线粒体tRNAThr及苏氨酰-tRNA合成酶基因突变导致线粒体脑肌病的机理研究

Mitochondrial translation apparatus regulates mitochondrial metabolism and various key cellular pathways. As core components of mitochondrial translation machinery, tRNA and aminoacyl-tRNA synthetases directly control mitochondrial functions and play a critical role. Clinical studies have established that 6 mutations in mitochondrial tRNAThr genes cause 5 human mitochondrial encephalomyopathies; meanwhile, a P282L mutation in mitochondrial threonyl-tRNA synthetase (ThrRS) leads to mitochondrial encephalomyopathy. However, the detailed pathogenic mechanism needs to be explored. This project will compare the interaction of native tRNAThr and its mutants with various proteins to study influence of these mutations on formation, maturation, modification, aminoacylation and affinity with elongation factor on mitochondrial tRNAThr; based on recent progress, study on the biological function of newly identified isoform of ThrRS, SV; the impact of the P282L mutation on the protein structure, its interaction with other proteins and enzymatic activities; try to resolve crystal structure of ThrRS-tRNAThr complex to further understand the interaction between eukaryotic/mitochondrial ThrRS and its cognate tRNA and the influence of the various disease-causing mutations on synthesis of Thr-tRNAThr. These studies will uncover the pathogenic mechanism of tRNAThr and ThrRS gene mutations and provide basis to diagnose and give treatment for the relative disorders.

线粒体翻译系统控制线粒体代谢与重大细胞生命活动。tRNA及氨基酰-tRNA合成酶是线粒体翻译机器的核心分子，直接决定线粒体正常功能的发挥，具有重要的生物学意义。临床研究确认，人线粒体tRNAThr基因6种突变造成5种疾病；线粒体苏氨酰-tRNA合成酶(ThrRS) P282L突变导致线粒体脑肌病。但致病机制尚不明确。本项目将从分子水平上比较线粒体tRNAThr及其突变体与相关蛋白质的相互作用，研究各突变对于tRNAThr生命周期中多步骤的影响；在前期研究基础上，阐释最新发现的ThrRS同源异形体SV的功能、P282L突变对蛋白质间相互作用及酶活力影响的机制；解析ThrRS-tRNAThr共晶结构，在原子水平上揭示真核类ThrRS识别底物的方式及突变点对形成Thr-tRNAThr的影响。这些研究将加深人们对tRNA和ThrRS基因突变导致线粒体疾病的机理认识、为该类疾病的诊治提供理论基础。

tRNA及氨基酰-tRNA合成酶(aaRS)是线粒体翻译机器的核心分子，直接决定线粒体正常功能的发挥，具有重要的生物学意义。人线粒体tRNAThr (hmtRNAThr)及苏氨酰-tRNA合成酶(ThrRS)突变导致多种人类疾病，但致病机制尚不清楚。在本面上项目资助下，我们系统揭示人线粒体tRNAThr致病点突变对于tRNA生命周期多个步骤，包括tRNA折叠、加工、成熟以及功能的影响；针对线粒体ThrRS的研究首次揭示亚硝基化修饰调控哺乳动物蛋白质合成的速度与精确性。此外，针对人翻译系统的若干tRNA相互作用蛋白质，获得了一系列重要创新性研究进展，包括：发现新的导致耳聋、智障与脑白质病变的赖氨酰-tRNA合成酶(KARS)基因突变；发现线粒体翻译质量控制对于胚胎发育的重要性；发现一种新的aaRS基因；揭示人线粒体丙氨酰-tRNA合成酶(hmtAlaRS)识别tRNA的独特机制；发现哺乳动物细胞质多aaRS复合物新成员；揭示KARS遗传变异导致听力障碍的分子基础；揭示人线粒体tRNA t6A修饰的分子机制等成果。这些研究结果系统揭示了人细胞质与线粒体蛋白质合成及其导致相关疾病的分子基础。共发表研究论文9篇，包括Nucleic Acids Research 7篇；Human Mutation以及SCIENCE CHINA Life Sciences各1篇。项目主持人晋升研究员职称、晋升中科院分子细胞科学卓越创新中心/生化与细胞所研究组长、获得基金委优青项目资助；培养博士研究生3名。圆满并超额完成目标任务。