Fig. 7.

Effect of MNF treatment, cell confluence, and AhR knockdown on luciferase expression from a reporter plasmid containing 7073 nt of the SULT1E1 5′-flanking sequence. A, MCF10A cells were stably transfected with the SULT1E1-luciferase reporter plasmid, and preconfluent cells were treated with 1 to 10 μM MNF for 24 h, after which they were harvested for measurement of firefly luciferase activity and protein content. Each bar represents the mean relative luciferase activity (normalized to cellular protein content) ± S.D. (three wells per treatment group). Groups that do not share a capital letter are significantly different from each other (p < 0.001). B, preconfluent and confluent MCF10A cells stably transfected with the SULT1E1-luciferase reporter were harvested for measurement of luciferase activity as described in A. ***, p < 0.001 compared with preconfluent cells. C, preconfluent MCF10A cells were transiently transfected with the SULT1E1-luciferase reporter containing either an intact (□) or mutated (■) predicted DRE at −3476, treated for 24 h with 1 to 10 μM MNF, and harvested for measurement of luciferase activities. Each bar represents the mean relative luciferase activity (firefly/Renilla) ± S.D. (three wells per treatment group). Groups that do not share a capital letter are significantly different from each other (p < 0.05). D, preconfluent MCF10A cells engineered for conditional knockdown of AhR were treated with either 0.1% DMSO or 1 μg/ml doxycycline for 96 h and transiently transfected with SULT1E1-luciferase reporter containing intact or mutant DRE. At 24 h after transfection, cells were harvested for measurement of firefly and Renilla luciferase activities. Groups that do not share a capital letter are significantly different from each other (p < 0.001).