Frascati-mediated Mitochondrial Metabolism

弗拉斯卡蒂介导的线粒体代谢

• 批准号: 8205189
• 负责人: BARRY H PAW
• 金额: $ 35.68万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-15 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8205189
• 关键词: Anabolism; Anemia; Bioinformatics; Candidate Disease Gene; Cell Line; Cell physiology; Cells; Defect; Developmental Biology; Embryo; Endosomes; Epitopes; Erythrocytes; Erythroid; Erythroid Cells; Erythropoiesis; Fetal Liver; Follow-Up Studies; Gene Expression; Generations; Genes; Genetic; Goals; Heme; Heme Iron; Hemoglobin; Hepatocyte; Human; Instruction; Iron; Iron Metabolism Disorders; Knock-out; Liver; Mammalian Cell; Mammals; Mass Spectrum Analysis; Mediating; Messenger RNA; Metabolism; Mitochondria; Mitochondrial Membrane Protein; Mitochondrial Proteins; Mus; Organ; Orthologous Gene; Phenotype; Population; Primary Cell Cultures; Principal Investigator; Prosthesis; Proteins; Proteome; Role; Structural Genes; Structural Protein; Structure; Validation; Zebrafish; ferrochelatase; heme biosynthesis; insight; interest; loss of function; novel; protein complex; tool; trafficking

The objectives of this proposal are to identify and characterize novel proteins involved in mitochondrial iron/heme metabolism identified from differentiating fetal liver population by mRNA sequencing (RNAseq) and bioinformatics approaches. Heme serves as a prosthetic group in hemo-proteins for a wide array of crucial cellular processes, and of these, hemoglobin synthesis in red cells is the most well known. Despite advances in our understanding of cytosolic iron trafficking and proto-prophyrin biosynthesis, significant gaps remain, especially, with respect to components involving the egress of iron from the endosomes to the mitochondria, the trafficking of iron/heme within the mitochondria, and the eventual export of heme from the mitochondria for its incorporation in hemoglobin. In recent years, we have characterized the role ofthe l\/litoferrin1 (l\/lfrn1, Slc25a37) iron importer and its interaction with other mitochondrial proteins, AbcblO and ferrochelatase, in the acquisition of mitochondrial iron and its utilization in heme synthesis. In an attempt to dentify additional, unknown components important for heme synthesis, we screened thousands of microarrays for genes that were tightly co-expressed and co-regulated with previously known heme biosynthesis genes to identify potentially interesting, novel candidate genes. Follow up studies of these candidate genes in the zebrafish showed that gene-specific knockdown, using anti-sense morpholinos, resulted in profound anemia in all cases. In a complementary approach, we recently analyzed the expression of structural proteins with predicted transmembrane motifs, transporter function, or localization to the mitochondria, which were identified by RNAseq analysis from differentiating fetal liver cells. We identified 9 additional strongly induced genes, whose function in erythroid iron/heme metabolism has not been previously studied. We propose to study the expression and loss-of-function phenotype of these 9 structural genes (c20orf108, Snx3, Slc43a1, Slc43a3, Slc7a5, Ehbplll, Tmcc2, Slc38a5, Tmem14c) in the zebrafish and mammalian cells. In particular, we plan to focus on one of these newly identified genes from the two bioinformatics screens, Tmem14c, a small mitochondrial membrane protein of unknown function. RELEVANCE (See instructions): Elucidating the function of these 8 structural proteins, Tmem14c and its interacting protein partners may give insight into unknown steps in mitochondrial heme metabolism and provide new genetic tools for exploring human disorders of iron/heme metabolism and erythropoiesis.

这项提案的目标是识别和表征与线粒体有关的新蛋白质。 用mRNA测序(RNAseq)鉴定区分胎肝群体的铁/血红素代谢 和生物信息学方法。血红素是血液蛋白中的一个修饰性基团，具有广泛的 在这些关键的细胞过程中，红细胞中的血红蛋白合成是最广为人知的。尽管 细胞质铁转运和原卟啉生物合成的研究进展 保留，尤其是涉及铁从内体向外排出的成分 线粒体，铁/血红素在线粒体内的运输，以及最终从线粒体输出的血红素 线粒体为其与血红蛋白的结合。近年来，我们已经确定了 L Litoferrin1(L，SLC25a37)铁导入蛋白及其与其他线粒体蛋白的相互作用 铁络合酶，在线粒体铁的获取及其在血红素合成中的应用。为了试图 鉴定对血红素合成重要的其他未知成分，我们筛选了数千种 与已知的血红素紧密共表达和共同调控的基因微阵列 生物合成基因以确定潜在的有趣的、新的候选基因。对这些问题的后续研究 斑马鱼中的候选基因表明，使用反义吗啉， 在所有病例中都导致严重贫血。在一种补充方法中，我们最近分析了 具有预测跨膜基序、转运蛋白功能或定位的结构蛋白表达 经RNAseq分析鉴定为分化胎肝细胞的线粒体。我们 鉴定了另外9个强诱导基因，它们在红系铁/血红素代谢中的功能尚未见报道 已经被研究过了。我们建议研究这9个基因的表达和功能丧失表型。 结构基因(c20orf108、Snx3、Slc43a1、Slc43a3、Slc7a5、Ehbplll、Tmcc2、Slc38a5、Tem14c) 斑马鱼和哺乳动物细胞。特别是，我们计划将重点放在这些新发现的来自 这两个生物信息学筛选，Tem14c，一个功能未知的小线粒体膜蛋白。 相关性(请参阅说明)： 为了阐明这8种结构蛋白的功能，Tem14c及其相互作用的蛋白质伙伴可能会给出 深入了解线粒体血红素代谢的未知步骤，为探索提供新的遗传工具 人类铁/血红素代谢紊乱与红细胞生成。