In:
Proceedings of the National Academy of Sciences, Proceedings of the National Academy of Sciences, Vol. 109, No. 51 ( 2012-12-18)
Abstract:
Mosquitoes and other blood-feeding arthropods are important vectors for emerging and reemerging viral infectious diseases. RNAi has previously been shown to control vector-borne RNA virus infection in mosquitoes ( 4 ). Our results show that the insect RNAi machinery also controls DNA virus infection, expanding the breadth of the RNAi-mediated antiviral response to all major classes of viruses. Understanding how the RNAi mechanism functions as a nucleic acid-based immune system in insects could help the development of new methods to control arthropod-borne viruses. Our data indicate that the RNAi machinery controls infection by DNA viruses in Drosophila . We propose that sense and antisense viral transcripts base-pair, thus forming viral dsRNA that is recognized as nonself and processed into small RNAs by the antiviral RNAi machinery ( Fig. P1 ). Here, we used D. melanogaster to address these questions. We first analyzed the course of infection by the DNA virus Invertebrate iridescent virus 6 (IIV-6) in wild-type (WT) flies. The number of infectious virus particles increased rapidly during the first 6 days (d) and remained stable thereafter. Nevertheless, the flies did not succumb to infection over the course of 30 d. We then analyzed whether the RNAi machinery affected the course of infection by infecting flies that lack a functional RNAi response due to a disruption of the genes encoding the central RNAi components Dicer-2 and Argonaute-2. These flies died earlier than the WT, suggesting that RNAi indeed controls infection by IIV-6. We then analyzed whether viral small RNAs are produced by the RNAi machinery. In infected flies, viral small RNAs were readily detectable and, as expected, their presence depended on the function of the dsRNA-processing enzyme Dicer-2. What is the source of these small RNAs? The small RNA sequences allowed us to determine their origin by aligning them to the viral genome. We found that small RNAs mapped along the entire viral genome to both the viral sense and antisense transcripts. Further, using assays that directly detect both transcripts, we confirmed the occurence of antisense transcription in flies infected with IIV-6. Double-stranded (ds) RNA is a typical danger signal that alerts the immune system to ongoing virus infection. Uninfected cells produce single-stranded (ss) RNA but not dsRNA. The genome of RNA viruses goes through a double-stranded intermediate stage during the amplification cycle. The RNAi mechanism recognizes viral dsRNA and cuts it into smaller pieces called viral small interfering RNA. These small RNAs are then used to find and destroy viral ssRNAs. By degrading viral RNAs, the RNAi machinery prevents infection of a cell and reduces the amount of virus produced. Although DNA viruses do not rely on an intermediate that is composed of dsRNA, they are known to produce dsRNA during infection ( 3 ). To produce viral proteins, the viral DNA genome is transcribed into mRNAs that are the template for protein synthesis. Nevertheless, transcription of the complementary strand that does not encode a functional protein (antisense transcription) also seems to occur. Overlapping sense and antisense transcripts have the potential to base-pair and form dsRNA. Open questions are as follows: Are DNA viruses recognized by an antiviral RNAi response? Can we detect viral small RNAs in DNA virus infection? How does the RNAi machinery affect the course of infection? Viruses can infect and cause disease in virtually all organisms. Survival of the infected host requires an efficient defense that is able to distinguish components and structures of the virus (nonself) from those of the host (self). Insects, including the model organism Drosophila melanogaster (fruit fly), rely on an RNA-based response called RNA interference (RNAi) for their defense against viruses with an RNA genome (RNA virus) ( 1 , 2 ). However, it is not clear whether viruses with a DNA genome are fought by the same antiviral mechanism, and if so, how these viruses are recognized. We showed here that the RNAi machinery controls infection by DNA viruses in Drosophila.
Type of Medium:
Online Resource
ISSN:
0027-8424
,
1091-6490
DOI:
10.1073/pnas.1207213109
Language:
English
Publisher:
Proceedings of the National Academy of Sciences
Publication Date:
2012
detail.hit.zdb_id:
209104-5
detail.hit.zdb_id:
1461794-8
SSG:
11
SSG:
12
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