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Figure 2 | Veterinary Research

Figure 2

From: DHAV 3CD targets IRF7 and RIG-I proteins to block the type I interferon upstream signaling pathway

Figure 2

The 3CD protein interacts with IRF7 protein and inhibits the expression of IRF7 protein.A pCAGGS-3CD-HA and pCAGGS-IRF7-Flag were co-transfected or both were separately co-transfected with pCAGGS into 24-well plates (transfection ratio of 1:1), and cell samples were collected 36 h after transfection for indirect immunofluorescence experiments to observe the intracellular localization of the 3CD protein and IRF7 protein. B Cells were grown in 6-well plates and each well was transfected with a total of 1 µg pCAGGS-3CD-HA and 1 µg pCAGGS-IRF7-Flag. Cells were lysed 36 h after transfection, and lysates were immunoprecipitated with mouse anti-Flag or mouse IgG antibodies and subjected to Western blotting. C 0.4 µg pCAGGS-IRF7-Flag was co-transfected into 12-well plates with 0.6 µg pCAGGS-3CD-HA, pCAGGS-3D-HA, pCMV-3 C-HA or empty, respectively, and cell samples were collected 36 h after transfection to detect the expression of IRF7 protein. D Similar transfection and quantitative PCR experiments were performed to analyze the effect of 3CD protein on Poly(I:C)-induced IRF7 mRNA levels as described in Figure 1A. * P < 0.05, ** P < 0.01, *** P < 0.001, compared with the control group. E DEFs were grown in 12-well plates, and different doses of 3CD-HA expression plasmids were transferred into the cells (the amount transfected into each group was 1 µg, and pCAGGS was used to supplement the insufficient group). The expression of 3CD protein and endogenous IRF7 protein was detected by Western blotting at 36 h. F DEFs were grown in 12-well plates, and 0.4 µg of pCAGGS-IRF7-Flag was co-transfected with different doses of pCAGGS-3CD-HA (the amount transfected into each group was 1 µg, and pCAGGS was used to supplement the insufficient group). 36 h after transfection, cell samples were collected, and the protein expressions of 3CD and IRF7 were detected.

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