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<p>RNA oxidation is an important yet understudied process, partly because methods to localize oxidized residues in RNA are lacking. We introduce OAbSeq, a deep-sequencing approach that maps oxidized sites with high sensitivity by exploiting aniline-induced strand scission at noncanonical nucleosides to generate unique ligation-competent fragments utilized for library preparation. Applied to yeast RNA, OAbSeq detects widespread signals predominating at purines, especially at guanosines. Exogenous oxidation increased signal intensity but preserved the guanosine-dominated pattern. Parallel quantification of 8-oxoguanosine (oxo<sup>8</sup>G) and abasic sites revealed that abasic sites are more abundant than oxo<sup>8</sup>G following oxidative treatment in vitro and under physiological conditions. These data support a model in which guanosine oxidation proceeds via transient oxo<sup>8</sup>G yielding abasic sites that can be mapped at nucleotide resolution by OAbSeq. Our findings also suggest abasic sites may be a more informative marker of RNA oxidative damage than oxo<sup>8</sup>G, facilitating studies of RNA oxidation dynamics in cells.</p>

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Mapping of HOCl-oxidized RNA identifies abasic sites as major damage and oxidation product of oxo8G

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年11月21日

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  • Marlies Weber,
  • Kasturi Raorane,
  • Clara Johanna Grampp,
  • Valérie Bourguignon,
  • Lea-Marie Kilz,
  • David Glänzer,
  • Virginie Marchand,
  • Christoph Kreutz,
  • Yuri Motorin,
  • Mark Helm

摘要

RNA oxidation is an important yet understudied process, partly because methods to localize oxidized residues in RNA are lacking. We introduce OAbSeq, a deep-sequencing approach that maps oxidized sites with high sensitivity by exploiting aniline-induced strand scission at noncanonical nucleosides to generate unique ligation-competent fragments utilized for library preparation. Applied to yeast RNA, OAbSeq detects widespread signals predominating at purines, especially at guanosines. Exogenous oxidation increased signal intensity but preserved the guanosine-dominated pattern. Parallel quantification of 8-oxoguanosine (oxo8G) and abasic sites revealed that abasic sites are more abundant than oxo8G following oxidative treatment in vitro and under physiological conditions. These data support a model in which guanosine oxidation proceeds via transient oxo8G yielding abasic sites that can be mapped at nucleotide resolution by OAbSeq. Our findings also suggest abasic sites may be a more informative marker of RNA oxidative damage than oxo8G, facilitating studies of RNA oxidation dynamics in cells.