Stablization of Fragile Human Transfer RNAs

脆弱人类转移 RNA 的稳定

基本信息

批准号：
9769070
负责人：
PAUL R SCHIMMEL
金额：
$ 38.7万
依托单位：
SCRIPPS RESEARCH INSTITUTE, THE
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-08-23 至 2022-06-30
项目状态：
已结题

项目摘要

Project Summary This proposal is aimed at understanding enough to eventually have a therapeutic intervention for a large group of human mitochondrial diseases associated with transfer RNAs. These diseases arise from mutations in mitochondrial tRNAs that cause instability and susceptibility to nuclease degradation. We focus on stabilizing these mutant tRNAs by identifying and utilizing a pan-specific mitochondrial tRNA binding protein that stabilize the fragile tRNA structure. We have found one such natural protein from the recently discovered ensemble of human tRNA synthetase splice variants. Because defects in mitochondrial (mt) protein synthesis have an immediate impact on cellular metabolism, it is perhaps not surprising that more than 50% of all identified disease-causing mutations in mtDNA are located within mtDNA genes for tRNAs. And yet, these genes constitute only 10% of the mitochondrial genome. A significant number of mt disease-linked mutations, which cause myopathies, neurodegenerative diseases, and multisystemic disorders, are located in the mt tRNAs. Our laboratory has published extensively on aaRSs from bacteria to humans. However, we did not previously have the insight to suggest a path to stabilize disease-causing mutant tRNAs. aaRSs have progressively acquired new domains in evolution. These new domains are dispensable for the aminoacylation function and are mobilized for specific, novel functions outside of translation. We discovered over 250 splice variants (SVs) of human aaRSs. The majority ablate the catalytic activity but retain the novel motifs. Importantly, most are stable as recombinant proteins. Amongst the SVs, we focus on the few that have the appearance of being chaperones for tRNAs, that is, proteins that bind to and stabilize tRNAs, but with a structure- but not sequence-specific recognition of the outside corner (elbow) of the L-shaped tRNA structure. The goal is to select one or more easy- to-purify, stable, recombinant corner- binding splice variants. These will be tested for their ability, when added in trans, to bind defective mutant mitochondrial tRNAs. Another criterion is that the chosen domain can be applied exogenously to cells and enter the mitochondria. In preliminary work we identified at least one SV that fulfills the criteria we established. If successful, this proposal will suggest a new path for therapeutic intervention of the most prevalent human mitochondrial diseases. !

项目摘要该建议旨在足够理解以最终进行治疗干预对于与转移RNA相关的大量人线粒体疾病。这些疾病由线粒体TRNA的突变引起，导致核酸酶降解的不稳定性和敏感性。我们专注于通过识别和利用泛特异性线粒体tRNA来稳定这些突变体TRNA 稳定脆弱tRNA结构的结合蛋白。我们从最近发现的人tRNA合成酶剪接变体的合奏。因为线粒体（MT）蛋白合成的缺陷对细胞有直接影响代谢，也许不足为奇的是，在所有鉴定出疾病的突变中超过50％在mtDNA中，位于mtDNA基因内的TRNA。然而，这些基因仅占10％线粒体基因组。大量MT疾病连接的突变引起肌病，神经退行性疾病和多系统疾病位于MT TRNA中。我们的实验室已广泛发表有关从细菌到人类的AARS的广泛发表。但是，我们没有以前有洞察力建议稳定引起疾病的突变体TRNA的途径。 AARS逐渐获得了进化的新领域。这些新领域是可用于氨基酰化函数的可分配，并动员特定的新功能。翻译。我们发现了人类AARS的250多个剪接变体（SV）。大多数消融催化活性，但保留了新型基序。重要的是，大多数作为重组蛋白稳定。在SVS中，我们专注于少数具有TRNA伴侣的伴侣的人， IS，与TRNA结合并稳定的蛋白质，但没有结构 - 但不是序列特异性识别 L形tRNA结构的外角（肘）。目标是选择一个或多个要纯化，稳定，重组角结合剪接变体。这些将对它们的能力进行测试，当在反式中添加，以结合有缺陷的突变体线粒体TRNA。另一个标准是选择域可以外源应用于细胞并进入线粒体。在初步工作中确定了至少一项符合我们确定标准的SV。如果成功，该提议将提出了最普遍的人类线粒体疾病治疗干预的新途径。呢