Effects of arsenite on estrogen receptor-alpha expression and activity in MCF-7 breast cancer cells.
To determine whether arsenite has estrogen-like activities, the effects of this compound on estrogen receptor-alpha (ERalpha) and other estrogen-regulated genes were measured in the human breast cancer cell line MCF-7. Treatment of cells with 1 microM arsenite resulted in a 60% decrease in the amount of ERalpha and in a parallel decrease of 40% in ERalpha messenger RNA. Progesterone receptor concentration increased 22-fold after arsenite treatment. pS2 messenger RNA also increased 2. 1-fold after treatment. The induction of progesterone receptor and pS2 was blocked by the antiestrogen ICI-182,780. In transient cotransfection experiments of wild-type ERalpha and an estrogen response element-reporter construct, arsenite stimulated chloramphenicol acetyltransferase (CAT) activity. In growth assays, arsenite significantly stimulated the proliferation of MCF-7 cells compared with cells grown in estrogen-depleted medium. Addition of an antiestrogen blocked growth stimulation by arsenite. In binding assays, arsenite blocked the binding of estradiol to ERalpha (Ki = 5 +/- 0.5 nM; n = 3), suggesting that the compound interacts with the hormone-binding domain of the receptor. To determine whether interaction of arsenite with the hormone-binding domain results in receptor activation, COS-1 cells were transiently cotransfected with the chimeric receptors GAL-ER, which contains the hormone-binding domain of ERalpha and the DNA-binding domain of the transcription factor GAL4, and a GAL4-responsive CAT reporter gene. Treatment of cells with estradiol or arsenite resulted in a 4-fold increase in CAT activity. The effects of arsenite on the chimeric receptor were blocked by the antiestrogen, suggesting that arsenite activates ERalpha through an interaction with the hormone-binding domain of the receptor. Transfection assays with ERalpha mutants identified C381, C447, H524, and N532 as interaction sites of arsenite with the hormone-binding domain.
Stoica
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Department of Biochemistry and Molecular Biology, Lombardi Cancer Center, Georgetown University, Washington, DC 20007, USA.
Pentecost
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Martin
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MB
D000818:Animals; D018053:Arsenites / Q000494:pharmacology*; D001943:Breast Neoplasms / Q000378:metabolism* / Q000473:pathology; D019556:COS Cells; D002455:Cell Division / Q000187:drug effects; D004958:Estradiol / Q000378:metabolism; D047628:Estrogen Receptor alpha; D005260:Female; D006706:Homeostasis; D006728:Hormones / Q000378:metabolism; D006801:Humans; D009154:Mutation / Q000187:drug effects / Q000502:physiology; D009994:Osmolar Concentration; D011506:Proteins / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D011960:Receptors, Estrogen / Q000235:genetics / Q000378:metabolism*; D011980:Receptors, Progesterone / Q000378:metabolism; D020218:Response Elements / Q000502:physiology; D000071163:Trefoil Factor-1; D014407:Tumor Cells, Cultured; D025521:Tumor Suppressor Proteins
D018053:Arsenites; D047628:Estrogen Receptor alpha; D006728:Hormones; D011506:Proteins; D012333:RNA, Messenger; D011960:Receptors, Estrogen; D011980:Receptors, Progesterone; C051838:TFF1 protein, human; D000071163:Trefoil Factor-1; D025521:Tumor Suppressor Proteins; D004958:Estradiol; C015001:arsenite