Activation of estrogen receptor-alpha by the anion nitrite.
In this study, the ability of nitrite and nitrate to mimic the effects of estradiol on growth and gene expression was measured in the human breast cancer cell line MCF-7. Similar to estradiol, treatment of MCF-7 cells with either 1 mumol/L nitrite or 1 mumol/L nitrate resulted in approximately 4-fold increase in cell growth and 2.3-fold to 3-fold increase in progesterone receptor (PgR), pS2, and cathepsin D mRNAs that were blocked by the antiestrogen ICI 182,780. The anions also recruited estrogen receptor-alpha (ERalpha) to the pS2 promoter and activated exogenously expressed ERalpha when tested in transient cotransfection assays. To determine whether nitrite or nitrate was the active anion, diphenyleneiodonium was used to inhibit oxidation/reduction reactions in the cell. The ability of diphenyleneiodonium to block the effects of nitrate, but not nitrite, on the induction of PgR mRNA and the activation of exogenously expressed ERalpha suggests that nitrite is the active anion. Concentrations of nitrite, as low as 100 nmol/L, induced a significant increase in PgR mRNA, suggesting that physiologically and environmentally relevant doses of the anion activate ERalpha. Nitrite activated the chimeric receptor Gal-ER containing the DNA-binding domain of GAL-4 and the ligand-binding domain of ERalpha and blocked the binding of estradiol to the receptor, suggesting that the anion activates ERalpha through the ligand-binding domain. Mutational analysis identified the amino acids Cys381, His516, Lys520, Lys529, Asn532, and His547 as important for nitrite activation of the receptor.
Veselik
David J
DJ
Department of Biochemistry and Molecular and Cellular Biology, Georgetown University, Washington, District of Columbia, USA.
Divekar
Shailaja
S
Dakshanamurthy
Sivanesan
S
Storchan
Geoffrey B
GB
Turner
Jasmine M A
JM
Graham
Kelly L
KL
Huang
Li
L
Stoica
Adriana
A
Martin
Mary Beth
MB
D000596:Amino Acids / Q000737:chemistry; D000818:Animals; D000838:Anions; D019556:COS Cells; D002402:Cathepsin D / Q000378:metabolism; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D004958:Estradiol / Q000031:analogs & derivatives / Q000494:pharmacology; D047628:Estrogen Receptor alpha / Q000378:metabolism*; D000077267:Fulvestrant; D006801:Humans; D008968:Molecular Conformation; D009573:Nitrites / Q000737:chemistry*; D011401:Promoter Regions, Genetic; D011980:Receptors, Progesterone / Q000096:biosynthesis; D014162:Transfection
D000596:Amino Acids; D000838:Anions; C506487:ESR1 protein, human; D047628:Estrogen Receptor alpha; D009573:Nitrites; D011980:Receptors, Progesterone; D000077267:Fulvestrant; D004958:Estradiol; D002402:Cathepsin D