2'-O-Methyl-5-hydroxymethylcytidine: A Second Oxidative Derivative of 5-Methylcytidine in RNA

5-Hydroxymethylcytidine (hm5C) was recently identified as a direct metabolite of m5C in RNA. We investigated the stability of hm5C in human cells using bio-isotopologues and LC-MS/HRMS. This has led to the discovery of a second oxidative metabolite of m5C in RNA, namely 2′-O-methyl-5-hydroxymethylcytidine (hm5Cm). Subsequent quantitative analysis of total RNA from higher organisms revealed varying levels and TET-independent formation of this new RNA modification.