Zhang Ping, Huang Junhong, Zheng Wujian, Chen Lifan, Liu Shurong, Liu Anrui, Ye Jiayi, Zhou Jie, Chen Zhirong, Huang Qiaojuan, Liu Shun, Zhou Keren, Qu Lianghu, Li Bin, Yang Jianhua
MOE Key Laboratory of Gene Function and Regulation, State Key Laboratory of Biocontrol, School of Life Sciences, The Fifth Affiliated Hospital, Sun Yat-sen University, Guangzhou, 510275, China.
Department of Chemistry, The University of Chicago, Chicago, IL, 60637, USA.
Sci China Life Sci. 2023 Apr;66(4):800-818. doi: 10.1007/s11427-022-2210-0. Epub 2022 Oct 28.
2'-O-methylation (Nm) is one of the most abundant RNA epigenetic modifications and plays a vital role in the post-transcriptional regulation of gene expression. Current Nm mapping approaches are normally limited to highly abundant RNAs and have significant technical hurdles in mRNAs or relatively rare non-coding RNAs (ncRNAs). Here, we developed a new method for enriching Nm sites by using RNA exoribonuclease and periodate oxidation reactivity to eliminate 2'-hydroxylated (2'-OH) nucleosides, coupled with sequencing (Nm-REP-seq). We revealed several novel classes of Nm-containing ncRNAs as well as mRNAs in humans, mice, and drosophila. We found that some novel Nm sites are present at fixed positions in different tRNAs and are potential substrates of fibrillarin (FBL) methyltransferase mediated by snoRNAs. Importantly, we discovered, for the first time, that Nm located at the 3'-end of various types of ncRNAs and fragments derived from them. Our approach precisely redefines the genome-wide distribution of Nm and provides new technologies for functional studies of Nm-mediated gene regulation.
2'-O-甲基化（Nm）是最丰富的RNA表观遗传修饰之一，在基因表达的转录后调控中起着至关重要的作用。目前的Nm定位方法通常仅限于高丰度RNA，在mRNA或相对罕见的非编码RNA（ncRNA）中存在重大技术障碍。在此，我们开发了一种新方法，利用RNA外切核糖核酸酶和高碘酸盐氧化反应性来消除2'-羟基化（2'-OH）核苷，结合测序（Nm-REP-seq）来富集Nm位点。我们在人类、小鼠和果蝇中揭示了几类新的含Nm的ncRNA以及mRNA。我们发现一些新的Nm位点存在于不同tRNA的固定位置，并且是由小核仁RNA（snoRNA）介导的纤维蛋白原（FBL）甲基转移酶的潜在底物。重要的是，我们首次发现Nm位于各种类型ncRNA及其衍生片段的3'末端。我们的方法精确地重新定义了全基因组范围内Nm的分布，并为Nm介导的基因调控功能研究提供了新技术。
