@article{186666,
  author = {Wei Dai and Ang Li and Nathan J. Yu and Thao Nguyen and Robert W. Leach and Martin W{\"u}hr and Ralph E. Kleiner},
  title = {Activity-based RNA-modifying enzyme probing reveals DUS3L-mediated dihydrouridylation},
  abstract = {<p>Epitranscriptomic RNA modifications can regulate RNA activity; however, there remains a major gap in our understanding of the RNA chemistry present in biological systems. Here we develop RNA-mediated activity-based protein profiling (RNABPP), a chemoproteomic strategy that relies on metabolic RNA labeling, mRNA interactome capture and quantitative proteomics, to investigate RNA-modifying enzymes in human cells. RNABPP with 5-fluoropyrimidines allowed us to profile 5-methylcytidine (m5C) and 5-methyluridine (m5U) methyltransferases. Further, we uncover a new mechanism-based crosslink between 5-fluorouridine (5-FUrd)-modified RNA and the dihydrouridine synthase (DUS) homolog DUS3L. We investigate the mechanism of crosslinking and use quantitative nucleoside liquid chromatography{\textendash}tandem mass spectrometry (LC{\textendash}MS/MS) analysis and 5-FUrd-based crosslinking and immunoprecipitation (CLIP) sequencing to map DUS3L-dependent dihydrouridine (DHU) modifications across the transcriptome. Finally, we show that DUS3L-knockout (KO) cells have compromised protein translation rates and impaired cellular proliferation. Taken together, our work provides a general approach for profiling RNA-modifying enzyme activity in living cells and reveals new pathways for epitranscriptomic RNA regulation.</p>
},
  year = {2021},
  journal = {Nature Chemical Biology},
  month = {09/2021},
  issn = {1552-4469},
  url = {https://doi.org/10.1038/s41589-021-00874-8},
  doi = {10.1038/s41589-021-00874-8},
  language = {eng},
}