Metal Homeostatic Mechanisms in Yeast

酵母中的金属稳态机制

基本信息

批准号：
8067894
负责人：
Dennis R. Winge
金额：
$ 25.81万
依托单位：
UNIVERSITY OF UTAH
依托单位国家：
美国
项目类别：
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/8067894
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 Health 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 respiratory structural biology superoxide dismutase 1 trafficking transcription factor

项目摘要

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）：该提案提出了作为模型真核生物的酿酒酵母中新的线粒体-细胞质信号传导途径。在第一个线粒体-细胞质途径中，我们发现了一种不同于金属伴侣铜穿梭的铜运输新范式。该途径涉及铜配体 (CuL) 复合物的候选易位，该复合物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) 的金属化。除酵母外，CuL 复合物也是人成纤维细胞、小鼠肝脏和拟南芥中线粒体铜的重要来源。我们假设 Cu(I) 与细胞质中丰富的游离配体结合后，CuL 复合物易位到线粒体中是通过尚未识别的线粒体转运蛋白发生的。我们的三个主要目标是阐明 CuL 复合物的结构、验证其易位到线粒体基质中并确定转运蛋白。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，并结合细胞和体外线粒体输入研究来表征易位途径。我们将尝试通过基因筛选和对呼吸缺陷突变体库的评估来识别转运蛋白。第二条线粒体-细胞质途径与 Aft1 转录激活因子的铁感应有关。在铁充足的细胞中，线粒体输出蛋白 Atm1 发出的信号会抑制 Aft1 的转录激活。 Aft1 功能与线粒体铁状态相关，因为许多表现出线粒体铁积累的酵母突变体显示出持续活跃的 Aft1。我们的目的是确定在这些突变体中 Aft1 的铁抑制被阻断的机制。 Aft1 的铁感应依赖于两种谷氧还蛋白和两种新鉴定的蛋白质 Fra1 和 Fra2。我们的目标是阐明这些蛋白质如何介导 Aft1 的 Fe 抑制。人类细胞与 Atm1 和 Grx3/4 具有同源物，其中 Atm1 的人类直系同源物 ABCB7 在铁稳态中具有明显的作用。我们最近的研究表明存在可扩展的基质铁池。在铁硫簇生物合成受损的酵母突变体中，该铁库会扩大，并可能导致超氧化物歧化酶 2 (Sod2) 的金属错误和铁响应性转录激活剂 Aft1 的组成型激活。这项资助的统一主题是基质中存在生物可利用的铜和铁池，以及线粒体-细胞质串扰作为包括人类在内的真核生物铜和铁稳态的关键组成部分。线粒体和细胞核之间的串扰对许多代谢疾病和紊乱具有广泛的影响。公共卫生相关性：线粒体内铜配体复合物对细胞色素氧化酶和超氧化物歧化酶 1 的铜金属化与人体生理学相关。细胞色素氧化酶组装缺陷会导致人类呼吸窘迫，表现为新生儿心肌病。线粒体超氧化物歧化酶-1 突变体与家族性肌萎缩侧索硬化症的病因有关。已知人类 ABCB7 线粒体铁输出蛋白功能障碍会导致 X 连锁铁粒幼细胞贫血伴共济失调。