Import of RNA into Mitochondria

将 RNA 导入线粒体

• 批准号: 6603457
• 负责人: MARK E SCHMITT
• 金额: $ 25.54万
• 依托单位: UPSTATE MEDICAL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-07-01 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6603457
• 关键词: RNA; RNA biosynthesis; Saccharomyces cerevisiae; binding proteins; endoribonucleases; gene mutation; intracellular transport; membrane potentials; mitochondria; mitochondrial DNA; nucleic acid quantitation /detection; nucleic acid sequence; ribonuclease P; ribonucleoproteins

Title: Import of RNA into Mitochondria. The main goal of the research outlined in this proposal is to understand how an RNA molecule is imported into the mitochondrial compartment and to identify the machinery involved in this process. As a model system we have been using the Saccharomyces cerevisiae RNase MRP, a ribonucleoprotein endoribonuclease that has at least two roles in cellular RNA processing, one in the mitochondrial compartment where it plays a direct role in the initiation of mitochondrial DNA replication and a second in the nucleolus. The specific objective for this proposal is to understand the mechanism and pathway by which the RNA subunit of the ribonucleoprotein enzyme RNase MRP is imported into mitochondria. Loss of mitochondrial DNA through mutation, deletion or depletion has been implicated in heart disease, diabetes, muscle myopathies, and more recently aging. Understanding the role of RNase MRP in mitochondrial DNA replication and function may provide better treatments for patients with these diseases. Understanding how RNAs are imported into mitochondria would provide a mode of gene therapy for people with mitochondrial DNA mutations without the need to transform mitochondria. Experimentation is based on the rationale that the genes for the RNA component of RNase MRP is nuclear encoded; hence, the MRP RNA needs to be transported from its site of transcription in the nucleus to mitochondria and be assembled with its cognate proteins. The RNase MRP enzyme in the yeast offers an ideal system to study RNA import into the mitochondrion using both biochemical and genetic methodologies. The specific aims enumerated below are designed to use both biochemistry and genetics to identify the mechanism and machinery of RNA import into mitochondria. Specific aims will include. 1) Use of a newly developed in vitro RNA import system to elucidate the biochemical and molecular requirements for RNA import. 2) Isolation of mutations in the MRP RNA that are specific for mitochondrial import to determine the sequences required for mitochondrial substrate binding, mitochondrial specific protein binding and import into the mitochondrial matrix. 3) Characterization of these mutants for various aspects of mitochondrial metabolism including loss of mitochondrial respiration, DNA maintenance, translation, processing of RNAs and import of the mutant MRP RNA in our in vitro system. 4) Identification of other RNA molecules imported into mitochondria. These experiments should provide a wealth of knowledge about this fundamental process.

标题：将RNA导入线粒体。 该提案中概述的研究的主要目标是了解RNA分子如何输入线粒体区室，并确定参与这一过程的机制。 作为一个模型系统，我们一直在使用酿酒酵母RNase MRP，核糖核蛋白内切核糖核酸酶，在细胞RNA加工中至少有两个作用，一个在线粒体区室中，它在启动线粒体DNA复制中发挥直接作用，第二个在核仁中。 该提案的具体目标是了解核糖核蛋白酶RNase MRP的RNA亚基导入线粒体的机制和途径。 线粒体DNA的突变、缺失或耗竭导致的缺失与心脏病、糖尿病、肌肉肌病以及最近的衰老有关。 了解RNase MRP在线粒体DNA复制和功能中的作用可能为这些疾病患者提供更好的治疗。 了解RNA如何导入线粒体将为线粒体DNA突变的人提供一种基因治疗模式，而无需转化线粒体。实验基于RNase MRP的RNA组分的基因是核编码的基本原理;因此，MRP RNA需要从其在核中的转录位点转运到线粒体，并与其同源蛋白组装。 酵母中的RNase MRP酶提供了一个理想的系统，使用生物化学和遗传学方法研究RNA输入到酵母中。 下面列举的具体目标旨在利用生物化学和遗传学来确定RNA输入线粒体的机制和机制。具体目标包括。 1)使用新开发的体外RNA导入系统来阐明RNA导入的生物化学和分子要求。2)分离MRP RNA中对线粒体输入具有特异性的突变，以确定线粒体底物结合、线粒体特异性蛋白结合和输入线粒体基质所需的序列。3)对这些突变体的线粒体代谢各个方面进行表征，包括线粒体呼吸丧失、DNA维持、翻译、RNA加工以及在我们的体外系统中导入突变MRP RNA。4)识别其他导入线粒体的RNA分子。 这些实验应该提供了关于这个基本过程的丰富知识。