Quantitative sequencing using BID-seq uncovers abundant pseudouridines in mammalian mRNA at base resolution

• DOI: 10.1038/s41587-022-01505-w
• Publication Date: 2022-10-27T16:06:35Z
• Authors: Qing Dai; Li-Sheng Zhang; Hui-Lung Sun; Kinga Pajdzik; Lei Yang; Chang Ye; Cheng-Wei Ju; Shun Liu; Yuru Wang; Zhong Zheng; Linda Zhang; Bryan T. Harada; Xiaoyang Dou; Iryna Irkliyenko; Xinran Feng; Wen Zhang; Tao Pan; Chuan He
• Keywords: Mammals; Base Composition; 0303 health sciences; Sequence Analysis, RNA; Article; Mice; 03 medical and health sciences; Animals; Humans; RNA; Sulfites; RNA, Messenger; RNA Processing, Post-Transcriptional; Pseudouridine

AbstractFunctional characterization of pseudouridine (Ψ) in mammalian mRNA has been hampered by the lack of a quantitative method that maps Ψ in the whole transcriptome. We report bisulfite-induced deletion sequencing (BID-seq), which uses a bisulfite-mediated reaction to convert pseudouridine stoichiometrically into deletion upon reverse transcription without cytosine deamination. BID-seq enables detection of abundant Ψ sites with stoichiometry information in several human cell lines and 12 different mouse tissues using 10–20 ng input RNA. We uncover consensus sequences for Ψ in mammalian mRNA and assign different ‘writer’ proteins to individual Ψ deposition. Our results reveal a transcript stabilization role of Ψ sites installed by TRUB1 in human cancer cells. We also detect the presence of Ψ within stop codons of mammalian mRNA and confirm the role of Ψ in promoting stop codon readthrough in vivo. BID-seq will enable future investigations of the roles of Ψ in diverse biological processes.