2. Academic Validation
  3. Chemical pulldown reveals dynamic pseudouridylation of the mammalian transcriptome

Chemical pulldown reveals dynamic pseudouridylation of the mammalian transcriptome

  • Nat Chem Biol. 2015 Aug;11(8):592-7. doi: 10.1038/nchembio.1836.
Xiaoyu Li 1,  Ping Zhu 2,  Shiqing Ma 3,  Jinghui Song 1,  Jinyi Bai 3,  Fangfang Sun 1,  Chengqi Yi 4
Affiliations

Affiliations

  • 1 State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences and Peking-Tsinghua Center for Life Sciences, Peking University, Beijing, China.
  • 2 Biodynamic Optical Imaging Center, School of Life Sciences, Peking University, Beijing, China.
  • 3 1] State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences and Peking-Tsinghua Center for Life Sciences, Peking University, Beijing, China. [2] Academy for Advanced Interdisciplinary Studies, Peking University, Beijing, China.
  • 4 1] State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences and Peking-Tsinghua Center for Life Sciences, Peking University, Beijing, China. [2] Synthetic and Functional Biomolecules Center, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
PMID: 26075521 DOI: 10.1038/nchembio.1836
Abstract

Pseudouridine (Ψ) is the most abundant post-transcriptional RNA modification, yet little is known about its prevalence, mechanism and function in mRNA. Here, we performed quantitative MS analysis and show that Ψ is much more prevalent (Ψ/U ratio ∼0.2-0.6%) in mammalian mRNA than previously believed. We developed N3-CMC-enriched pseudouridine Sequencing (CeU-Seq), a selective chemical labeling and pulldown method, to identify 2,084 Ψ sites within 1,929 human transcripts, of which four (in ribosomal RNA and EEF1A1 mRNA) are biochemically verified. We show that hPUS1, a known Ψ synthase, acts on human mRNA; under stress, CeU-Seq demonstrates inducible and stress-specific mRNA pseudouridylation. Applying CeU-Seq to the mouse transcriptome revealed conserved and tissue-specific pseudouridylation. Collectively, our approaches allow comprehensive analysis of transcriptome-wide pseudouridylation and provide tools for functional studies of Ψ-mediated epigenetic regulation.

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