YTHDF2 Recognition of N-Methyladenosine (mA)-Modified RNA Is Associated with Transcript Destabilization.
Epitranscriptomic modifications play an important role in RNA function and can impact gene expression. Here, we apply a chemical proteomics approach to investigate readers of N-methyladenosine (mA), a poorly characterized modification on mammalian mRNA. We find that YTHDF proteins, known mA readers, recognize mA-modified sequences in a methylation-specific manner. We characterize binding of recombinant YTHDF1/2 proteins to mA-modified oligonucleotides to demonstrate that these interactions can exhibit comparable affinity to mA-recognition events and occur in diverse sequence contexts. Further, we demonstrate YTHDF2 interacts specifically with endogenously modified mA transcripts. Finally, we deplete cellular YTHDF2 to show that the abundance of mA-modified transcripts is increased in its absence. Similarly, increasing mA levels through depletion of ALKBH3, an mA eraser protein, destabilizes known mA-containing RNAs. Our results shed light on the function of mA on mRNA and provide a mechanistic framework to further evaluate the role of mA in biological processes.