Fig 1.
Divergence of piRNA pathways in Drosophila melanogaster and Aedes aegypti.
In Drosophila (left panel), PIWI proteins are almost exclusively expressed in gonadal tissues. Nuclear Piwi is expressed in both germ cells and ovarian somatic cells, whereas Aub and Ago3 expression is limited to germ cells specifically. In the nuage surrounding the nucleus of these cells, Aub and Ago3 form the ping-pong amplification complex, which is responsible for secondary piRNA production with the characteristic 1U/10A nucleotide bias (Box 1). Drosophila piRNAs are mainly derived from transposon sequences and to a lesser extent from mRNA. In Ae. aegypti (right panel), the PIWI protein family is expanded to eight members (Piwi 1–7 and Ago3), some of which are expressed in somatic tissues. Of these PIWI proteins, Piwi5 and Ago3 interact to produce piRNAs with the 1U/10A nucleotide bias indicative of secondary piRNA production through ping-pong amplification. In Aedes, piRNAs are produced from viral RNA, in addition to transposon sequences and mRNA.
Table 1.
vpiRNA production in infections with arboviruses and insect-specific viruses.
Fig 2.
piRNA distributions across the genomes of selected (A) alphaviruses, (B) flaviviruses, and (C) bunyaviruses. The plots depict published genome profiles of Sindbis virus (SINV) [29], chikungunya virus (CHIKV) [30], Semliki Forest virus (SFV) [31], dengue virus serotype 2 (DENV2) [35,36], cell fusing agent virus (CFAV) [34], Rift Valley fever virus (RVFV) [38], and Schmallenberg virus (SBV) [37]. For alphaviruses, the position of the subgenomic promoter is depicted. The piRNA coverage on the sense or antisense strand is shown as peaks above or below the x-axis, respectively. Please note that the plots are representations of piRNA profiles from multiple studies that used different ways of normalizing and presenting read counts. Therefore, the heights of the bars are arbitrary and do not allow a quantitative comparison between the different viruses.
Fig 3.
Model for piRNA biogenesis in Aedes aegypti.
RNA molecules from varying sources are processed differently by the piRNA machinery in Ae. aegypti. Upon acute infection, Sindbis virus RNA is processed into ping-pong–dependent piRNAs involving PIWI proteins Piwi5 and Ago3. In contrast, dengue virus RNA can also be processed into piRNAs by Piwi6. Transposon-derived piRNAs associate primarily with Piwi5 and Piwi6; however, some transposon RNAs feed into the ping-pong loop and give rise to Ago3-bound secondary piRNAs. Additionally, the production of transposon piRNAs is dependent on Piwi4 in an indirect manner, as transposon-derived piRNAs are not loaded in Piwi4, but knockdown of Piwi4 does reduce their numbers. Viral RNA may directly enter the piRNA machinery; additionally, viral RNA is reverse transcribed to produce a DNA form of the virus (vDNA). The vDNA may either remain episomal or integrate into the host genome. Putative vDNA-derived transcripts may serve as additional precursors for vpiRNA production. Moreover, when genome integration occurs in the germline, the vDNA fragment forms a novel endogenous viral element (EVE) that may lead to the production of EVE-derived piRNAs.