Transcriptome-wide distribution and function of RNA hydroxymethylcytosine

Our understanding of the posttranscriptional modifications that decorate RNA, a regulatory layer positioned between DNA and proteins, is in its infancy. Hydroxymethylcytosine, well described in DNA, occurs also in RNA, where its distribution, localization and function remain unexplored. Here, we show that hydroxymethylcytosine preferentially marks polyadenylated RNAs and is deposited by Tet in Drosophila. We provide the transcriptome-wide hydroxymethylation landscape, revealing hydroxymethylcytosine in the transcripts of many genes, notably in coding sequences, and identify consensus sites for hydroxymethylation. We find that RNA hydroxymethylation can favor mRNA translation. Tet-deficient fruitflies show impaired brain development, accompanied by decreased RNA hydroxymethylation. Together, these steps into uncharted territory uncover central roles for RNA hydroxymethylation in Drosophila, highlighting the importance of epitranscriptomics. Overall design: RNA hydroxymethylation landscape in S2 cell lines.