Avian oncogenic herpesvirus antagonizes the cGAS-STING DNA-sensing pathway to mediate immune evasion
Fig 2
Screening of MDV open reading frames (ORFs) that modulate the cGAS-STING pathway.
(A) DF-1 cells were cotransfected with IFN-β promoter luciferase reporter and various plasmids (pCAGGS or pCAGGS-cGAS-HA and pCAGGS-STING-HA combined). The luciferase activity was measured at 36 h posttransfection. (B) Schematic of the screening assay. DF-1 cells were transfected with the same amount of cGAS and STING expression plasmid, plus each of MDV ORF expression plasmid or the empty vector. (C) Heat map of the effects of MDV ORFs on the cGAS-STING pathway. Higher IFN-β promoter luciferase activation levels are indicated by red, whereas lower levels are indicated by blue, which corresponds to a higher degree of inhibition. (D) The top five MDV ORF inhibitors and the MDV gI ORF were cotransfected with cGAS and STING expression plasmids into DF-1 cells. At 36 h posttransfection, IFN-β mRNA levels were measured by real-time qPCR. The relative amount of IFN-β mRNA was normalized to the actin mRNA level in each sample, and the fold changes were compared with those in the mock controls. (E) The top five MDV ORF inhibitors and the gI ORF were cotransfected with cGAS and STING expression plasmids into DF-1 cells, and IFN-β protein levels were measured by enzyme-linked immunosorbent assay 36 h posttransfection. (F) Varying doses of the top five MDV ORF inhibitors and the gI ORF were cotransfected with cGAS and STING expression plasmids, and IFN-β promoter luciferase activity was measured at 36 h posttransfection. (G) The top five MDV ORF inhibitors and the gI ORF were transfected into DF-1 cells, and IFN-β promoter luciferase activity was measured at 36 h posttransfection. *: p < 0.05, **: p < 0.01, ***: p < 0.001; ns: no significant difference.