Human cytoplasmic tRNAs contain dihydrouridine modifications at positions 16 and 17 (D16/D17). The enzyme responsible for D16/D17 formation and its cellular roles remain elusive. Here, we identify DUS1L as the human tRNA D16/D17 writer. DUS1L knockout in the glioblastoma cell lines LNZ308 and U87 causes loss of D16/D17. D formation is reconstituted in vitro using recombinant DUS1L in the presence of NADPH or NADH. DUS1L knockout/overexpression in LNZ308 cells shows that DUS1L supports cell growth. Moreover, higher DUS1L expression in glioma patients is associated with poorer prognosis. Upon vector-mediated DUS1L overexpression in LNZ308 cells, 5' and 3' processing of precursor tRNATyr(GUA) is inhibited, resulting in a reduced mature tRNATyr(GUA) level, reduced translation of the tyrosine codons UAC and UAU, and reduced translational readthrough of the near-cognate stop codons UAA and UAG. Moreover, DUS1L overexpression increases the amounts of several D16/D17-containing tRNAs and total cellular translation. Our study identifies a human dihydrouridine writer, providing the foundation to study its roles in health and disease.

Center for Metabolic Regulation of Healthy Aging Faculty of Life Science Kumamoto University Kumamoto Japan

Department of Cell Pathology Faculty of Life Sciences Kumamoto University Kumamoto Japan

Department of Chemistry and Biotechnology Graduate School of Engineering University of Tokyo Tokyo Japan

Department of Modomics Biology and Medicine Institute of Development Aging and Cancer Tohoku University Sendai Japan

Department of Molecular Physiology Faculty of Life Sciences Kumamoto University Kumamoto Japan

Department of Neurosurgery Faculty of Life Sciences Kumamoto University Kumamoto Japan

Laboratory of Regulation of Gene Expression Institute of Microbiology of the Czech Academy of Sciences Prague Czech Republic

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