The Aedes aegypti siRNA pathway mediates broad-spectrum defense against human pathogenic viruses and modulates antibacterial and antifungal defenses

Abstract

The mosquito’s innate immune system defends against a variety of pathogens, and the conserved siRNA pathway plays a central role in the control of viral infections. Here, we show that transgenic overexpression of Dicer2 (Dcr2) or R2d2 resulted in an accumulation of 21-nucleotide viral sequences that was accompanied by a significant suppression of dengue virus (DENV), Zika virus (ZIKV), and chikungunya virus (CHIKV) replication, thus indicating the broad-spectrum antiviral response mediated by the siRNA pathway that can be applied for the development of novel arbovirus control strategies. Interestingly, overexpression of Dcr2 or R2d2 regulated the mRNA abundance of a variety of antimicrobial immune genes, pointing to additional functions of DCR2 and R2D2 as well as cross-talk between the siRNA pathway and other immune pathways. Accordingly, transgenic overexpression of Dcr2 or R2d2 resulted in a lesser proliferation of the midgut microbiota and increased resistance to bacterial and fungal infections.

Characterization of New Defensin Antimicrobial Peptides and Their Expression in Bed Bugs in Response to Bacterial Ingestion and Injection

“…Initially, it was reported that the Toll signalling pathway responded to Gr+ bacteria and fungi, the immune deficiency (IMD) pathway to Gr– bacteria, and the JAK-STAT and RNAi pathways to viruses [ 17 , 18 , 19 , 20 ]. More recent studies have demonstrated that there is plasticity and cross-talk among and between immune pathways that allow strong multifaceted responses to pathogens and parasites [ 21 , 22 , 23 , 24 ].…”

CRISPR Mediated Transactivation in the Human Disease Vector Aedes aegypti

“…The mosquitoToll and JAK-STAT immune signaling pathways and the siRNA immune pathway, play important roles in defending against arboviral infections [56,66,67] (reviewed in [68]). Here as proof of principle, we used CRISPRa to transactivate AaRel1, an NF-κB Relish-like transcription factor that mediates the Toll immune pathway's antipathogenic action including the suppression of Dengue and Zika viruses [54][55][56][57].…”

“…With the rapid development of the tissue-specific dCas9-VPR lines in combinations with targets-specific sgRNA lines in the future, these key immune factors can be studied specifically in a spatio-temporal manner. This tool can be used to boost mosquitoes’ immunity against viral infections by transactivating genes involved in various immune pathways, such as Relish-like transcription factors Rel1 and Rel2, the two key downstream regulators of the Toll and IMD immune pathways [54,72,73], or the insect cytokine-like factor Vago [74,75], or DOME and HOP in the JAK-STAT pathway [67], or Dicer2 and R2d2 in the siRNA immune pathway [66]. The knowledge gained from these basic studies will further strengthen the development of vector control strategies.…”

“…DENV serotype 2 New Guinea C strain (DENV2) was cultured in Aedes albopictus C6/36 cells (ATCC CRL-1660), and viral stocks were prepared as previously described in [66,67,84]. All infection procedures were performed under BSL2 conditions for DENV2.…”

“…Viral stocks were aliquoted and stored at -80°C for long-term storage. Seven-day-old mosquitoes were orally infected with DENV2 through artificial glass membrane feeders as previously described [66,67,84]. A portion of each blood meal was frozen, and back titrated by plaque assay on the BHK-21 cells at 37°C.…”