IFNs inhibit the association of transcription factor GABP with the E1A enhancer region.
(A and B) HDF-TERT cells were treated with IFNs or left untreated for 24 hr and then infected with dl309 at 200 virus particles/cell. ChIP assays were performed with samples isolated at 18 hr post-infection using an antibody directed against GABPα or an isotype-matched control (A), or using an antibody directed against Sp1 or an isotype-matched control (B). Immunoprecipitated DNA was quantified by qPCR and the enrichment is represented as percentage of input. The values are plotted as mean ± sd, n = 3. (C) HDF-TERT cells were treated with IFNs or left untreated. Cellular extracts were prepared at 24 hr post-treatment and analyzed by Western blot using antibodies directed against GABPα and GABPβ. γ-tubulin was used as a loading control for the samples. (D) HDF-TERT cells were treated with IFNs or left untreated for 24 hr and then infected with dl309 200 virus particles/cell or mock-infected. Nuclear (N) and cytoplasmic (C) fractions were prepared 18 hr post-infection. Western blot analyses were performed using antibodies against GABPα and GABPβ. Histone H3 was used a nuclear marker to verify the subcellular fractionation. (E) IFN treatment and infection of HDF-TERT cells was carried out as described in (D). Total cell extracts were isolated at 18 hr post-infection and used for immunoprecipitation (IP) with an antibody against GABPα or an isotype-matched control. Immunocomplexes were analyzed for GABPβ coIP by Western blot. The top panel shows GABPα and GABPβ protein levels in cell lysates (Input). The bottom panel shows the results of the IPs.