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Quantitative CRACI reveals transcriptome-wide distribution of RNA dihydrouridine at base resolution

Author

Listed:
  • Cheng-Wei Ju

    (The University of Chicago
    The University of Chicago
    The University of Chicago)

  • Han Li

    (The University of Chicago
    The University of Chicago)

  • Bochen Jiang

    (The University of Chicago
    The University of Chicago
    Shanghai Jiao Tong University)

  • Xuanhao Zhu

    (The University of Chicago
    The University of Chicago)

  • Liang Cui

    (Singapore-MIT Alliance for Research and Technology)

  • Zhanghui Han

    (National University of Singapore)

  • Junxi Zou

    (The University of Chicago
    The University of Chicago)

  • Yunzheng Liu

    (California Institute of Technology)

  • Shenghai Shen

    (The Hong Kong University of Science and Technology
    The Hong Kong University of Science and Technology)

  • Hardik Shah

    (The University of Chicago)

  • Chang Ye

    (The University of Chicago
    The University of Chicago)

  • Yuhao Zhong

    (The University of Chicago
    The University of Chicago)

  • Ruiqi Ge

    (The University of Chicago
    The University of Chicago)

  • Peng Xia

    (The University of Chicago
    The University of Chicago)

  • Yiyi Ji

    (The University of Chicago
    The University of Chicago)

  • Shun Liu

    (The University of Chicago
    The University of Chicago)

  • Fan Yang

    (The University of Chicago
    The University of Chicago)

  • Bei Liu

    (The University of Chicago
    The University of Chicago)

  • Yuzhi Xu

    (New York University)

  • Jiangbo Wei

    (National University of Singapore
    National University of Singapore)

  • Li-Sheng Zhang

    (The Hong Kong University of Science and Technology
    The Hong Kong University of Science and Technology)

  • Chuan He

    (The University of Chicago
    The University of Chicago
    The University of Chicago)

Abstract

Dihydrouridine (D) is an abundant RNA modification, yet its roles in mammals remain poorly understood due to limited detection methods. We even do not have a comprehensive profile of D site location and modification stoichiometry in tRNA. Here, we introduce Chemical Reduction Assisted Cytosine Incorporation sequencing (CRACI), a highly sensitive, quantitative approach for mapping D at single-base resolution. Using CRACI, we generate the transcriptome-wide maps of D in both cytoplasmic and mitochondrial tRNAs from mammals and plants. We uncover D sites in mitochondrial tRNAs and identify DUS2L as the ‘writer’ protein responsible for human mitochondrial tRNAs. Furthermore, we demonstrate that most D modifications have a limited impact on tRNA stability, except for D20a, which also exhibits cis-regulation of adjacent D20 sites. Application of CRACI to human mRNA reveals that D modifications are present but rare and occur at very low stoichiometry. CRACI thus provides a powerful platform for investigating D biology across species.

Suggested Citation

  • Cheng-Wei Ju & Han Li & Bochen Jiang & Xuanhao Zhu & Liang Cui & Zhanghui Han & Junxi Zou & Yunzheng Liu & Shenghai Shen & Hardik Shah & Chang Ye & Yuhao Zhong & Ruiqi Ge & Peng Xia & Yiyi Ji & Shun L, 2025. "Quantitative CRACI reveals transcriptome-wide distribution of RNA dihydrouridine at base resolution," Nature Communications, Nature, vol. 16(1), pages 1-14, December.
    • Handle: RePEc:nat:natcom:v:16:y:2025:i:1:d:10.1038_s41467-025-63918-w
    DOI: 10.1038/s41467-025-63918-w
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