metabolism in regulating life span and cell death. Iron is an essential nutrient, however, in the presence of naturally produced reactive oxygen intermediates, iron augments free radical formation. Free radicals are suspected of causing cellular damage, for example, DNA fragmentation associated with aging, and high cellular iron concentration and sensitivity to the oxidizing potential of iron is associated with cell death observed in post- ischemic reperfusion injury and Friedreich's ataxia. In the cell, mitochondria are the major site of iron utilization and the major site of reactive oxygen intermediate formation implying that the ability of the mitochondria to manage iron may be extremely important to cell viability. Disruption of the gene for a previously undefined yeast traffic ATPase (mtATPase) that is suspected of regulating mitochondrial iron uptake and/or metabolism based on preliminary results, was shown to cause morphological defects similar to those observed during cell senescence. The overall aim of the proposed work is to confirm that this traffic ATPase is a mitochondrial iron uptake protein and determine if this traffick ATPase affects the overall cell cycle, as suspected based on preliminary data. Specifically the aims are: 1) to verify that the traffick ATPase is disrupted in a previously constructed mutant cell line and prove that a decrease in mtATPase expression causes the mutant phenotype by introducing an ATPase expression vector into mutant cells, 2) to prove that mtATPase is involved in mitochondrial iron metabolism by determining iron uptake and iron management activity of mtATPase disruption mutants and mtATPase over-expressing strains, and to verify that mtATPase is a mitochondrial protein by localizing the protein in situ using immunoassays, and 3) to prove that the mtATPase-1 cell line has an altered mitochondrial content and an altered cell cycle and life span by cytometry, fluorometry, and gene expression analysis. Determining the underlying mechanisms for mitochondrial iron handling is essential for future studies aimed at defining the role of iron aging, and for better understanding diseases associated with free radical damage and maladaptive iron metabolism. Since preliminary data indicates that a mammalian mtATPase homologue exists, a long term consequence of this work may be discovery of more effective treatment for iron related disorders and diseases.

这项工作的长期目标是了解线粒体铁代谢在调节寿命和细胞死亡中的作用。 但是，在天然产生的活性氧中间体存在下，铁是一种必不可少的营养素，铁增强了自由基的形成。 自由基怀疑会引起细胞损伤，例如，与衰老相关的DNA碎片，高细胞铁浓度和对铁的氧化潜力的敏感性与在缺血后再灌注损伤和弗里德雷希的共济失调中观察到的细胞死亡有关。 在细胞中，线粒体是铁利用的主要部位，是反应性氧中间形成的主要部位，这意味着线粒体管理铁的能力可能对细胞生存能力非常重要。 基于先前未定义的酵母流量ATPase（MTATPase）的破坏，该基因涉嫌根据初步结果调节线粒体铁的摄取和/或代谢，被证明会导致与细胞衰老过程中观察到的类似的形态缺陷。 拟议工作的总体目的是确认该流量ATPase是一种线粒体铁摄取蛋白，并确定此运输ATPase是否会影响总体细胞周期，这是根据初步数据所怀疑的。 Specifically the aims are: 1) to verify that the traffick ATPase is disrupted in a previously constructed mutant cell line and prove that a decrease in mtATPase expression causes the mutant phenotype by introducing an ATPase expression vector into mutant cells, 2) to prove that mtATPase is involved in mitochondrial iron metabolism by determining iron uptake and iron management activity of mtATPase disruption mutants and MTATPase过表达的菌株，并通过使用免疫测定定位将蛋白定位到原位，以及3）来证明MTATPase-1细胞系具有改变的线粒体含量，细胞周期和细胞范围改变的细胞周期和生命分析，并证明MTATPase是一种线粒体蛋白。 确定线粒体铁处理的基本机制对于旨在定义铁衰老的作用的未来研究至关重要，并更好地理解与自由基损害和不良适应性铁代谢相关的疾病。 由于初步数据表明存在哺乳动物MTATPase同源物，因此这项工作的长期后果可能是发现针对铁相关疾病和疾病的更有效治疗。