tRNA片段及RNA磷硫酰化修饰在细胞应激反应中的动态变化规律研究

细胞内tRNA分子的拷贝数、修饰类型、衍生片段的动态变化，调控着细胞诸多代谢过程。然而，tRNA分子的高度修饰等特性，导致现有技术无法对tRNA和其衍生片段进行精确定量，导致目前关于tRNA的调控功能这一关键科学问题知之甚少。我们课题组开发了一种新型RNA定量技术AQRNA-seq，首次实现了对细胞内所有RNA分子的高通量绝对定量。本项目将利用AQRNA-seq: 一方面，绘制不同等级的原核和真核细胞内tRNA组的精确图谱，尤其在细胞应激反应过程中产生的新型tRNA衍生片段，探索tRNA组的代谢图谱和调控网络；另一方面，深入研究最新发现的RNA磷硫酰化修饰，对不同RNA分子上的该修饰进行精确定量、定位及动态变化分析。本项目将为tRNA组学、RNA磷硫酰化修饰等研究建立通用的精确定量技术平台，并揭示两者在细胞内的动态变化规律，为进一步研究其生理功能、寻找新型分子靶点等提供指导方向。

Emerging evidence shows that tRNA abundance, tRNA modifications and tRNA-derived fragments (tRFs) perform regulatory functions in numerous metabolic and cellular processes in both prokaryotes and eukaryotes. Despite their biological importance, tRNA and tRFs have not been accurately quantified due to their abundant modifications, rigid secondary structure and high sequence similarity. The lack of high-throughput technique for tRNA quantification has hindered our understanding of tRNA dynamics and their biological functions. Our group has developed a novel RNA sequencing technology, AQRNA-seq, which enables absolute quantification of the copy number of all RNA species in cell. In this project, we will apply AQRNA-seq technology to quantitatively analyse the dynamics of tRNA pool in both prokaryotes and eukaryotes, especially focusing on its dynamic changes and novel tRNA-derived fragments during cell stress response; Meanwhile, we will also apply AQRNA-seq to quantitatively profile RNA phosphorothioate (PT) modification, a novel RNA modification discovered in both prokaryotes and eukaryotes. This project will provide a universal approach for absolute quantification of tRNA pools and RNA PT modification in all organisms and, meanwhile, provide insights of their regulatory mechanisms of cellular stress response.