Figure 3.

HMBA augments prostratin-activated NF-κB by promoting prostratin-induced IκBα degradation. (a) HMBA prolongs prostratin-induced nuclear translocation of RelA. HeLa cells with indicated treatments were assayed by anti-RelA immunofluorescence for the nuclear translocation of RelA. (b) HMBA enhances prostratin-activated recruitment of RelA on HIV-1 promoter. HIV-LTR-Luc cells with indicated treatments were assayed by anti-RelA chromatin immunoprecipitation (ChIP) for the enrichment of RelA on HIV-1 promoter as in Figure 2(b). (c) HMBA promotes prostratin-induced IκBα degradation. The levels of IκBα were analyzed by Western Blot (WB) using the cell lysates prepared from HeLa cells with indicated treatments. (d) Inhibiting IκBα degradation by MG-132 blocks HMBA and prostratin cotreatment- (H+P-) activated HIV-1 expression. HIV-LTR-Luc cells with indicated pretreatment and treatment were subjected to luciferase assay for HIV-1 expression as in Figure 1(d). (e) Inhibiting IKK blocks H+P-induced IκBα degradation. The levels of IκBα were analyzed by WB using the cell lysates derived from HeLa cells with IKK inhibitor BAY preincubation, followed by indicated treatments.