Nm-Mut-seq: a base-resolution quantitative method for mapping transcriptome-wide 2′-O-methylation

Li Chen, Li-Sheng Zhang, Chang Ye, Huiqing Zhou, Bei Liu, Boyang Gao, Zixin Deng, Changming Zhao, Chuan He & Bryan C. Dickinson

Cell Research (2023)

https://www.nature.com/articles/s41422-023-00836-w

Abstract

2′-O-methylation (Nm) (Fig. 1a) is a prevalent post-transcriptional RNA modification present in many cellular RNAs and plays critical roles in modulating both physical properties and functions of eukaryotic RNA. Studies of Nm modifications in RNA have long been hampered by a lack of effective mapping methods. Previously reported approaches can work well for detecting Nm modifications on abundant RNAs,1,2,3,4,5,6,7 but face challenges when applied to less abundant RNAs such as mRNA, lack stoichiometric information, and suffer from RNA sample degradation due to chemical treatment. Here, we present Nm-Mut-seq, a mutation signature-based Nm mapping method, which uses a custom reverse transcriptase (RT) that installs mutations at Am-, Cm-, and Gm-modified sites (Um is undetectable by this method). Our work provides a much-needed approach to detect Nm at base resolution in low abundant RNAs and to estimate the stoichiometry of each modified site transcriptome-widely.