Arbovirus-Derived piRNAs Exhibit a Ping-Pong Signature in Mosquito Cells

Abstract

The siRNA pathway is an essential antiviral mechanism in insects. Whether other RNA interference pathways are involved in antiviral defense remains unclear. Here, we report in cells derived from the two main vectors for arboviruses, Aedes albopictus and Aedes aegypti, the production of viral small RNAs that exhibit the hallmarks of ping-pong derived piwi-associated RNAs (piRNAs) after infection with positive or negative sense RNA viruses. Furthermore, these cells produce endogenous piRNAs that mapped to transposable elements. Our results show that these mosquito cells can initiate de novo piRNA production and recapitulate the ping-pong dependent piRNA pathway upon viral infection. The mechanism of viral-piRNA production is discussed.

Antiviral responses of arthropod vectors: an update on recent advances

“…Increased knowledge about the piRNA pathway in mosquitoes reveals many apparent differences from that in Drosophila (reviewed by [32]). Virus-derived piRNAs have been shown to be produced in whole Aedes mosquitoes upon infection with CHIKV [82] and DENV [45,104], in Aedes mosquito soma upon infection with CHIKV [82] and in the cell lines U4.4, C6/36 (both A. albopictus) and Aag2 (A. aegypti) upon infection with Sindbis virus (SINV) [123]. Virus-derived piRNAs were also observed in C6/36 cells infected with the bunyavirus La Crosse virus (LACV) [123].…”

“…Virus-derived piRNAs have been shown to be produced in whole Aedes mosquitoes upon infection with CHIKV [82] and DENV [45,104], in Aedes mosquito soma upon infection with CHIKV [82] and in the cell lines U4.4, C6/36 (both A. albopictus) and Aag2 (A. aegypti) upon infection with Sindbis virus (SINV) [123]. Virus-derived piRNAs were also observed in C6/36 cells infected with the bunyavirus La Crosse virus (LACV) [123]. In another study, both U4.4 and Aag2 cells produced piRNAs after infection with the alphavirus Semliki Forest virus (SFV), and knockdown of one of the PIWI proteins, PIWI4, resulted in an increase in SFV replication, demonstrating that activity of a PIWI protein was involved in controlling arbovirus replication [102].…”

“…Hoa et al [47] have shown that the PIWI protein Ago-3 plays a role in gene silencing mediated by dsRNA treatment in A. gambiae cells and Morazzani et al [82] suggested that dsRNA is responsible for the piRNA-like molecules induced by CHIKV in C6/36 cells. However, Schnettler et al [102] showed that transfection of dsRNA alone is not sufficient to induce production of piRNA-like molecules, thus providing evidence that ssRNA as a result of active virus replication is required, and Vodovar et al [123] also argue that piRNAs are derived from viral ssRNA generating primary piRNAs from the (-) strand and secondary piRNAs from the abundant (?) sense mRNA.…”

“…While the generation of exogenous siRNAs has been implicated in antiviral defences in arthropods, miRNAs and piRNAs have mainly been associated with regulation of gene expression and control of transposons in germlines (reviewed by [5,55]); however, recently their role in virus infection has become more apparent. Numerous small RNA profiles of arbovirus-infected mosquitoes or mosquito cells have been published [45,68,84,111,123] and have provided insights into the importance of the different small RNA pathways during arbovirus infection. Knowledge gained about antiviral RNAi over the last decade has also led to new techniques for virus discovery.…”

Everybody wins! Poland hosts thrilling competitions of viruses, RNAi and football teams

“…In both cases, Dicer-2-dependent vsiRNAs with a typical length of 21 nt were observed, scattered across the genome and mapping in similar numbers to the viral (+) and (-) strands. Strikingly, in Aedes cells, an additional population of viral small RNAs showed the characteristics of ping-pong-derived piwi-associated RNAs [3]. Arboviruses replicate efficiently in both their mosquito and vertebrate hosts, which provides an excellent opportunity to study viral small RNAs from the same virus in two disparate hosts.…”