N-methyladenosine modification of the Aedes aegypti transcriptome and its alteration upon dengue virus infection in Aag2 cell line.

The N-methyladenosine (mA) modification of RNA has been reported to affect viral infections. Studies have confirmed the role of mA in replication of several vector-borne flaviviruses, including dengue virus (DENV), in mammalian cells. Here, we explored the role of mA in DENV replication in the mosquito Aedes aegypti Aag2 cell line. We first determined the presence of mA on the RNAs from mosquito cells and using methylated RNA immunoprecipitation and sequencing (MeRIP-Seq) identified mA modification of the mosquito transcriptome and those that changed upon DENV infection. Depletion of mA methyltransferases and the mA binding protein YTHDF3 RNAs decreased the replication of DENV. In particular, we found that the Ae. aegypti ubiquitin carrier protein 9 (Ubc9) is mA modified and its expression increases after DENV infection. Silencing of the gene and ectopic expression of Ubc9 led to reduced and increased DENV replication, respectively. The abundance of Ubc9 mRNA and its stability were reduced with the inhibition of mA modification, implying that mA modification of Ubc9 might enhance expression of the gene. We also show that the genome of DENV is mA modified at five sites in mosquito cells. Altogether, this work reveals the involvement of mA modification in Ae. aegypti-DENV interaction.