Over 150 types of RNA modifications are identified in RNA molecules. Transcriptome profiling is one of the key steps in decoding the epitranscriptomic panorama of these chemical modifications and their potential functions. N 7-methylguanosine (m 7 G) is one of the most abundant modifications present in tRNA, rRNA and mRNA 5′cap, and has critical roles in regulating RNA processing, metabolism and function. Besides its presence at the cap position in mRNAs, m 7 G is also identified in internal mRNA regions. However, its transcriptome-wide distribution and dynamic regulation within internal mRNA regions remain unknown. Here, we have established m 7 G individual-nucleotide-resolution cross-linking and immunoprecipitation with sequencing (m 7 G miCLIP-seq) to specifically detect internal mRNA m 7 G modification. Using this approach, we revealed that m 7 G is enriched at the 5′UTR region and AG-rich contexts, a feature that is well-conserved across different human/mouse cell lines and mouse tissues. Strikingly, the internal m 7 G modification is dynamically regulated under both H 2 O 2 and heat shock treatments, with remarkable accumulations in the CDS and 3′UTR regions, and functions in promoting mRNA translation efficiency. Consistently, a PCNA 3′UTR minigene reporter harboring the native m 7 G modification site displays both enriched m 7 G modification and increased mRNA translation upon H 2 O 2 treatment compared to the m 7 G site-mutated minigene reporter (G to A). Taken together, our findings unravel the dynamic profiles of internal mRNA m 7 G methylome and highlight m 7 G as a novel epitranscriptomic marker with regulatory roles in translation.

The RNA modification N6-methyladenosine (m6A) plays a key role in the life cycles of several RNA viruses. Whether this applies to SARS-CoV-2 and whether m6A affects the outcome of COVID-19 disease is still poorly explored. Here we report that the RNA demethylase FTO strongly affects both m6A marking of SARS-CoV-2 and COVID-19 severity. By m6A profiling of SARS-CoV-2, we confirmed in infected cultured cells and showed for the first time in vivo in hamsters that the regions encoding TRS_L and the nucleocapsid protein are multiply marked by m6A, preferentially within RRACH motifs that are specific to β-coronaviruses and well conserved across SARS-CoV-2 variants. In cells, downregulation of the m6A demethylase FTO, occurring upon SARS-CoV-2 infection, increased m6A marking of SARS-CoV-2 RNA and slightly promoted viral replication. In COVID-19 patients, a negative correlation was found between FTO expression and both SARS-CoV-2 expression and disease severity. FTO emerged as a classifier of disease severity and hence a potential stratifier of COVID-19 patients.