Role of NF-κB pathway in K13-induced CCL20 promoter activation. (A) Dominant-negative mutants of IκBα (IκBαΔN and IκBαSS32/36AA) block K13-induced CCL20 promoter activity. The 293T cells were transfected either with an empty vector or K13, along with a CCL20 luciferase reporter construct and a β-galactosidase reporter construct, as described in Figure 3A. The amount of IκBα mutant plasmids (500 ng/well) was 5 times the amount of vector or K13 (100 ng/well) plasmid, and the total amount of transfected DNA was kept constant by adding empty vector. The values shown are averages (mean ± SE) of 1 representative experiment of 3 in which each transfection was performed in duplicate. (B,C) Pharmacologic inhibitors of NF-κB block K13-induced CCL20 promoter activation. The 293T cells were transfected with an empty vector or a vector encoding K13, and 30 minutes after transfection treated with dimethyl sulfoxide (vehicle) or the indicated compounds for 16 hours before cell lysis. Reporter assay was performed as described for Figure 3A. (D) HUVECs were pretreated with Bay-11-7082 (5 μM) or As₂O₃ (2.5 μM) for 2 hours and subsequently infected with KSHV as described previously.³¹  Four hours after infection, the medium was changed with fresh medium containing Bay-11-7082 or As₂O₃. After overnight incubation, supernatant was collected and CCL20 secretion was measured as described in Figure 1B.