The siRNA pathway controls the innate immunity against FHV in adult flies. (A) Survival of wt, dcr-2 and r2d2 flies after FHV infection. Data shown represented mean of triplicates and the error bars indicate standard derivation. (B, 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-2, 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 miR-8 and U6 RNA.

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 co-transfected with pFR1gfp and a plasmid as indicated (see Fig. s2), and total RNA 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; ΔA, 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).

Antiviral silencing in Drosophila embryos requires Dcr-2 and Ago-2. Northern blot detection of FHV RNA accumulation in wt, dcr-2, and ago-2 embryos micro-injected with in vitro synthesized transcripts R1, R1fs and R1ΔB2 as shown in Fig. s1.

dcr-2 and r2d2 flies exhibit enhanced disease susceptibility to CrPV. (A) Survival of wt, dcr-2 and r2d2 flies after CrPV infection. Data shown represented mean of triplicates and the error bars indicate standard derivation. (B) Accumulation of CrPV genomic RNA in wt, dcr-2, and r2d2 adults injected with CrPV.