Differential action of polycyclic aromatic hydrocarbons on endogenous estrogen-responsive genes and on a transfected estrogen-responsive reporter in MCF-7 cells.
Polycyclic aromatic hydrocarbons (PAHs) are common environmental pollutants that have been extensively studied for multiple toxicological endpoints in both laboratory animals and humans. The purpose of this study was to investigate the estrogenicity of PAHs in the human breast cancer cell line MCF-7. We investigated 14 PAHs for their ability to bind either the estrogen receptor (ER) or the aryl hydrocarbon receptor (AhR) and to activate target gene expression. PAHs were tested in a human recombinant estrogen receptor (hrER) competitive binding assay, and in both an estrogen response element (ERE)- and xenobiotic response element (XRE)-mediated reporter gene assay. We used quantitative RT-PCR to examine selected PAHs that showed activity in the ERE reporter gene assay for their ability to upregulate estrogen-responsive genes HEM45, progesterone receptor, and pS2, and the aryl hydrocarbon-responsive CYP1A1 gene. None of the 14 PAHs bound the hrER, but five of the PAHs (anthracene, B[a]A, chrysene, B[b]F, and B[a]P) induced ER-reporter activity. This activity was dependent on the metabolism of PAHs in MCF-7 cells via the AhR pathway, which resulted in the formation of metabolites that bound the ER. None of the five PAHs that induced the ER-reporter were found to upregulate estrogen-responsive genes, yet four of the five PAHs induced AhR-dependent CYP1A1 gene expression. In contrast, a metabolite of B[a]P, 3'OH-B[a]P, and a PCB metabolite, 4'OH-2,4,6-BP, did weakly upregulate all three estrogen-responsive genes. Data from these studies indicate that induction of ER-reporter activity alone does not necessarily parallel endogenous gene transcription, and that the reporter gene assay may detect interactions that are not functional in vivo.
Gozgit
Joseph M
JM
Department of Veterinary and Animal Sciences, University of Massachusetts Amherst, Amherst, MA 01003-9298, USA.
Nestor
Kathleen M
KM
Fasco
Michael J
MJ
Pentecost
Brian T
BT
Arcaro
Kathleen F
KF
D001667:Binding, Competitive; D001943:Breast Neoplasms / Q000378:metabolism / Q000473:pathology; D045744:Cell Line, Tumor; D015870:Gene Expression / Q000187:drug effects*; D017930:Genes, Reporter / Q000235:genetics*; D006801:Humans; D011084:Polycyclic Aromatic Hydrocarbons / Q000737:chemistry / Q000633:toxicity*; D018336:Receptors, Aryl Hydrocarbon / Q000235:genetics / Q000378:metabolism; D011960:Receptors, Estrogen / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D020218:Response Elements / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
D011084:Polycyclic Aromatic Hydrocarbons; D018336:Receptors, Aryl Hydrocarbon; D011960:Receptors, Estrogen; D011994:Recombinant Proteins