Modulation of Oxidative phosphorylation by mitochondrial soluble adenylyl cyclase

线粒体可溶性腺苷酸环化酶对氧化磷酸化的调节

基本信息

批准号：
8332758
负责人：
Giovanni Manfredi
金额：
$ 35.49万
依托单位：
WEILL MEDICAL COLL OF CORNELL UNIV
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-09-01 至 2015-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8332758
关键词：
Adenylate Cyclase Affect Amino Acids Basal metabolic rate Biochemical Biochemistry Carbon Dioxide Cells Characteristics Citric Acid Cycle Clinical Consumption Cyclic AMP Cyclic AMP-Dependent Protein Kinases Cytochrome-c Oxidase Deficiency Data Defect Disease Disease Progression Electron Transport Enzymes Excision Exercise Fatty acid glycerol esters Genetic Glucose Goals Health Heart Homeostasis Homozygote Intervention Investigation Knock-in Mouse Laboratories Life Metabolic Metabolic Pathway Metabolism Mitochondria Mitochondrial Diseases Mitochondrial Matrix Mitochondrial Proteins Modeling Molecular Mus Nutrient Outcome Oxidative Phosphorylation Pacemakers Pathway interactions Phosphorylation Physiological Production Protein Dephosphorylation Proteins Regulation Regulatory Pathway Running Signal Pathway Signal Transduction Site Skeletal Muscle Speed System Thermogenesis Tissues Transgenic Mice Work clinical phenotype coping cytochrome c oxidase human disease in vivo mitochondrial dysfunction mouse model mutant oxidation response sensor

项目摘要

DESCRIPTION (provided by applicant): Phosphorylation of mitochondrial proteins is a rapid and efficient way to regulate oxidative phosphorylation (OXPHOS) and maintain energy homeostasis. Mitochondrial PKA modulates enzymes of the electron transfer chain through protein phosphorylation, which is regulated by a signaling pathway, involving mitochondrial soluble adenylyl cyclase (sAC), generating cAMP that activates PKA. This signaling cascade serves as a metabolic sensor that modulates energy conversion in mitochondria. Cytochrome oxidase (COX) is a pacemaker of ETC fluxes, whose activity is modulated by phosphorylation of its subunits and by ATP allosteric inhibition. We showed that COX is a target of the signaling pathway and identified subunit IV of COX (COXIV) as a target for phosphorylation. We also demonstrated that phosphorylation of S58 in COXIV- 1 is responsible for modulation of COX activity. We propose that this regulatory pathway participates to metabolic adaptive responses to OXPHOS defects and could be a target for pharmacological intervention. The goal of this project is to understand the physiological implications of intramitochondrial sAC-cAMP- PKA signaling and S58 COXIV-1 phosphorylation, in vivo. We will understand how this system behaves in healthy tissues and in tissues affected by mitochondrial defects. To this end, we will generate and study mice, in which the molecular players of the regulatory pathway are genetically modified and assess the outcomes on clinical phenotype and mitochondrial biochemistry. In aim 1, we will generate transgenic mice expressing inducible sAC selectively targeted to the mitochondrial matrix (mito-sAC), which is expected produce high basal metabolism. In aim2, we will generate COXIV-1 S58A knockin mice, lacking the phosphorylated site, and thus incapable of up-regulating ETC fluxes and enhancing ATP production, resulting in exercise intolerance, decreased glucose utilization, impaired thermogenesis, and increased fat storage. In aim 3, we will investigate sAC-cAMP-PKA modulation in a mouse model of OXPHOS defect caused by conditional and inducible genetic disruption of COX. These mice recapitulate biochemical and clinical characteristics of mitochondrial diseases. We will assess how sAC-cAMP-PKA modulation and protein phosphorylation in mitochondria from affected tissues and correlate it with disease progression.

描述（由申请人提供）：线粒体蛋白的磷酸化是一种调节氧化磷酸化（OXPHOS）并维持能量稳态的快速有效方法。线粒体PKA通过蛋白质磷酸化调节电子转移链的酶，该磷酸化受信号传导途径调节，涉及线粒体可溶性腺苷酸环化酶（SAC），产生了激活PKA的cAMP。该信号级联反应是一种代谢传感器，可调节线粒体中的能量转换。细胞色素氧化酶（COX）是ETC通量的起搏器，其活性是通过其亚基的磷酸化和ATP变构抑制来调节的。我们表明Cox是信号通路的目标，并确定Cox（Coxiv）的亚基IV作为磷酸化的靶标。我们还证明，Coxiv-1中S58的磷酸化负责调节Cox活性。我们建议该调节途径参与对Oxphos缺陷的代谢自适应反应，并且可能是药理干预的目标。该项目的目的是在体内了解体内囊膜内囊 - cAMP-PKA信号传导和S58 coxiv-1磷酸化的生理意义。我们将了解该系统在健康组织和受线粒体缺陷影响的组织中的行为。为此，我们将生成和研究小鼠，其中调节途径的分子参与者经过遗传修饰，并评估临床表型和线粒体生物化学的结果。在AIM 1中，我们将产生表达诱导囊的转基因小鼠选择性地靶向线粒体基质（Mito-SAC），这预计会产生高基础代谢。在AIM2中，我们将产生Coxiv-1 S58A敲击蛋白小鼠，缺乏磷酸化的位点，因此无法上调ETC的ETC磁通量并增强ATP产生，从而导致运动不耐症，葡萄糖利用率降低，热生成受损和脂肪存储增加。在AIM 3中，我们将研究由COX的条件和诱导遗传破坏引起的OXPHOS缺陷的小鼠模型中的SAC-camp-PKA调制。这些小鼠概括了线粒体疾病的生化和临床特征。我们将评估SAC-PKA的调节和蛋白质的磷酸化如何与受影响组织的线粒体中的线粒体中，并将其与疾病进展相关联。