Cancer mutations rewire the RNA methylation specificity of METTL3-METTL14

Chemical modification of RNAs is important for post-transcriptional gene regulation. The METTL3-METTL14 complex generates most N6-methyladenosine (m6A) modifications in mRNAs, and dysregulated methyltransferase expression has been linked to cancers. Here we show that a changed sequence context for m6A can promote oncogenesis. A gain-of-function missense mutation from cancer patients, METTL14R298P, increases malignant cell growth in culture and transgenic mice, without increasing global m6A levels in mRNAs. The mutant methyltransferase preferentially modifies noncanonical sites containing a GGAU motif, in vitro and in vivo. The m6A in GGAU context is detected by the YTH family of readers similarly to the canonical sites but is demethylated less efficiently by an eraser, ALKBH5. Combining the biochemical and structural data we provide a model for how the cognate RNA sequences are selected for methylation by METTL3-METTL14. Our work highlights that sequence-specific m6A deposition is important and that increased GGAU methylation can promote oncogenesis. Overall design: We generated four stable HepG2 liver cancer cell lines by integrating lentivirus cassettes expressing EGFP, METTL14 (WT), METTL14 (R298P), or METTL14 (D312A). m6A-seq was performed using three biological replicates from each cell line. The sequencing data included the reads of input sample and RNA immunoprecipitation sample per replicate per cell line. The RNA-seq analysis was performed using the reads of input samples from three replicates of METTL14 (WT), METTL14 (R298P) expressing cell lines.