Chemistry of installing epitranscriptomic 5-modified cytidines in RNA oligomers.

Studies of 5-hydroxymethylcytidine (hm 5 C), 5-formylcytidine (f 5 C) and 5-carboxycytidine (ca 5 C) modifications as products of the 5-methylcytidine (m 5 C) oxidative demethylation pathway in cellular mRNAs constitute an important element of the new epitranscriptomic field of research. The dynamic process of m 5 C conversion and final turnover to the parent cytidine is considered a post-transcriptional layer of gene-expression regulation. However, the regulatory mechanism associated with epitranscriptomic cytidine modifications remains largely unknown. Therefore, oligonucleotides containing m 5 C oxidation products are of great value for the next generation of biochemical, biophysical, and structural studies on their function, metabolism, and contribution to human diseases. Herein, we summarize the synthetic strategies developed for the incorporation of hm 5 C, f 5 C and ca 5 C into RNA oligomers by phosphoramidite chemistry, including post-synthetic C5-cytidine functionalization and enzymatic methods.