FIGURE 7.

SN50, but not SN50M, inhibits Nrf2-induced transcription from ARE-driven gene promoters. In A, K562 cells were co-transfected with hNQO1-ARE-luc reporter gene construct (0.3 μg) without or with plasmid (0.2 μg) containing cDNA for Nrf2 (pCI-Nrf2) or the empty vector (pCI-Neo) followed by the addition of 5 μm SN50 or SN50M 1 h later. In B, the cells were co-transfected with pGα_i2(–1214/+115)-luc reporter gene construct (0.2 μg) without or with plasmid (0.3 μg) containing cDNA for Nrf2 (PCI-Nrf2) or the empty vector (PCI-Neo) followed by the addition of 5 μm SN50 or SN50M 1 h later. For both panels, the cells were harvested after 24 h and processed for luciferase assay as described under “Experimental Procedures.” The hNQO1-ARE-luc reporter construct contains a single copy of ARE, derived from the human NQO1 promoter, placed upstream of a minimal promoter containing a TATA box fused to the luciferase gene (41, 42). The Gα_i2-luc reporter gene construct contains only one ARE sequence (5′-TGACTGGGC-3′) that maps at –84/–76 in the promoter (27, 43). Values shown are means ± S.E. for duplicate assays from four different experiments. Values obtained for cells that were transfected with the empty vector (pCI-Neo) were set as 1.0. Con, control (no peptide added).