RNA oxidation HOCl

RNA oxidation by hypochlorous acid (HOCl) is of major interest in vitro as well as in vivo settings. Conducting a thorough analysis of HOCl oxidation of ribonucleosides, oligoribonucleotides and native RNAs by LC-MS, we characterized dominant oxidation events and products in vitro. In addition to extensive cytidine chlorination, the predominant reactivity was guanosine oxidation leading to 8-oxoguanosine as an intermediate, which was further oxidized, thereby acting as a scavenger in protecting adjacent residues from oxidation, and eventually resulting in an abasic site, or a derivative with similar reactivity. We show that, while 8 oxoguanosine does not undergo ?-elimination under conventional aniline treatment, a combination with prior alkaline treatment does. On this basis, we developed a method that allows mapping of oxidative guanosine lesions in RNA sequences, not only detecting abasic sites but including direct detection of 8-oxoguanosine in a sequence context. An application to yeast ribosomal RNA oxidized as pure RNA, or in ribosome particles, or in living cells, revealed idiosyncratic features in each of these RNA preparations.