FIG 1.

VP23 inhibits IFN-β activation induced by cGAS-STING. (A) CEFs were infected with MDV at a multiplicity of infection (MOI) of 0.1. IFN-β mRNA and protein levels were measured by real-time qPCR and ELISA, respectively, from 4 h to 72 h after infection. The expression of MDV protein VP23 and gI during viral infection was monitored by immunoblotting. (B) CEFs were transfected with cGAS siRNA (sicGAS), STING siRNA (siSTING), or a nonspecific control (NC) siRNA (siNC) and then infected with MDV (MOI = 0.1). IFN-β mRNA was examined by real-time qPCR at 12 h postinfection, and the IFN-β protein was examined by ELISA at 24 h postinfection. (C) HD11 cells were transfected with cGAS or STING siRNA or a nonspecific control siRNA and then transfected with MDV DNA fragments. IFN-β mRNA was examined by real-time qPCR at 8 h posttransfection, and the IFN-β protein was examined by ELISA at 24 h posttransfection. (D) The knockdown efficiency of cGAS and STING in CEFs was monitored by real-time qPCR and immunoblotting. (E) The knockdown efficiency of cGAS and STING in HD11 cells was monitored by immunoblotting. (F) Various doses of the VP23 expression plasmid or MDV gI expression plasmid were cotransfected with the IFN-β-luc reporter with or without cGAS-STING stimuli into DF-1 cells, and IFN-β promoter luciferase activity was measured at 24 h posttransfection. (G) VP23, the gI expression plasmid, or the empty vector was cotransfected with cGAS and STING plasmids into DF-1 cells. IFN-β mRNA and protein levels were measured by real-time qPCR or ELISA at 24 h posttransfection. (H) Various doses of the VP23 expression plasmid or the MDV gI expression plasmid were cotransfected with the IFN-β-luc reporter with or without cGAS-STING stimuli into HD11 cells, and IFN-β promoter luciferase activity was measured at 24 h posttransfection. (I) VP23, the gI expression plasmid, or the empty vector was cotransfected with the cGAS and STING plasmids into HD11 cells. IFN-β mRNA and protein levels were measured by real-time qPCR or ELISA at 24 h posttransfection. (J) The expression of the transfected plasmids in DF-1 and HD11 cells was detected by Western blotting. The relative amounts of IFN-β, cGAS, and STING mRNA were normalized to the actin mRNA level in each sample, and the fold differences compared with the amounts in the mock-transfected samples were determined. Data are presented as the mean ± SD from three independent experiments. Statistical analysis was performed by using Student's t test (**, P < 0.01; ns, no significant difference).