AHR and GPER mediate the stimulatory effects induced by 3-methylcholanthrene in breast cancer cells and cancer-associated fibroblasts (CAFs).

Abstract

BACKGROUND:

The chemical carcinogen 3-methylcholanthrene (3MC) binds to the aryl hydrocarbon receptor (AHR) that regulates the expression of cytochrome P450 (CYP) enzymes as CYP1B1, which is involved in the oncogenic activation of environmental pollutants as well as in the estrogen biosynthesis and metabolism. 3MC was shown to induce estrogenic responses binding to the estrogen receptor (ER) α and stimulating a functional interaction between AHR and ERα. Recently, the G protein estrogen receptor (GPER) has been reported to mediate certain biological responses induced by endogenous estrogens and environmental compounds eliciting an estrogen-like activity.

METHODS:

Molecular dynamics and docking simulations were performed to evaluate the potential of 3MC to interact with GPER. SkBr3 breast cancer cells and cancer-associated fibroblasts (CAFs) derived from breast tumor patients were used as model system. Real-time PCR and western blotting analysis were performed in order to evaluate the activation of transduction mediators as well as the mRNA and protein levels of CYP1B1 and cyclin D1. Co-immunoprecipitation studies were performed in order to explore the potential of 3MC to trigger the association of GPER with AHR and EGFR. Luciferase assays were carried out to determine the activity of CYP1B1 promoter deletion constructs upon 3MC exposure, while the nuclear shuttle of AHR induced by 3MC was assessed through confocal microscopy. Cell proliferation stimulated by 3MC was determined as biological counterpart of the aforementioned experimental assays. The statistical analysis was performed by ANOVA.

RESULTS:

We first ascertained by docking simulations the ability of 3MC to interact with GPER. Thereafter, we established that 3MC activates the EGFR/ERK/c-Fos transduction signaling through both AHR and GPER in SkBr3 cells and CAFs. Then, we found that these receptors are involved in the up-regulation of CYP1B1 and cyclin D1 as well as in the stimulation of growth responses induced by 3MC.

CONCLUSIONS:

In the present study we have provided novel insights regarding the molecular mechanisms by which 3MC may trigger a physical and functional interaction between AHR and GPER, leading to the stimulation of both SkBr3 breast cancer cells and CAFs. Altogether, our results indicate that 3MC may engage both GPER and AHR transduction pathways toward breast cancer progression.

• Authors: Francesca Cirillo, Rosamaria Lappano, Leonardo Bruno, Bruno Rizzuti, Fedora Grande, Rita Guzzi, Sara Briguori, Anna Maria Miglietta, Miki Nakajima, Maria Teresa Di Martino, Marcello Maggiolini
• Journal: Journal of experimental & clinical cancer research : CR (J Exp Clin Cancer Res) Vol. 38 Issue 1 Pg. 335 (Aug 01 2019) ISSN: 1756-9966 [Electronic] England
• PMID: 31370872 (Publication Type: Journal Article)

Chemical References

• AHR protein, human
• Basic Helix-Loop-Helix Transcription Factors
• GPER1 protein, human
• Receptors, Aryl Hydrocarbon
• Receptors, Estrogen
• Receptors, G-Protein-Coupled
• Methylcholanthrene
• CYP1B1 protein, human
• Cytochrome P-450 CYP1B1
• EGFR protein, human
• ErbB Receptors
• Extracellular Signal-Regulated MAP Kinases

Topics

• Basic Helix-Loop-Helix Transcription Factors (chemistry, metabolism)
• Breast Neoplasms (genetics, metabolism, pathology)
• Cancer-Associated Fibroblasts (drug effects, metabolism, pathology)
• Cell Line, Tumor
• Cell Proliferation (drug effects)
• Cytochrome P-450 CYP1B1 (genetics, metabolism)
• ErbB Receptors (metabolism)
• Extracellular Signal-Regulated MAP Kinases (metabolism)
• Female
• Gene Expression Regulation, Neoplastic
• Humans
• Methylcholanthrene (chemistry, pharmacology)
• Molecular Conformation
• Molecular Docking Simulation
• Molecular Dynamics Simulation
• Protein Binding
• Protein Transport
• Receptors, Aryl Hydrocarbon (chemistry, metabolism)
• Receptors, Estrogen (chemistry, metabolism)
• Receptors, G-Protein-Coupled (chemistry, metabolism)
• Signal Transduction (drug effects)
• Structure-Activity Relationship