Background
Emerging evidence indicates that microRNAs (miRNAs) are involved in host-virus interaction. We previously reported that some miRNAs were differentially expressed in sugar-fed and blood-fed females of Aedes albopictus (Ae. albopictus). Here, we analysis the role in the host-virus system of an abundant midgut-specific miRNA in the mosquito Ae. albopictus.
Conclusion
These findings confirm that miR-281, an abundant midgut-specific miRNA, facilitates DENV-2 replication.
Methods
The expression profiles of miR-281 in different body parts of Ae. albopictus and following dengue virus infection were determined using RT-qPCR and Northern blot. miR-281 mimics, antagomiRs and corresponding negative controls were designed and their overexpression and knock-down efficiency were analyzed by qRT-PCR after transfecting the mosquito cell lines C6/36, and also by injecting female mosquitoes. Dengue virus serotype-2 (DENV-2) viral genomic RNA abundance was determined by RT-qPCR. The levels of DENV-2 E protein were detected using Western blot. Virus titers were tested using TCID50. RNAhybrid was used to predict targets of miR-281 in the DENV-2 genome. The EGFP plasmid-based reporter system was used to investigate the interaction between miR-281 and the predicted binding site in the C6/36 cell line.
Results
miR-281 is specifically expressed in the female midgut where dengue virus first invades. After DENV-2 infection, this miRNA is up-regulated in response to viral infection. Functional intervention analyses in vitro with specifically designed miR-281 mimics and corresponding antagomiRs indicated that miR-281 enhances DENV-2 viral replication. Further depletion of miR-281 in female mosquitoes by injection of its specific antagomiRs led to a significant reduction in DENV-2 abundance. The interaction between miR-281 and its predicted target sequence, the DENV-2 genomic 5'-untranslated region (UTR), is confirmed in the context of a plasmid-based reporter system.