Abstract

In eukaryotes, N6-methyladenosine (m 6 A) is one of the most abundant modifications on RNAs , and it plays important roles in many biological processes and diseases such as cancer. While most m 6 A researches focus on message RNAs and long non-coding RNAs, recent studies have reported the presence of m 6 A in small RNAs. Nevertheless, current knowledge about m 6 A prevalence in mature microRNAs (miRNA) is extremely limited and the functional significance of m 6 A methylation in miRNAs remains to be elucidated. Here, we demonstrated cell-specific m 6 A profiles of miRNAs in A549 human non-small cell lung cancer (NSCLC) cells and HEK293A cells by using miRNA m 6 A immunoprecipitation sequencing and constructed the consensus motif in m 6 A-enriched miRNAs de novo . We found that miR-21–5p, an oncogenic miRNA, showed the highest m 6 A enrichment in NSCLC cells. Depletion of the demethylase ALKBH5 did not change the expression level of miR-21–5p, but altered the m 6 A abundance of miR-21–5p, thereby changing the expression levels of its target gene. We further synthesized m 6 A modified miR-21–5p mimics in vitro and demonstrated that in NSCLC cells, m 6 A marks in mature miR-21–5p could directly affect its silencing potency towards target genes, which finally impaired its promotion to proliferation and motility. Together, our findings reveal the landscape of m 6 A modification in mature miRNAs, and provide the first evidence that it may contribute to the mRNA responses to cancer-related miRNAs.