miRNA Modulation of Insect Virus Replication.

The outcome of virus infection in insects is impacted by regulation of both host and virus gene expression. A class of small RNAs called microRNAs (miRNA) have emerged as important regulators of gene expression that can influence the outcome of virus infection. miRNA regulation occurs at a comparatively late stage of gene expression, allowing for rapid control and fine-tuning of gene expression levels. Here we discuss the biogenesis of miRNAs from both host and virus genomes, the interactions that lead to regulation of gene expression, and the miRNA-mRNA interactions that lead to either antivirus or provirus consequences in the course of virus infection in insects.

miRNAs in Insects Infected by Animal and Plant Viruses.

Viruses vectored by insects cause severe medical and agricultural burdens. The process of virus infection of insects regulates and is regulated by a complex interplay of biomolecules including the small, non-coding microRNAs (miRNAs). Considered an anomaly upon its discovery only around 25 years ago, miRNAs as a class have challenged the molecular central dogma which essentially typifies RNAs as just intermediaries in the flow of information from DNA to protein. miRNAs are now known to be common modulators or fine-tuners of gene expression. While recent years has seen an increased emphasis on understanding the role of miRNAs in host-virus associations, existing literature on the interaction between insects and their arthropod-borne viruses (arboviruses) is largely restricted to miRNA abundance profiling. Here we analyse the commonalities and contrasts between miRNA abundance profiles with different host-arbovirus combinations and outline a suggested pipeline and criteria for functional analysis of the contribution of miRNAs to the insect vector-virus interaction. Finally, we discuss the potential use of the model organism, Drosophila melanogaster, in complementing research on the role of miRNAs in insect vector-virus interaction.

Wolbachia-mediated protection of Drosophila melanogaster against systemic infection with its natural viral pathogen Drosophila C virus does not involve changes in levels of highly abundant miRNAs.

The presence of Wolbachia confers virus protection to insects. The molecular mechanism underlying Wolbachia-mediated protection in this tripartite host-endosymbiont-virus interaction is not yet fully understood. In the bipartite association between Drosophila melanogaster and Drosophila C virus (DCV), changes in the expression of microRNAs (miRNAs) influence the outcome of viral pathogenesis. Here we examined whether changes in miRNA expression are similarly involved in the Drosophila-Wolbachia-DCV association. The levels of highly abundant miRNAs in D. melanogaster, Wolbachia-mono-infected D. melanogaster, and DCV- and Wolbachia-bi-infected D. melanogaster were quantified using RT-qPCR and compared. The results show that the abundance of the 17 tested D. melanogaster miRNAs is not affected by Wolbachia endosymbiosis or by bi-infection of Wolbachia and DCV. These results suggest that the in vivo protection conferred by Wolbachia to its native host against D. melanogaster's natural pathogen DCV is not likely to be dependent on or associated with changes in the levels of highly expressed miRNAs.

Drosophila microRNA modulates viral replication by targeting a homologue of mammalian cJun.

MicroRNAs (miRNAs) are important regulators of biological processes, including host-virus interaction. This study investigated the involvement of Drosophila melanogaster miR-8-5p in host-virus interaction. Drosophila flies and cells challenged with Drosophila C virus (DCV) were found to have lower miR-8-5p abundance compared to uninfected samples. Lowering miR-8-5p abundance by experimental inhibition of the miRNA led to an increase in viral accumulation, suggesting that the observed decrease in the miR-8-5p abundance during DCV infection enhances viral replication. miR-8-5p putative targets were identified and included dJun, a transcription factor gene whose mammalian homologue cJun is induced by various viruses through kinase activation. Increasing miR-8-5p abundance using miR-8-5p mimics resulted in a decrease in dJun and GFP reporter levels. Furthermore, when the putative target in dJun was mutated, addition of miR-8-5p mimics did not result in the same antagonistic effect on dJun. These results show negative regulation of dJun by miR-8-5p and suggest that an miRNA-mediated pathway is involved in dJun regulation during viral infection. To analyse the role of dJun during DCV infection, dJun was knocked down in cells prior to DCV infection. Knockdown of dJun decreased DCV replication, providing evidence that dJun up-regulation that is concomitant with miR-8-5p down-regulation during DCV infection supports viral replication. These results highlight the role of miRNA in regulating the transcription factor gene dJun and uncover a previously unrecognized mechanism by which dJun is regulated during host-virus interaction.

Drosophila miR-956 suppression modulates Ectoderm-expressed 4 and inhibits viral replication.

Small non-coding microRNAs (miRNAs) can modulate the outcome of virus infection. Here we explore the role of miRNAs in insect-virus interactions, in vivo, using the natural Drosophila melanogaster-Drosophila C virus (DCV) model system. Comparison of the miRNA expression profiles in DCV-infected and uninfected flies showed altered miRNA levels due to DCV infection, with the largest change in abundance observed for miR-956-3p. Knockout of miR-956 resulted to delayed DCV-induced mortality and decreased viral accumulation compared to wild-type flies. A screen of 84 putative miR-956-3p target genes identified regulation of Ectoderm-expressed 4 (Ect4) in miR-956 knockout flies and, separately, DCV infection. In Ect4 knockdown flies DCV-induced mortality occurred more quickly and virus accumulation was increased. Taken together, results suggest that the host-protective and antiviral consequences of miR-956 suppression during in vivo infection of D. melanogaster with its natural pathogen DCV is conferred through miR-956-3p induction of its target Ect4.