PKACα or PKACβ inhibits SeV-induced and VISA-mediated signaling.

(A) Screening of kinase cDNA clones that inhibit VISA-mediated activation of the IFN-β promoter. HEK293 cells were transfected with the IFN-β promoter luciferase plasmid, mammalian expression plasmids for VISA and the kinase cDNA clones for 20 h before luciferase assays were performed. Representative data is shown. (B) PKACα and PKACβ inhibit SeV-triggered activation of the IFN-β promoter and ISRE but not NF-κB. HEK293 cells were transfected with the indicated reporter plasmids and increased amounts of Flag-PKACα/β plasmids for 20 h, then infected with SeV (MOI = 1) or left untreated for 12 h before luciferase assays were performed. The lower blots show the expression levels of transfected PKACα/β as detected by anti-Flag antibody. (C) PKACα and PKACβ inhibit SeV-triggered induction of downstream antiviral genes. HEK293 cells were transfected with the indicated expression plasmids for 20 h, then infected with SeV (MOI = 1) or left untreated for 8 h before qPCR was performed. (D) Effects of PKA subunits and mutants on SeV-triggered activation of the IFN-β promoter. HEK293 cells were transfected with the IFN-β promoter luciferase plasmid and the indicated Flag-tagged expression plasmids for 20 h, then infected with SeV (MOI = 1) or left untreated for 12 h before luciferase assays were performed. The lower blot shows the expression levels of the transfected proteins.