Transcriptome-wide profiling of RNA N4-cytidine acetylation in Arabidopsis thaliana and Oryza sativa

Acetylation of N4-cytidine (ac4C) has recently been discovered as a novel modification of mRNA. RNA ac4C modification has been shown to be a key regulator of RNA stability, RNA translation, and the thermal stress response. However, its existence in eukaryotic mRNAs is still controversial. In plants, the existence, distribution pattern, and potential function of RNA ac4C modification are largely unknown. Here we report the presence of ac4C in the mRNAs of both Arabidopsis thaliana and rice (Oryza sativa). By comparing two ac4C sequencing methods, we found that RNA immunoprecipitation and sequencing (acRIP-seq), but not ac4C sequencing, was suitable for plant RNA ac4C sequencing. We present transcriptome-wide atlases of RNA ac4C modification in A. thaliana and rice mRNAs obtained by acRIP-seq. Analysis of the distribution of RNA ac4C modifications showed that ac4C is enriched near translation start sites in rice mRNAs and near translation start sites and translation end sites in Arabidopsis mRNAs. The RNA ac4C modification level is positively correlated with RNA half-life and the number of splicing variants. Similar to that in mammals, the translation efficiency of ac4C target genes is significantly higher than that of other genes. Our in vitro translation results confirmed that RNA ac4C modification enhances translation efficiency. We also found that RNA ac4C modification is negatively correlated with RNA structure. These results suggest that ac4C is a conserved mRNA modification in plants that contributes to RNA stability, splicing, translation, and secondary structure formation.