Molecular Genetics of Caloric Restriction in Aging Flies

描述（由申请人提供）：热量限制（CR）是延长动物寿命和延迟年龄相关症状发作的最可靠方法。在目前的资助期间，我们做出了重大贡献，我们的年龄和热量摄入对成年果蝇的生理和寿命的影响的理解。果蝇（Drosophila melanogaster）中Indy基因的突变显著延长了寿命。INDY是克雷布斯循环中间体的二羧酸转运蛋白，主要存在于对中间代谢重要的组织中。已提出Indy活性降低的寿命延长效应导致一种遗传CR形式，这一假设得到了在本资助期间进行的生物化学、分子和遗传研究的支持。Indy长寿果蝇基因组转录反应的测定揭示了代谢功能基因的下调，特别值得注意的是线粒体氧化磷酸化（OP）复合物I和III的组分表达的瞬时减少。我们发现，在印第蝇OP I和III复合物具有较低的酶活性，产生较少的活性氧（ROS），并引起较低的氧化损伤。然而，印第蝇中ATP的产生与对照相似，这一结果可以通过印第蝇中发现的线粒体密度增加来解释。考虑到线粒体在能量产生和细胞内稳态中的关键作用，我们的初步数据提供了代谢和长寿途径之间的额外联系，并形成了我们假设的基础，即OP复合物的瞬时变化介导了Indy突变果蝇的长寿。与此同时，我们已经表明，下调rpd 3组蛋白脱乙酰酶，或过表达的dSir 2组蛋白脱乙酰酶基因，或增加dSir 2活性喂养苍蝇白藜芦醇，延长寿命在果蝇的机制类似于CR。这使我们有机会确定线粒体生理学的类似变化是否是三种遗传CR果蝇模型中寿命延长的潜在途径的一部分。在本提案的目标1中，我们将确定Indy、rpd 3和Sir 2基因或CR的遗传操作是否通过下调OP I和III组分在每种寿命延长条件下的水平和活性来影响寿命。在目标2中，我们将确定降低复合物I和III的组分的水平是否对果蝇生理学和寿命有影响。在目标3中，我们将研究OP I和III组分与已建立的Indy/rpd 3/Sir 2长寿途径之间的遗传相互作用。在目标4中，我们将通过评估Indy/rpd 3/Sir 2长寿途径中的线粒体生理学和生物发生来进一步阐明线粒体在CR寿命延长中的作用。由于线粒体在能量稳态，应激反应和寿命中的作用是众所周知的，我们提出的实验将扩展现有的知识，OP组分在CR途径中的新作用，并可能提供治疗干预的基础。公共卫生相关性：热量限制已经成为保护脊椎动物和无脊椎动物物种免受衰老有害影响的最有效方法。本项目将研究限制热量摄入延长果蝇寿命的分子机制。它将揭示减少热量摄入如何影响新陈代谢和寿命，并为开发治疗人类年龄相关疾病的新疗法提供基础。

DESCRIPTION (provided by applicant): Caloric restriction (CR) is the surest way of increasing life span and delaying the onset of age-related symptoms in animals. During the current funding period, we made significant contributions to our understanding of the effects of age and caloric intake on physiology and longevity of adult Drosophila. Mutation in the Indy gene in the fruit fly, Drosophila melanogaster, dramatically extends life span. INDY is a dicarboxylate transporter of Krebs cycle intermediates primarily found in the tissues important for intermediary metabolism. The life extending effect of reduced Indy activity has been proposed to result in a form of genetic CR, a hypothesis supported by biochemical, molecular and genetic studies carried out during this funding period. Determination of the genomic transcriptional responses of Indy long-lived flies reveal down-regulation of genes that function in metabolism-particularly noteworthy is a transient decrease in the expression of components of the mitochondrial oxidative phosphorylation (OP) complexes I and III. We showed that in Indy flies OP I and III complex have lower enzyme activity, produced less reactive oxygen species (ROS), and caused lower oxidative damage. However, production of ATP in Indy flies is similar to the control, a result that could be explained by increased mitochondrial density found in Indy flies. Considering the crucial role of mitochondria in energy production and cellular homeostasis, our preliminary data provide additional links between metabolic and longevity pathways and form the basis for our hypothesis that transient change in the OP complexes mediate longevity in Indy mutant flies. In parallel, we have shown that down-regulation of the rpd3 histone deacetylase, or overexpression of dSir2 histone deacetylase genetically, or increasing dSir2 activity by feeding flies resveratrol, extends life span in Drosophila by a mechanism similar to CR. This give us an opportunity to determine if similar changes in mitochondrial physiology are part of the pathway underlying life span extension in three fly models of genetic CR. In aim 1 of this proposal, we will determine if genetic manipulations of Indy, rpd3, and Sir2 genes, or CR, effect longevity by downregulation of the levels and activity of the OP I and III components in each life span extending condition. In aim 2, we will determine if decreasing the levels of components of complex I and III have effects on fly physiology and longevity. In aim 3, we will examine genetic interactions between OP I and III components and the established Indy/rpd3/Sir2 longevity pathway. In aim 4, we will further elucidate the role mitochondria play in CR life span extension by assessing the mitochondrial physiology and biogenesis in the Indy/rpd3/Sir2 longevity pathway. Since the role of mitochondria in energy homeostasis, stress response and longevity is well known, our proposed experiments will extend current knowledge to the novel role of the OP components in the CR pathway and potentially provide a basis for therapeutic intervention. PUBLIC HEALTH RELEVANCE: Caloric restriction has emerged as the most efficient way to protect the organism against deleterious effects of aging in both vertebrate and invertebrate species. This project will study the molecular mechanism underlying life span extension in fruit flies, Drosophila melanogaster, by caloric restriction. It will reveal how reduced caloric intake affects metabolism and life span, and provide the foundation for the development of new therapies for the treatments of age-associated diseases in humans.