Metal Homeostatic Mechanisms in Yeast

酵母中的金属稳态机制

• 批准号: 7663956
• 负责人: Dennis R. Winge
• 金额: $ 26.34万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-24 至 2012-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7663956
• 关键词: ATP phosphohydrolase; Abbreviations; Address; Affect; Aminopeptidase P; Arabidopsis; Ataxia; Binding; Bioavailable; Biogenesis; Cardiomyopathies; Cell Nucleus; Cell physiology; Cells; Chelating Agents; Complex; Copper; Coupled; Cuprozinc Superoxide Dismutase; Cytoplasm; Disease; Enzymes; Etiology; Eukaryota; Evaluation; Exhibits; Familial Amyotrophic Lateral Sclerosis; Family; Fibroblasts; Functional disorder; Genes; Genetic Screening; Golgi Apparatus; Grant; High Pressure Liquid Chromatography; Homeostasis; Homologous Gene; Human; Immunoprecipitation; In Vitro; Inner mitochondrial membrane; Ions; Iron; Lead; Letters; Ligands; Link; Liver; Manganese Superoxide Dismutase; Mediating; Membrane; Metabolic Diseases; Metals; Mitochondria; Mitochondrial Matrix; Modeling; Mus; Neonatal; Nomenclature; Nuclear; Orthologous Gene; Outer Mitochondrial Membrane; Pathway interactions; Phase; Physiology; Printing; Process; Proteins; Reaction; Reading; Respiratory distress; Role; Saccharomyces cerevisiae; Sideroblastic Anemia; Signal Pathway; Signal Transduction; Source; Specificity; Structure; Sulfur; Transcription Coactivator; Transcriptional Activation; Vesicle; Yeasts; bathocuproine sulfonate; bathophenanthroline; cofactor; cytochrome c oxidase; frataxin; glutaredoxin; human SOD2 protein; mutant; novel; public health relevance; respiratory; structural biology; superoxide dismutase 1; trafficking; transcription factor

DESCRIPTION (provided by applicant): This proposal addresses novel mitochondrial-cytoplasm signaling pathways in Saccharomyces cerevisiae as a model eukaryote. In the first mitochondrial-cytoplasm pathway, we discovered a new paradigm in copper trafficking distinct from metallochaperone copper shuttling. This pathway involves the candidate translocation of a copper-ligand (CuL) complex used in metallation of cytochrome c oxidase (CcO) and superoxide dismutase-1 (Sod1) within the mitochondria. In addition to yeast, the CuL complex is a significant source of mitochondrial copper in human fibroblasts, mouse liver and Arabidopsis. We postulate that translocation of the CuL complex into the mitochondria occurs through a yet unidentified mitochondrial transporter after Cu(I) is bound to an abundant free ligand in the cytoplasm. We aim to elucidate the structure of the CuL complex, verify its translocation into the mitochondrial matrix and identify the transporters as three major objectives. Studies in these objectives will involve analytical and structural biology approaches to deduce the ligand structure and a combination of cellular and in vitro mitochondrial import studies to characterize the translocation pathway. We will attempt to identify the transporters through a genetic screen and the evaluation of a bank of respiratory deficient mutants. A second mitochondrial-cytoplasm pathway relates to iron sensing by the Aft1 transcriptional activator. Transcriptional activation by Aft1 is inhibited in iron-replete cells by a signal emanating from the mitochondrial exporter Atm1. Aft1 function is coupled to the mitochondrial iron status, as a number of yeast mutants exhibiting mitochondrial iron accumulation show constitutively active Aft1. We aim to determine the mechanism by which iron-inhibition of Aft1 is blocked in these mutants. Iron sensing by Aft1 is dependent on two glutaredoxins and two newly identified proteins Fra1 and Fra2. We aim to elucidate how these proteins mediate Fe-inhibition of Aft1. Human cells have homologs to Atm1 and Grx3/4 with the human ortholog of Atm1, ABCB7, having a clear role in iron homeostasis. Our recent studies suggest that an expandable matrix iron pool exists. This iron pool is expanded in yeast mutants impaired in iron-sulfur cluster biogenesis and can lead to mis-metallation of superoxide dismutase-2 (Sod2) and constitutive activation of the iron-responsive transcriptional activator Aft1. The unifying themes in this grant are the presence of bioavailable pools of copper and iron within the matrix and mitochondrial-cytoplasm cross talk as a key component in copper and iron homeostasis in eukaryotes including humans. Cross talk between mitochondria and the nucleus has wide implications in a number of metabolic diseases and disorders. PUBLIC HEALTH RELEVANCE: Copper metallation of cytochrome oxidase and superoxide dismutase-1 by the copper- ligand complex within the mitochondria is relevant in human physiology. Deficiencies in cytochrome oxidase assembly lead to respiratory distress in humans presenting as neonatal cardiomyopathies. Mutants of the mitochondrial superoxide dismutase-1 are implicated in the etiology of familial amyotrophic lateral sclerosis. Dysfunction in the human ABCB7 mitochondrial iron exporter is known to cause X-linked sideroblastic anemia with ataxia.

描述（由申请人提供）：该提案介绍了酿酒酵母中新型的线粒体 - 细胞质信号通路作为模型真核生物。在第一个线粒体 - 细胞质途径中，我们在铜运输中发现了一种新的范式，与金属伴侣铜班车不同。该途径涉及用于线粒体内细胞色素C氧化酶（CCO）和超氧化物歧化酶1（SOD1）的铜 - 配体（CUL）复合物的候选易位。除酵母外，CUL复合物是人成纤维细胞，小鼠肝脏和拟南芥中线粒体铜的重要来源。我们假设CuL络合物进入线粒体的易位是通过Cu（i）结合到细胞质中丰富的自由配体后的尚未确定的线粒体转运蛋白发生的。我们旨在阐明CUL复合物的结构，验证其易位到线粒体基质中，并将转运蛋白识别为三个主要目标。这些目标的研究将涉及分析和结构生物学方法来推断配体结构以及细胞和体外线粒体进口研究的结合，以表征易位途径。我们将尝试通过遗传筛查和评估呼吸不足突变体的评估。第二个线粒体 - 细胞质途径与AFT1转录激活剂的铁感应有关。 AFT1的转录激活通过线粒体出口商ATM1散发的信号在铁恢复细胞中抑制。 AFT1功能与线粒体铁的状态耦合，因为表现出线粒体铁积累的许多酵母突变体显示了组成性活性Aft1。我们旨在确定这些突变体中阻断AFT1的铁抑制的机制。 AFT1的铁感应取决于两种谷歌蛋白和两种新鉴定的蛋白FRA1和FRA2。我们旨在阐明这些蛋白如何介导FE抑制AFT1。人类细胞与ATM1和GRX3/4的同源物与ATM1的人类直系同源物ABCB7具有明显的作用。我们最近的研究表明，存在可扩展的基质铁池。该铁池在铁硫簇生物发生下损害的酵母突变体中扩展，并可能导致超氧化物歧化酶2（SOD2）和构型激活的铁反应转录激活剂AFT1。这笔赠款中的统一主题是在基质和线粒体 - 细胞质中存在可生物利用的铜和铁池，这是铜和铁稳态的关键组成部分，以及包括人类在内的真核生物中。线粒体与细胞核之间的串扰对多种代谢疾病和疾病具有广泛的影响。公共卫生相关性：通过线粒体内的铜配合体对细胞色素氧化酶和超氧化物歧化酶1的铜金属化在人类生理学中是相关的。细胞色素氧化酶组装的缺陷导致人类作为新生儿心肌病的呼吸窘迫。线粒体超氧化物歧化酶1的突变体与家族性肌萎缩性侧硬化症的病因有关。已知人类ABCB7线粒体导出蛋白的功能障碍会导致共济失调引起X连锁的sideroblastic贫血。