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Effects of Benzophenone-3 and Propylparaben on Estrogen Receptor-Dependent R-Loops and DNA Damage in Breast Epithelial Cells and Mice.

ABSTRACT: BACKGROUND:Endocrine-disrupting chemicals have been shown to have broad effects on development, but their mutagenic actions that can lead to cancer have been less clearly demonstrated. Physiological levels of estrogen have been shown to stimulate DNA damage in breast epithelial cells through mechanisms mediated by estrogen-receptor alpha (ER?). Benzophenone-3 (BP-3) and propylparaben (PP) are xenoestrogens found in the urine of >96% of U.S. POPULATION: OBJECTIVES:We investigated the effect of BP-3 and PP on estrogen receptor-dependent transactivation and DNA damage at concentrations relevant to exposures in humans. METHODS:In human breast epithelial cells, DNA damage following treatment with 17?-estradiol (E2), BP-3, and PP was determined by immunostaining with antibodies against ?-H2AX and 53BP1. Estrogenic responses were determined using luciferase reporter assays and gene expression. Formation of R-loops was determined with DNA: RNA hybrid-specific S9.6 antibody. Short-term exposure to the chemicals was also studied in ovariectomized mice. Immunostaining of mouse mammary epithelium was performed to quantify R-loops and DNA damage in vivo. RESULTS:Concentrations of 1?M and 5?M BP-3 or PP increased DNA damage similar to that of E2 treatment in a ER?-dependent manner. However, BP-3 and PP had limited transactivation of target genes at 1?M and 5?M concentrations. BP-3 and PP exposure caused R-loop formation in a normal human breast epithelial cell line when ER? was introduced. R-loops and DNA damage were also detected in mammary epithelial cells of mice treated with BP-3 and PP. CONCLUSIONS:Acute exposure to xenoestrogens (PP and BP-3) in mice induce DNA damage mediated by formation of ER?-dependent R-loops at concentrations 10-fold lower than those required for transactivation. Exposure to these xenoestrogens may cause deleterious estrogenic responses, such as DNA damage, in susceptible individuals. https://doi.org/10.1289/EHP5221.

SUBMITTER: Majhi PD

PROVIDER: S-EPMC7015622 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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Effects of Benzophenone-3 and Propylparaben on Estrogen Receptor-Dependent R-Loops and DNA Damage in Breast Epithelial Cells and Mice.

Majhi Prabin Dhangada PD Sharma Aman A Roberts Amy L AL Daniele Elizabeth E Majewski Aliza R AR Chuong Lynn M LM Black Amye L AL Vandenberg Laura N LN Schneider Sallie S SS Dunphy Karen A KA Jerry D Joseph DJ

Environmental health perspectives 20200115 1

<h4>Background</h4>Endocrine-disrupting chemicals have been shown to have broad effects on development, but their mutagenic actions that can lead to cancer have been less clearly demonstrated. Physiological levels of estrogen have been shown to stimulate DNA damage in breast epithelial cells through mechanisms mediated by estrogen-receptor alpha (ERα). Benzophenone-3 (BP-3) and propylparaben (PP) are xenoestrogens found in the urine of >96% of U.S.<h4>Population</h4><h4>Objectives</h4>We investi ...[more]

PMID: 31939680

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Project description:BACKGROUND:Breast cancer (BC) is highly heterogeneous with ~?60-70% of estrogen receptor positive BC patient's response to anti-hormone therapy. Estrogen receptors (ERs) play an important role in breast cancer progression and treatment. Estrogen related receptors (ERRs) are a group of nuclear receptors which belong to orphan nuclear receptors, which have sequence homology with ERs and share target genes. Here, we investigated the possible role and clinicopathological importance of ERR? in breast cancer. METHODS:Estrogen related receptor ? (ERR?) expression was examined using tissue microarray slides (TMA) of Breast Carcinoma patients with adjacent normal by immunohistochemistry and in breast cancer cell lines. In order to investigate whether ERR? is a direct target of ER?, we investigated the expression of ERR? in short hairpin ribonucleic acid knockdown of ER? breast cancer cells by western blot, qRT-PCR and RT-PCR. We further confirmed the binding of ER? by electrophoretic mobility shift assay (EMSA), chromatin immunoprecipitation (ChIP), Re-ChIP and luciferase assays. Fluorescence-activated cell sorting analysis (FACS) was performed to elucidate the role of ERR? in cell cycle regulation. A Kaplan-Meier Survival analysis of GEO dataset was performed to correlate the expression of ERR? with survival in breast cancer patients. RESULTS:Tissue microarray (TMA) analysis showed that ERR? is significantly down-regulated in breast carcinoma tissue samples compared to adjacent normal. ER?+?ve breast tumors and cell lines showed a significant expression of ERR? compared to ER-ve tumors and cell lines. Estrogen treatment significantly induced the expression of ERR? and it was ER? dependent. Mechanistic analyses indicate that ER? directly targets ERR? through estrogen response element and ERR? also mediates cell cycle regulation through p18, p21cip and cyclin D1 in breast cancer cells. Our results also showed the up-regulation of ERR? promoter activity in ectopically co-expressed ER? and ERR? breast cancer cell lines. Fluorescence-activated cell sorting analysis (FACS) showed increased G0/G1 phase cell population in ERR? overexpressed MCF7 cells. Furthermore, ERR? expression was inversely correlated with overall survival in breast cancer. Collectively our results suggest cell cycle and tumor suppressor role of ERR? in breast cancer cells which provide a potential avenue to target ERR? signaling pathway in breast cancer. CONCLUSION:Our results indicate that ERR? is a negative regulator of cell cycle and a possible tumor suppressor in breast cancer. ERR? could be therapeutic target for the treatment of breast cancer.

Dataset Information

Effects of Benzophenone-3 and Propylparaben on Estrogen Receptor-Dependent R-Loops and DNA Damage in Breast Epithelial Cells and Mice.

Publications

Effects of Benzophenone-3 and Propylparaben on Estrogen Receptor-Dependent R-Loops and DNA Damage in Breast Epithelial Cells and Mice.

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