title: Functional specialization of the small interfering RNA pathway in response to virus infection.
abstract: In Drosophila, post-transcriptional gene silencing occurs when exogenous or endogenous double stranded RNA (dsRNA) is processed into small interfering RNAs (siRNAs) by Dicer-2 (Dcr-2) in association with a dsRNA-binding protein (dsRBP) cofactor called Loquacious (Loqs-PD). siRNAs are then loaded onto Argonaute-2 (Ago2) by the action of Dcr-2 with another dsRBP cofactor called R2D2. Loaded Ago2 executes the destruction of target RNAs that have sequence complementarity to siRNAs. Although Dcr-2, R2D2, and Ago2 are essential for innate antiviral defense, the mechanism of virus-derived siRNA (vsiRNA) biogenesis and viral target inhibition remains unclear. Here, we characterize the response mechanism mediated by siRNAs against two different RNA viruses that infect Drosophila. In both cases, we show that vsiRNAs are generated by Dcr-2 processing of dsRNA formed during viral genome replication and, to a lesser extent, viral transcription. These vsiRNAs seem to preferentially target viral polyadenylated RNA to inhibit viral replication. Loqs-PD is completely dispensable for silencing of the viruses, in contrast to its role in silencing endogenous targets. Biogenesis of vsiRNAs is independent of both Loqs-PD and R2D2. R2D2, however, is required for sorting and loading of vsiRNAs onto Ago2 and inhibition of viral RNA expression. Direct injection of viral RNA into Drosophila results in replication that is also independent of Loqs-PD. This suggests that triggering of the antiviral pathway is not related to viral mode of entry but recognition of intrinsic features of virus RNA. Our results indicate the existence of a vsiRNA pathway that is separate from the endogenous siRNA pathway and is specifically triggered by virus RNA. We speculate that this unique framework might be necessary for a prompt and efficient antiviral response.
authors.affiliation: Department of Molecular Biosciences, Northwestern University, Evanston, Illinois, United States of America. jtm@ufmg.br
authors.forename: Joao Trindade
authors.initials: JT
authors.lastname: Marques
authors.forename: Ji-Ping
authors.initials: JP
authors.lastname: Wang
authors.forename: Xiaohong
authors.initials: X
authors.lastname: Wang
authors.forename: Karla Pollyanna Vieira
authors.initials: KP
authors.lastname: de Oliveira
authors.forename: Catherine
authors.initials: C
authors.lastname: Gao
authors.forename: Eric Roberto Guimaraes Rocha
authors.initials: ER
authors.lastname: Aguiar
authors.forename: Nadereh
authors.initials: N
authors.lastname: Jafari
authors.forename: Richard W
authors.initials: RW
authors.lastname: Carthew
mesh_terms: D000818:Animals; D060565:Argonaute Proteins / Q000235:genetics / Q000378:metabolism; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism; D004331:Drosophila melanogaster; D020365:RNA Helicases / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D012328:RNA Viruses / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism*; D012367:RNA, Viral / Q000096:biosynthesis* / Q000235:genetics; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D043244:Ribonuclease III / Q000235:genetics / Q000378:metabolism; D014777:Virus Diseases / Q000235:genetics / Q000378:metabolism*
chemical_list: C433807:AGO2 protein, Drosophila; D060565:Argonaute Proteins; D029721:Drosophila Proteins; C503213:Loqs protein, Drosophila; C478068:R2D2 protein, Drosophila; D034741:RNA, Small Interfering; D012367:RNA, Viral; D016601:RNA-Binding Proteins; C478069:DCR-2 protein, Drosophila; D043244:Ribonuclease III; D020365:RNA Helicases