Activation of estrogen receptor-α by the heavy metal cadmium

Previous studies from this laboratory have shown that the heavy metal cadmium (Cd) mimics the effects of estradiol in estrogen-responsive breast cancer cell lines. To understand the mechanism by which cadmium activates estrogen receptor-α (ER-α), the ability of cadmium to bind to and activate wild-type and various mutants of ER-α was examined. When tested in transient cotransfection assays in COS-1 cells, cadmium concentrations as low as 10^-11 M activated ER-α. Scatchard analysis employing either purified human recombinant ER-α or extracts from ER-containing MCF-7 cells demonstrated that ¹⁰⁹Cd binds to the ER with an equilibrium dissociation constant of approximately 4 to 5 × 10^-10 M. Cadmium also blocks the binding of estradiol to ER-α in a noncompetitive manner (K_i = 2.96 × 10^-10 M), suggesting that the heavy metal interacts with the hormone-binding domain of the receptor. To study the role of the hormone-binding domain in cadmium activation, COS-1 cells were transiently cotransfected with GAL-ER, a chimeric receptor containing the DNA-binding domain of the transcription factor GAL4 and the hormone-binding domain of ER-α, and a GAL4-responsive reporter gene. Treatment of the transfected cells with either 10^-6 M cadmium or 10^-9 M estradiol resulted in a 4-fold increase in reporter gene activity. The effect of cadmium on the chimeric receptor was blocked by the antiestrogen, ICI-164,384, suggesting that cadmium activates ER-α through an interaction with the hormone-binding domain of the receptor. Transfection and binding assays with ER-α mutants identified C381, C447, E523, H524, and D538 as possible interaction sites of cadmium with the hormone-binding domain of ER-α.