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. 2020 Jan 17;94(3):e01233-19. doi: 10.1128/JVI.01233-19

FIG 1.

FIG 1

SFV capsid protein is a potential VSR. (A) S2 cells were cotransfected with a plasmid encoding EGFP (0.1 μg) and dsEGFP (0.3 μg), together with either empty plasmid or a plasmid encoding SFV protein or FHV B2 (FB2) (1 μg for each). At 48 hpt, total RNAs were extracted, and the level of EGFP mRNA was examined by Northern blotting with a DIG-labeled RNA probe targeting nt 500 to 720 of the EGFP ORF region. Rp49 mRNA was used as loading control. (B) The expression of SFV proteins was detected by Western blotting. (C) S2 cells were transfected with pMT FHV RNA1 (FHV RNA1) (0.01 μg) or pMT FHV ΔB2 RNA1 (FR1 ΔB2) (0.3 μg), together with a plasmid encoding SFV capsid or FB2, as indicated above. At 48 hpt, FHV RNA transcription was induced by incubation with CuSO4 (0.5 mM). At 24 h after induction, the total RNAs were harvested for Northern blot analysis. The band between RNA1 and RNA3 was the B2 mRNA transcribed from the expression plasmid. The dsRNAs targeting AGO2 (dsAGO2) and Dicer2 (dsDicer2) were used as positive controls. (D) 293T cells were cotransfected with a plasmid encoding EGFP (0.1 μg) and EGFP-specific shRNA (shEGFP) (0.3 μg), together with either empty plasmid or a plasmid encoding SFV capsid protein, nsP2, nsP3, or NB2 (1 μg for each). At 48 hpt, the total RNAs were extracted, and the level of EGFP mRNA was examined by Northern blotting. (E) The expression of SFV nsP2, nsP3, and capsid proteins in 293T cells was detected by Western blotting. (F) S2 cells were cotransfected with a plasmid encoding EGFP (0.1 μg) and EGFP-dsRNA (dsEGFP) (0.3 μg), together with either empty plasmid or a plasmid encoding SINV capsid protein, SFV capsid protein, or FB2 (1 μg for each). At 48 hpt, the total RNAs were extracted, and the level of EGFP mRNA was examined by Northern blotting. Rp49 mRNA was used as the loading control. (G) 293T cells were cotransfected with a plasmid encoding EGFP (0.1 μg) and EGFP-specific shRNA (shEGFP) (0.3 μg), together with either empty plasmid or a plasmid encoding SINV capsid protein, SFV capsid protein, or NB2 (1 μg for each). At 48 hpt, the total RNAs were extracted, and the level of EGFP mRNA was examined by Northern blotting. GAPDH mRNA was used as the loading control.