RNA trafficking in mitochondria

线粒体是大多数真核细胞所必需的细胞器。它们提供不同的代谢功能 包括生产能源和用于生物合成过程的专门代谢物。装配，装配 由线粒体和核基因组编码的功能性线粒体、蛋白质和RNA是 必填项。在人类中，至少有1000种核编码蛋白被输入线粒体，而只有13种 电子传输链的蛋白质由线粒体基因编码。这几个字的翻译 必需的线粒体编码蛋白需要线粒体核糖体、翻译因子和转移 RNA，其子集必须从细胞质输入到线粒体基质中。最近的研究 进化上不同的生物已经证明，多个RNA在细胞核内编码并输入 从胞浆进入线粒体。我们对核线粒体输入途径的总体理解 编码的蛋白质非常详细。相比之下，人们对线粒体导入机制知之甚少。 对于核编码的RNA，或者如何利用这一新知识将特定的RNA传递到线粒体。 我们的竞争性R01续订申请的修订总体目标是剖析主要和 可能只有RNA进入线粒体的途径，才能开发针对核编码的策略 报告结构中的RNA到线粒体的作用以及用于治疗由基因突变引起的疾病 线粒体DNA(MtDNA)在单独的研究中，我们将确定导入到 使用下一代RNA-Seq和耦合生物信息学方法的线粒体。此前，我们报道了 研究表明，RNA加工酶多核苷酸磷酸化酶(PNPase)竟然 定位于线粒体膜间间隙(IMS)，在那里它起着核守门人的作用 从胞浆中导入的编码RNA。有趣的是，PNPase只存在于 例如苍蝇、蠕虫和哺乳动物，这表明它的功能仅限于高等真核生物。 为了实现我们修改后的提案目标，以保留项目的原始本质，我们有 确定了两个具体的研究目标。在AIM 1中，PNPase以外的额外管制内容用于进口 核编码的RNA将被识别。在目标2中，报告和纠正性核编码的RNA结构将 被设计成确定RNA导入机制的RNA序列和结构要求以及 开发治疗线粒体DNA突变引起的疾病的策略。对于这些研究，我们将采取 我们已经建立了不同的模型系统来操纵PNPase的水平和活动。 除了增加我们对RNA导入机制的基础知识之外 线粒体和学习操纵这一RNA输入途径的规则，我们的更新应用程序可能 对公共健康有广泛的影响，因为我们的方法可以提供一种方法来改善或可能 治愈数十种由线粒体DNA突变引起的人类疾病。

Mitochondria are essential organelles for most eukaryotic cells. They provide diverse metabolic functions including the production of energy and specialized metabolites for biosynthetic processes. To assemble functional mitochondria, proteins and RNAs encoded by both the mitochondrial and nuclear genomes are required. In humans, at least 1,000 nuclear encoded proteins are imported into mitochondria, whereas only 13 proteins of the electron transport chain are encoded by mitochondrial genes. The translation of these few but essential mitochondrial encoded proteins requires mitochondrial ribosomes, translation factors, and transfer RNAs, a subset of which must be imported from the cytosol into the mitochondrial matrix. Recent studies in evolutionarily diverse organisms have shown that multiple RNAs are encoded within the nucleus and imported from the cytosol into mitochondria. Our overall understanding of mitochondrial import pathways for nuclear encoded proteins is quite detailed. By contrast, very little is known about the mitochondrial import mechanism for nuclear encoded RNAs or how this new knowledge can be used to deliver specific RNAs into mitochondria. The revised overall goals of our competitive R01 renewal application are to dissect the main and perhaps only pathway of RNA import into mitochondria and to develop strategies that target nuclear encoded RNAs to mitochondria in reporter constructs and for the treatment of diseases caused by mutations of mitochondrial DNA (mtDNA). In separate studies, we will identify the cohort of RNAs that are imported into mitochondria using next generation RNA-Seq and coupled bioinformatics approaches. Previously, we reported studies that showed that the RNA processing enzyme polynucleotide phosphorylase (PNPase) is unexpectedly localized to the mitochondrial intermembrane space (IMS), where it functions as a gatekeeper for nuclear encoded RNAs that are imported from the cytosol. Interestingly, PNPase is only present in the genomes of organisms such as flies, worms, and mammals, suggesting that its functions are limited to higher eukaryotes. To accomplish our revised proposal goals that retain the original essence of the project, we have identified two specific study aims. In Aim 1, additional regulatory components beyond PNPase for the import of nuclear encoded RNAs will be identified. In Aim 2, reporter and corrective nuclear encoded RNA constructs will be engineered to determine the RNA sequence and structural requirements of the RNA import machinery and to develop strategies to treat diseases caused by mutations in mtDNA. For these studies we will take advantage of different model systems that we have established to manipulate PNPase levels and activities. In addition to increasing our fundamental knowledge about the mechanisms of RNA import into mitochondria and learning the rules for manipulating this RNA import pathway, our renewal application may have a broad impact on public health because our approach could provide a method to ameliorate or possibly cure dozens of human diseases caused by mtDNA mutations.