Juvenile hormone regulates silk-associated genes expression in Bombyx mori though m6A modification.

N6-methyladenosine (m6A) is the most abundant modification of RNA that regulates RNA fate in eukaryotic organisms. Juvenile Hormone (JH) is an indispensable insect hormone that is critical in regulating insect development, growth, and physiology. However, the relationship of m6A and JH remains an enigma. Here, global transcriptional patterns revealed that m6A-related genes are specifically regulated by JH in the posterior silk gland (PSG) of Bombyx mori. m6A sequencing of silkworm PSG found that many m6A-containing genes are involved in mRNA stability, translation, and degradation. GO enrichment analysis revealed that m6A-containing genes are relating to nucleic acid binding, nucleus, and nucleobase-containing compound metabolism. By analyzing transcriptome sequencing and m6A sequencing data conjointly, we discovered that JH significantly regulated the m6A modification of mRNA in PSG of B. mori, especially in silk-associated genes including sericin2, seroin1, BmSPI5, BmSPI4, Ldb, etc. Moreover, we found that the expression of silk-associated genes of are significantly regulated by METTL3 knockdown, validating silk-associated genes are targets of m6A modification. We identified that m6a modification of BmSPI4 and BmSPI5 mediates the JH-enhancing expression of BmSPI4 and BmSPI5 though mutagenesis of m6A sites in BmN cells. Taken together, these results suggest that m6a modification widely distributes the mRNA of silkworm PSG and plays a novel epigenetic mechanism for mediating JH effects on PSG of B. mori, particularly in silk production. Our findings provide a novel insight into the role of m6A in insects.