权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Regulation Of Intracellular Iron Metabolism

细胞内铁代谢的调节

基本信息

批准号：
9150088
负责人：
TRACEY A. ROUAULT
金额：
$ 123.11万
依托单位：
EUNICE KENNEDY SHRIVER NATIONAL INSTITUTE OF CHILD HEALTH & HUMAN DEVELOPMENT
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/9150088
关键词：
3&apos Untranslated Regions 5&apos Untranslated Regions Aconitate Hydratase Adult Affect Anemia Animal Model Animals Axonal Transport Beryllium Binding Blood Vessels Bone Marrow Transplantation Cancer cell line Cell Line Cells Cessation of life Disease Elements Embryo Endothelium Enzymes Erythroblasts Erythropoietin Fatty Acids Ferritin Fibroblasts Functional disorder Gait abnormality Gene Expression Glycogen Goals Growth H ferritin HIF1A gene Heme Human Indium Infusion procedures Inherited Iron Iron Regulatory Protein 1 Iron Regulatory Protein 2 Iron-Sulfur Proteins Kidney Knockout Mice Laboratories Lead Leiomyomatosis Liver Lung Maintenance Malignant Neoplasms Mammals Mediating Messenger RNA Metabolic Metabolism Metformin Mitochondria Molecular Motor Neurons Mucous Membrane Mus Neoplasm Metastasis Nerve Degeneration Neurologic Neurons Nutrient Oligodendroglia Oxygen Pharmaceutical Preparations Phenotype Physiological Physiology Polycythemia Production Protein Binding Protein p53 Proteins Pulmonary Hypertension RNA RNA Splicing Regulation Renal carcinoma Role SLC11A2 gene Spleen Staging Stimulus Sulfur Syndrome System Tissues Transcript Transferrin Receptor Translational Repression Translations Tumor Suppressor Proteins Work aerobic glycolysis axonal degeneration axonopathy base blastomere structure cancer cell cell type heme oxygenase-1 homologous recombination human subject insight interest interstitial iron deficiency iron metabolism loss of function mutation macrophage metal transporting protein 1 monocyte mouse model multicatalytic endopeptidase complex oxidative damage prevent progressive neurodegeneration protein function sensor small molecule stem tempol tumor metabolism tumor xenograft

项目摘要

This project aims to understand the molecular basis for regulation of intracellular iron metabolism. The cis and trans elements mediating the iron-dependent alterations in abundance of ferritin and the transferrin receptor have been identified and characterized in previous years in this laboratory. Iron- responsive elements (IREs) are RNA stem-loops found in the 5' end of ferritin mRNA and the 3' end of transferrin receptor mRNA. We have previously cloned, expressed, and characterized two essential iron- sensing proteins, Iron Regulatory Protein 1 (IRP1) and Iron Regulatory Protein 2 (IRP2). IRPs bind IREs when iron levels are depleted, resulting in the inhibition of translation of ferritin mRNA and other transcripts that contain an IRE in the 5' untranslated regions, or in stabilization of the transferrin receptor mRNA and possibly other transcripts that contain IREs in the 3'UTR. The IRE-binding activity of IRP1 depends on whether the protein contains an iron-sulfur cluster (see project HD008814-01). IRP2 also binds IREs in iron-depleted cells, but unlike IRP1, IRP2 is degraded in cells that are iron- replete. There are nine major IRE-containing mRNAs, and many have very important functions, such as the iron exporter, ferroportin, and the oxygen sensor, HIF2 alpha. We discovered that one alternatively spliced transcript of the iron exporter, ferroportin, lacks an IRE, and expression of the non-IRE form in duodenal mucosa and erythroblasts explains several important aspects of physiology. In iron-replete cells, IRP2 is selectively ubiquitinated by FBXL5 and degraded by the proteasome. To approach questions about the physiology of iron metabolism, loss of function mutations of IRP1 and IRP2 have been generated in mice through homologous recombination in embryonic cell lines. In the absence of provocative stimuli, there are subtle abnormalities in iron metabolism associated with loss of IRP1 function. IRP2-/- mice develop a progressive neurologic syndrome characterized by gait abnormalities and axonal degeneration. Ferritin over-expression occurs in affected neurons, and in protrusions of oligodendrocytes into the space created by axonal degeneration. We are culturing neurons from Irp null mice to better characterize their metabolic abnormalities. In animals that lack IRP1, IRP 2 compensates for loss of IRP1 regulatory activity, whereas animals that null for both IRP1 and IRP2 die as early embryos. The adult-onset neurodegeneration of adult IRP2-/- mice is exacerbated when one copy of IRP1 is also deleted. IRP2-/- mice offer a unique example of spontaneous adult-onset slowly progressive neurodegeneration, and analyses of gene expression and iron status at various stages of disease are ongoing. We have found that lower motor neurons are very adversely affected, developing axonopathy and death. In addition, small molecule treatment with the stable nitroxide, Tempol, prevents neurodegeneration in IRP2-/- animals. We characterized metabolism in an HLRCC cell line and discovered that AMPK is down, which leads to reduced p53 and DMT1, an iron importer. The iron deficiency that arises as a consequence promotes the switch to aerobic glycolysis. Only HIF1 alpha is significantly elevated, whereas HIF2 alpha expression is repressed by IRP activation. These metabolic changes lead to high storage of glycogen and fatty acids, which enables these cancer cells to store large amounts of energy that may fuel them during when they metastasize and temporarily lose access to nutrients. We discovered that treatment of cells with metformin in combination with an experimental drug that interferes with vascular growth eliminates growth of mouse xenograft tumors formed from the HLRCC cell line. We are also working to characterize Chuvash polycythemia in animal models and human subjects with particular emphasis on the underlying molecular pathophysiology. We discovered that Irp1-/- mice develop erythropoietin-driven polycythemia and pulmonary hypertension, and Irp1 is important for modulating expression of HIF2 alpha in pulmonary endothelia and renal interstitial fibroblasts.

本项目旨在了解细胞内铁代谢调控的分子基础。在过去的几年中，本实验室已经确定并表征了介导铁蛋白和转铁蛋白受体丰度的铁依赖性改变的顺式和反式元素。铁响应元件（IRE）是存在于铁蛋白mRNA的5'端和转铁蛋白受体mRNA的3'端的RNA茎环。我们先前已经克隆、表达并鉴定了两种必需的铁敏感蛋白，铁调节蛋白1（IRP 1）和铁调节蛋白2（IRP 2）。当铁水平耗尽时，IRP结合IRE，导致铁蛋白mRNA和其他在5'非翻译区含有IRE的转录物的翻译受到抑制，或转铁蛋白受体mRNA和可能的其他在3' UTR含有IRE的转录物稳定。IRP 1的IRE结合活性取决于蛋白质是否含有铁硫簇（见项目HD 008814 -01）。 IRP 2也结合铁耗尽细胞中的IRE，但与IRP 1不同，IRP 2在铁充足的细胞中被降解。有九种主要的含IRE的mRNA，其中许多具有非常重要的功能，例如铁输出蛋白ferroportin和氧传感器HIF 2 α。我们发现铁输出蛋白ferroportin的一种选择性剪接转录本缺乏IRE，并且在十二指肠粘膜和成红细胞中非IRE形式的表达解释了生理学的几个重要方面。在充满铁的细胞中，IRP 2被FBXL 5选择性地泛素化并被蛋白酶体降解。为了探讨铁代谢的生理学问题，通过胚胎细胞系中的同源重组，在小鼠中产生了IRP 1和IRP 2的功能丧失突变。在没有刺激的情况下，与IRP 1功能丧失相关的铁代谢存在细微异常。IRP 2-/-小鼠发生以步态异常和轴突变性为特征的进行性神经系统综合征。铁蛋白过度表达发生在受影响的神经元中，以及少突胶质细胞突起到轴突变性产生的空间中。我们正在培养Irp缺失小鼠的神经元，以更好地表征其代谢异常。在缺乏IRP 1的动物中，IRP 2补偿IRP 1调节活性的损失，而IRP 1和IRP 2都无效的动物作为早期胚胎死亡。当IRP 1的一个拷贝也被删除时，成年IRP 2-/-小鼠的成年发病神经变性会加剧。IRP 2-/-小鼠提供了一个独特的自发性成人发病缓慢进行性神经变性的例子，在疾病的各个阶段的基因表达和铁状态的分析正在进行中。我们发现，下运动神经元受到非常不利的影响，发展轴突病变和死亡。此外，用稳定的氮氧化物Tempol进行的小分子治疗可预防IRP 2-/-动物的神经变性。我们表征了HLRCC细胞系中的代谢，发现AMPK下降，这导致p53和DMT 1（铁进口商）减少。因此出现的铁缺乏促进了有氧糖酵解的转变。只有HIF 1 α显著升高，而HIF 2 α表达被IRP激活抑制。这些代谢变化导致糖原和脂肪酸的高储存，这使得这些癌细胞能够储存大量的能量，这些能量可以在它们转移并暂时失去营养时为它们提供燃料。我们发现，二甲双胍与一种干扰血管生长的实验性药物联合治疗细胞可消除HLRCC细胞系形成的小鼠异种移植肿瘤的生长。我们还在努力在动物模型和人类受试者中表征楚瓦什红细胞增多症，特别强调潜在的分子病理生理学。我们发现Irp 1-/-小鼠发生红细胞生成素驱动的红细胞增多症和肺动脉高压，Irp 1对调节肺内皮细胞和肾间质成纤维细胞中HIF 2 α的表达很重要。