Determining the transrepression activity of xenoestrogen on nuclear factor-kappa B in Cos-1 cells by estrogen receptor-alpha.
Functional assays have been used to define the estrogenicity of xenoestrogens in cotransfection studies employing estrogen receptors in various cell lines. It is known that estrogen is able to affect transcription from other nuclear transcription factors, especially the nuclear factor-kappa B (NF-kappa B). The ability of selected xenoestrogens (methoxychlor [MXC], dieldrin, and o',p'-DDT) to transrepress the NF-kappa B-mediated transcription in Cos-1 cells was evaluated by cotransfection of human estrogen receptor-alpha (hERalpha). These xenoestrogens have been described as comparably potent xenoestrogens, whereas their relative binding activity (RBA) has been relegated to a lower order as compare to estrogen. The two NF-kappa B response element-containing SV40 promoter and -242/+54 cytomegalovirus (CMV)-expressing firefly luciferase (2 x NRE-PV-Luc and 2 x NRE-CMV-Luc, respectively) were transfected into Cos-1 cells with pRL-tk, expressing the renilla luciferase as internal control. The estrogen receptor was expressed from cytomegalovirus major immediate early promoter (CMV-MIEP) (CMV5-hERalpha). Treatment with 1 nM estrogen (E(2)) (26.2%), 5 nM E(2) (41.4%; p < .05), and xenoestrogens (methoxychlor [1 nM: 29.6%, p < .05; 10 nM: 22.6%), dieldrin [1 nM: 10.3%; 10 nM: 36.06%, p < .05], and o',p'-DDT [1 nM: 17.0%; 10 nM: 7.15%]) repressed transcription from 2 x NREX-PV-Luc. The antiestrogen, ICI 182,780, failed to antagonize the effects of xenoestrogens. The effects of xenoestrogens in transrepression of NF-kappa B by ERalpha were similar when 2 x NRE-CMV-Luc was employed as reporter. Statistically significant (p < .01) repression by 1 nM E(2) (69.2%), 5 nM E(2) (69.1%), 1 nM o',p'-DDT (51.4%), 1 nM dieldrin (47.3%), and 1 nM MXC (73.3%) were observed. The effect of these xenoestrogens without ERalpha cotransfection on 2 x NRE-PV-Luc- and 2 x NRE-CMV-Luc-mediated NF-kappa B transcription was not affected by the treatment alone. It is concluded that xenoestrogens, like estrogens, are capable of producing transrepression of NF-kappa B by hERalpha.
Ansari
R A
RA
Department of Pharmacology and Toxicology, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA. ansarirais@hotmail.com
Gandy
J
J
D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D004026:Dieldrin / Q000494:pharmacology; D024202:Electrophoretic Mobility Shift Assay; D004958:Estradiol / Q000031:analogs & derivatives / Q000494:pharmacology; D004965:Estrogen Antagonists / Q000494:pharmacology; D047628:Estrogen Receptor alpha / Q000235:genetics / Q000502:physiology*; D004967:Estrogens / Q000494:pharmacology; D000077267:Fulvestrant; D005786:Gene Expression Regulation / Q000187:drug effects; D049409:Luciferases, Firefly / Q000235:genetics / Q000378:metabolism; D008731:Methoxychlor / Q000494:pharmacology; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D010703:Phorbol Esters / Q000494:pharmacology; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D020218:Response Elements / Q000235:genetics; D020845:Selective Estrogen Receptor Modulators / Q000494:pharmacology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects; D014162:Transfection
D004965:Estrogen Antagonists; D047628:Estrogen Receptor alpha; D004967:Estrogens; D016328:NF-kappa B; D010703:Phorbol Esters; D011993:Recombinant Fusion Proteins; D020845:Selective Estrogen Receptor Modulators; D000077267:Fulvestrant; D004958:Estradiol; D049409:Luciferases, Firefly; D004026:Dieldrin; D013755:Tetradecanoylphorbol Acetate; D008731:Methoxychlor