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Originally published in Science Express on 23 March 2006
Science 21 April 2006:
Vol. 312. no. 5772, pp. 452 - 454
DOI: 10.1126/science.1125694
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Reports

RNA Interference Directs Innate Immunity Against Viruses in Adult Drosophila

Xiao-Hong Wang1*, Roghiyh Aliyari2*, Wan-Xiang Li2, Hong-Wei Li2, Kevin Kim4, Richard Carthew4, Peter Atkinson3 and Shou-Wei Ding1,2{dagger}

1 Graduate Program for Microbiology, University of California, Riverside, CA 92521, USA.
2 Department of Plant Pathology and Center for Plant Cell Biology, Institute for Integrative Genome Biology, University of California, Riverside, CA 92521, USA.
3 Department of Entomology, University of California, Riverside, CA 92521, USA.
4 Department of Biochemistry, Molecular Biology, and Cell Biology, Northwestern University, Evanston, IL 60208, USA.


Figure 1 Fig. 1. Antiviral silencing in Drosophila embryos requires Dcr-2 and Ago-2. Northern blot detection of FHV RNA accumulation in wt, dcr-2L811fsX, and ago-2414 embryos microinjected with in vitro synthesized transcripts R1, R1fs, and R1{Delta}B2, as shown in fig. S1. [View Larger Version of this Image (54K GIF file)]
 

Figure 2 Fig. 2. The siRNA pathway controls the innate immunity against FHV in adult flies. (A) Survival of wt, dcr-2L811fsX, and r2d2 mutant flies after FHV infection. Data shown represents mean of triplicates, and the error bars indicate standard deviation. (B and C) Detection of viral RNAs by a probe specific for RNAs 1 and 3 and viral coat protein (CP) by a rabbit anti-FHV serum (18) in wt, dcr-2L811fsX, and r2d2 flies injected with FHV. (D) Detection of FHV siRNAs in the infected adult flies as in (C) (d2, dcr-2; r2, r2d2). The membrane was also probed for microRNA-8 (miR-8) and U6 RNA. nt, nucleotides. [View Larger Version of this Image (35K GIF file)]
 

Figure 3 Fig. 3. Mutant dcr-2 and r2d2 flies exhibit enhanced disease susceptibility to CrPV. (A) Survival of wt, dcr-2L811fsX, and r2d2 mutant flies after CrPV infection. Data shown represents mean of triplicates, and the error bars indicate standard deviation. (B) Accumulation of CrPV genomic RNA in WT, dcr-2L811fsX, and r2d2 mutant adults injected with CrPV. [View Larger Version of this Image (37K GIF file)]
 

Figure 4 Fig. 4. Induction and suppression of antiviral silencing by CrPV. (A) Accumulation of the genomic RNA and siRNAs of CrPV in infected S2 cells. (B) pFR1gfp directed transcription of a recombinant FHV RNA1 in which the coding sequence for B2 was replaced by that of GFP. S2 cells were transfected with pFR1gfp alone (middle) or pFR1gfp plus either dsRNA-targeting Ago-2 (right) or CrPV superinfection (left). (C) Identification of CrPV RNAi suppressor. Cells were cotransfected with pFR1gfp and a plasmid as indicated on top of each lane (fig. S2), and total RNA was analyzed for the accumulation of pFR1gfp-encoded RNA1 and RNA3. P, the empty plasmid; VP, the virion protein precursor; V1, VP1; V2, VP2; V3, VP3; V4, VP4; V0, VP0 (precursor for VP3 and VP4); A, the first 140 codons of the upstream ORF; {Delta}A, a frameshift mutant of A; and b, the first 107 codons of the upstream ORF. NS1 is an RNAi suppressor of influenza A virus as described previously (11). [View Larger Version of this Image (25K GIF file)]
 

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