Protein Kinase D1 (PKD1) Is a New Functional Non-Genomic Target of Bisphenol A in Breast Cancer Cells Messaouda Merzoug-Larabi, Ilige Youssef, Ai Thu Bui, Christine Legay, Sophia Loiodice, Sophie Lognon, Sylvie Babajko, Jean-Marc Ricort To cite this version: Messaouda Merzoug-Larabi, Ilige Youssef, Ai Thu Bui, Christine Legay, Sophia Loiodice, et al.. Pro- tein Kinase D1 (PKD1) Is a New Functional Non-Genomic Target of Bisphenol A in Breast Cancer Cells. Frontiers in Pharmacology, Frontiers, 2020, 10, pp.1683. 10.3389/fphar.2019.01683. inserm- 03033637 HAL Id: inserm-03033637 https://www.hal.inserm.fr/inserm-03033637 Submitted on 1 Dec 2020 HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. 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ORIGINAL RESEARCH published: 31 January 2020 doi: 10.3389/fphar.2019.01683 Protein Kinase D1 (PKD1) Is a New Functional Non-Genomic Target of Bisphenol A in Breast Cancer Cells Messaouda Merzoug-Larabi 1,2, Ilige Youssef 1,2, Ai Thu Bui 3, Christine Legay 1,2, † † Sophia Loiodice 3, Sophie Lognon 2, Sylvie Babajko 3 and Jean-Marc Ricort 1,2* 1 Centre National de la Recherche Scientifique, CNRS UMR_8113, Laboratoire de Biologie et Pharmacologie Appliquée, Cachan, France, 2 École Normale Supérieure Paris-Saclay, Université Paris-Saclay, Cachan, France, 3 Centre de Recherche des Cordeliers, INSERM, Sorbonne Université, Université de Paris, Laboratoire de Physiopathologie Orale Moléculaire, Paris, France Exposure to bisphenol A (BPA), one of the most widespread endocrine disruptors present in our environment, has been associated with the recent increased prevalence and Edited by: severity of several diseases such as diabetes, obesity, autism, reproductive and George D. Loizou, neurological defects, oral diseases, and cancers such as breast tumors. BPA is Health and Safety Executive, suspected to act through genomic and non-genomic pathways. However, its precise United Kingdom molecular mechanisms are still largely unknown. Our goal was to identify and characterize Reviewed by: Detlef Woelfle, a new molecular target of BPA in breast cancer cells in order to better understand how this European Food Safety Authority compound may affect breast tumor growth and development. By using in vitro (MCF-7, (EFSA), Italy Heike Wulff, T47D, Hs578t, and MDA-MB231 cell lines) and in vivo models, we demonstrated that University of California, PKD1 is a functional non-genomic target of BPA. PKD1 specifically mediates BPA- Davis, United States induced cell proliferation, clonogenicity, and anchorage-independent growth of breast *Correspondence: tumor cells. Additionally, low-doses of BPA (≤10−8 M) induced the phosphorylation of Jean-Marc Ricort [email protected] PKD1, a key signature of its activation state. Moreover, PKD1 overexpression increased †These authors have contributed the growth of BPA-exposed breast tumor xenografts in vivo in athymic female Swiss nude equally to this work (Foxn1nu/nu) mice. These findings further our understanding of the molecular mechanisms of BPA. By defining PKD1 as a functional target of BPA in breast cancer cell proliferation Specialty section: This article was submitted to and tumor development, they provide new insights into the pathogenesis related to the Predictive Toxicology, exposure to BPA and other endocrine disruptors acting similarly. a section of the journal Frontiers in Pharmacology Keywords: endocrine disruptor, bisphenol A, protein kinase D1 (PKD1), breast cancer, pro-tumorigenic factor, non- genomic target Received: 08 February 2019 Accepted: 24 December 2019 Published: 31 January 2020 Citation: INTRODUCTION Merzoug-Larabi M, Youssef I, Bui AT, Legay C, Loiodice S, Lognon S, According to the World Health Organization’sdefinition (2002), endocrine disruptors are chemical Babajko S and Ricort J-M (2020) compounds that can interfere with the endocrine system and cause deleterious health effects to Protein Kinase D1 (PKD1) Is a New organisms or even their descendants. Among the most common endocrine disruptors, bisphenol A Functional Non-Genomic Target of fl Bisphenol A in Breast Cancer Cells. (BPA), used in epoxy resins and polycarbonate plastics, has been detected in biological uids of Front. Pharmacol. 10:1683. most of the population worldwide, essentially due to oral contamination by ingestion of BPA- doi: 10.3389/fphar.2019.01683 containing food and drinks (Pirard et al., 2012; Galloway et al., 2018). Epidemiological and Frontiers in Pharmacology | www.frontiersin.org 1 January 2020 | Volume 10 | Article 1683 Merzoug-Larabi et al. PKD1: A Bisphenol a Target experimental studies show that human BPA serum apoptosis, invasion, and motility (reviewed in (Sundram et al., − concentrations generally vary from 0.2 to 1.6 ng/mL 1 (0.88 to 2011) and plays a crucial role in cancer (reviewed in Youssef and 7 nM), but may reach higher values in workers who manipulate Ricort, 2019). We previously demonstrated that PKD1 high amounts of BPA (thermal paper or plastics industries) overexpression potentiates in vivo tumor growth of the MCF-7 (Hines et al., 2017). Many adverse health effects, such as adenocarcinoma-derived cell line, and regulates cell growth hyperactivity, obesity, fertility problems, enamel defects, and (Karam et al., 2012; Karam et al., 2014). Moreover, we cardiac arrhythmia have been associated with exposure to identified PKD1 as a poor prognostic factor in the whole BPA, especially during the perinatal period of life (Jedeon breast cancer population and in the triple-negative breast et al., 2013) (reviewed in Giulivo et al., 2016). Moreover, cancer (TNBC) subtype specifically (Spasojevic et al., 2018). exposure to BPA has been associated with the recent increased Therefore, due to its crucial role in breast tumor growth and incidence of prostate and breast cancer (reviewed in Seachrist development, we asked in this study whether PKD1 may be a et al., 2016). molecular target of BPA. Gestational or perinatal exposure of rodents and monkeys to BPA alters mammary gland development, increasing the risk of later onset of breast tumors (Munoz-de-Toro et al., 2005; Tharp MATERIALS AND METHODS et al., 2012; Mandrup et al., 2016). In women, BPA serum concentrations positively correlate with higher breast density, a Antibodies and Materials well-known risk factor for the subsequent development of breast Anti-PKD1 (1/1,000), anti-phospho-S910-PKD1 (1/1,000), anti- tumors (Sprague et al., 2013). Exposure to BPA also increases the phospho-S738/742-PKD1 (1/1,000), and anti-ERa (1/2,000) number of pre-cancerous mammary lesions and breast were purchased from Cell Signaling (Danvers, MA); anti-actin carcinomas and promotes metastasis (Murray et al., 2007; (1/1,000) and anti-GAPDH (1/2,000) from Santa Cruz Jenkins et al., 2011). Biotechnology (Santa Cruz, CA). The horseradish peroxidase- BPA interacts with intracellular estrogen receptors (ERa, conjugated secondary antibodies used were goat anti-rabbit IgG ERb) and ERRg (Takayanagi et al., 2006; Delfosse et al., 2012; (1/2,000; Dako, Glostrup, Denmark) and goat anti-mouse IgG Liu et al., 2012). It also modulates the activity of other (1/5,000; Rockland, Gilbertsville, PA). PRKD1-targeting (#5587) intracellular receptors, such as AR, PR, PPARg, RXRs, PXR, and control siRNAs were purchased from GE Healthcare- and TR enhancing cell proliferation and migration (reviewed in Dharmacon (Velizy-Villacoublay, France), Gö6976 and Acconcia et al., 2015). However, it acts differently than estrogens Gö6983 from Calbiochem (Darmstadt, Germany), MTT from on mammary gland (Speroni et al., 2017). In fact, aside from Sigma-Aldrich (St. Louis, MO) and BPA (purity 97%+) from these genomic processes, BPA also acts via non-genomic and Alfa Aesar (Haverhill, MA). ER-independent mechanisms through the regulation of intracellular signaling pathways. In breast cancer cells, BPA Tumorigenicity Assay in Athymic has been shown to activate ERK (Dong et al., 2011; Song et al., Nude Mice 2015), EGFR (Sauer et al., 2017), FAK, and Src (Castillo et al., Thirty 8-week old athymic female Swiss nude (Foxn1nu/nu) mice 2016), bind to small GTP binding proteins (Schopel et al., 2016), were purchased from Janvier Labs (Le Genest-Saint-Isle, France) modulate the phosphatidylinositol 3-kinase (PI3-K)/Akt and bred in our animal house for the tumorigenicity assay. All signaling pathway (Goodson et al., 2011), and down-regulate animals were fed ad libitum and maintained in accordance with PTEN expression (Wang et al., 2014). These signaling pathways the guidelines for the care and use of laboratory animals of the may be activated through binding of BPA to membrane French Ministry of Agriculture (A-75-06-12). All animals were receptors, such as GPR30 (Thomas and Dong, 2006; Dong treated humanely and with regard for alleviation of suffering. et al., 2011) or through metalloprotease-mediated shedding of Cages and bottles made of polypropylene were used to avoid any EGFR ligands, leading to EGFR activation (Sauer et al., 2017; BPA contamination. Mice were provided a phytoestrogens and Urriola-Munoz et al., 2017). Nowadays,