Figure 5.

Luciferase reporter assays of cloned genomic loci bound by ERα and displaying moderate, low, and no enhancer activity in MCF7 cells. Genomic loci (average length 776 bp) containing one or more ERE sequences were cloned into a minimal promoter reporter construct driving luciferase gene expression. Specific wild-type ERE sequences and mutated (i.e. E1M) ERE sequences are indicated above each clone being tested (repetitive element EREs are indicated by red font, mismatches from the classical ERE sequence are indicated by lower case lettering, and mutated sequences were changed to tttttt where indicated). Basal and E2-stimulated luciferase values are shown normalized to co-transfected β-galactosidase-expressing plasmid. D75 contains two repetitive element ERE sequences responsible for enhancer function (A). C31 contains two, nonrepetitive element EREs that are functional (B). Additional ERE-containing clones with low or no enhancer function in luciferase assays are indicated (C). Values are the average of three experiments, performed in triplicate, with SEM indicated. *P < 0.01 comparing E2-treated wild-type with E2-treated mutated reporter. ^§P < 0.01 comparing E2-treated empty vector with E2-treated mutated reporter. ^†P < 0.01 comparing vehicle control-treated with E2-treated reporter.