Effects of METTL3-METTL14 on primary microRNA processing by Drosha-DGCR8

Abstract

MicroRNAs modulate most protein-coding genes, and many are regulated during maturation. Chemical modifications of primary transcripts containing microRNAs have been implicated in altering Microprocessor processing efficiency, a key initiating endonucleolytic step performed by Drosha and DGCR8. METTL3-METTL14 produces N6-methyladenosine which is the most common methylation for mRNAs. Genetic experiments suggested that METTL3-METTL14 promotes primary microRNA processing by Microprocessor, but the molecular mechanism still needs to be elucidated. We tested the hypothesis that METTL3-METTL14 or m6A may directly impact Drosha or DGCR8 function during primary microRNA processing. After reconstituting the methyltransferase and processing activities, we show that the presence of METTL3-METTL14 complexes does not affect the processing efficiency of Drosha-DGCR8. We also established a method to prepare m6A-modified primary microRNAs and used them to show that the processing of the transcripts with m6A is similar to those without any modification. Recombinant METTL3-METTL14 and DGCR8 do not form stable complexes, challenging the previous model that depends on enhanced DGCR8 recruitment. Therefore, METTL3-METTL14 or m6A modification does not generally promote Microprocessor-mediated microRNA processing, although they may impact certain cases.