Éditeur(s) : HAL CCSD Endocrine Society
Résumé : International audience
Transcriptional regulation by the estrogen receptor-alpha(ER) has been investigated mainly in breast cancer cell lines, but estrogens such as 17 beta-estradiol (E2) exert numerous extrareproductive effects, particularly in the liver, where E2 exhibits both protective metabolic and deleterious thrombotic actions. To analyze the direct and early transcriptional effects of estrogens in the liver, we determined the E2-sensitive transcriptome and ER cistrome in mice after acute administration of E2 or placebo. These analyses revealed the early induction of genes involved in lipid metabolism, which fits with the crucial role of ER in the prevention of liver steatosis. Characterization of the chromatin state of ER binding sites (BSs) in mice expressing or not ER demonstrated that ER is not required per se for the establishment and/or maintenance of chromatin modifications at the majority of its BSs. This is presumably a consequence of a strong overlap between ER and hepatocyte nuclear factor 4 alpha BSs. In contrast, 40% of the BSs of the pioneer factor forkhead box protein a (Foxa2) were dependent upon ER expression, and ER expression also affected the distribution of nucleosomes harboring dimethylated lysine 4 of Histone H3 around Foxa2 BSs. We finally show that, in addition to a network of liver-specific transcription factors including CCAAT/enhancer-binding protein and hepatocyte nuclear factor 4 alpha, ER might be required for proper Foxa2 function in this tissue.
ISSN: 0888-8809

hal-01357623
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01357623
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01357623/document
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01357623/file/Changes%20in%20gene%20expression%201%20and%20Estrogen%20Receptor_accepted.pdf
DOI : 10.1210/me.2015-1311

Éditeur(s) : HAL CCSD Elsevier
Résumé : International audience
Xeno-estrogens, a class of endocrine disrupting chemicals (EDCs), can disturb estrogen receptor-dependent pathways involved in differentiation, proliferation or protection. Multiple methods have been developed to characterize the disturbances induced by EDCs in different cells or organs. In this study we have developed a new tool for the assessment of estrogenic compounds on differentiation. For this purpose we used the global model of NGF-induced neurite outgrowth of a pseudoneuronal PC12 cell line stably transfected with estrogen receptor alpha (PC12 ER). This new test evidences a new selectivity in which estradiol, genistein and 4-hydroxytamoxifen increased the NGF-induced neurite outgrowth of PC12 ER cells in a dose-dependent manner. In contrast, the strong estrogen agonist 17α-ethynylestradiol, the strong antagonist raloxifene and the agonist bisphenol A were unable to modify the neuritogenesis of PC12 ER cells. Therefore, the analysis of neuritogenesis in PC12 ER cells constitutes a complementary tool for the characterization of xeno-estrogen activity and also serves as a basis for further studies focusing on the mechanisms of EDCs in a neuronal context. Moreover, this test constitutes an alternative to animal testing.
ISSN: 0045-6535

hal-01063907
https://hal.archives-ouvertes.fr/hal-01063907
https://hal.archives-ouvertes.fr/hal-01063907/document
https://hal.archives-ouvertes.fr/hal-01063907/file/Differentiationsur PC12_cells_accepted.pdf
DOI : 10.1016/j.chemosphere.2014.03.101

Éditeur(s) : HAL CCSD Elsevier
Résumé : International audience
: Until recently, synonymous mutations (which do not change amino acids) have been much neglected. Some evidence suggests that this kind of mutations could affect mRNA secondary structure or stability, translation kinetics and protein structure. To explore deeper the role of synonymous mutations, we studied their consequence on the functional activity of the estrogen receptor alpha (ERα). The ERα is a ligand-inducible transcription factor that orchestrates pleiotropic cellular effects, at both genomic and non-genomic levels in response to estrogens. In this work we analyzed in transient transfection experiments, the activity of ERα carrying the synonymous mutation Ala87, a polymorphism involving about 5-10% of the population. In comparison to the wild type receptor, our results show that ERαA87 mutation reduces the transactivation efficiency of ERα on an ERE reporter gene while its expression level remains similar. This mutation enhances 4-OHT-induced transactivation of ERα on an AP1 reporter gene. Finally, the mutation affects the subcellular localization of ERα in a cell type specific manner. It enhances the cytoplasmic location of ERα without significant changes in non-genomic effects of E2. The functional alteration of the ERαA87 determined in this work highlights the relevance of synonymous mutations for biomedical and pharmacological points of view.
Journal of steroid biochemistry and molecular biology

inserm-01015532
http://www.hal.inserm.fr/inserm-01015532
DOI : 10.1016/j.jsbmb.2014.02.016
PUBMED : 24607813

Éditeur(s) : HAL CCSD Elsevier
Résumé : International audience
Archives of Cardiovascular Diseases Supplements
Strasbourg, France

hal-01118035
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01118035
DOI : 10.1016/S1878-6480(14)71283-2

Éditeur(s) : HAL CCSD American Society for Investigative Pathology
Résumé : International audience
Based on both experimental and clinical data, tamoxifen has been proposed to have cardiovascular benefits, although the mechanism(s) contributing to that protective effect are still poorly understood. In vitro experiments demonstrated that tamoxifen elicits its transcriptional effect through estrogen receptor (ER) α, but other targets can participate in its actions. However, although tamoxifen selectively activates the activating function (AF)-1 of ERα, we recently showed that this ERα subfunction is dispensable for the atheroprotective action of 17β-estradiol (E2), the main ligand of ERs. The goal of the present work is to determine to which extent ERα and its AF-1 mediate the vasculoprotective action of tamoxifen. Our data confirm that tamoxifen exerts an atheroprotective action on low density lipoprotein receptor (LDL-r(-/-)) female mice, but, in contrast to E2, it fails to accelerate reendothelialization after carotid electric injury. Tamoxifen and E2 elicit differences in gene expression profiles in the mouse aorta. Finally, the atheroprotective action of tamoxifen is abrogated in ERα(-/-)LDL-r(-/-) mice and in LDL-r(-/-) mice selectively deficient in ERαAF-1 (ERαAF-1(0/0)LDL-r(-/-)). Our results demonstrate, for the first time to our knowledge, that tamoxifen mediates its actions in vivo through the selective activation of ERαAF-1, which is sufficient to prevent atheroma, but not to accelerate endothelial healing.
ISSN: 0002-9440

hal-00874317
https://hal.archives-ouvertes.fr/hal-00874317
DOI : 10.1016/j.ajpath.2013.03.010
PUBMED : 23669343

Éditeur(s) : HAL CCSD
Résumé : International audience
Estrogen receptor alpha (ERα) has been recognized now for several decades as playing a key role in reproduction and exerting functions in numerous nonreproductive tissues. In this review, we attempt to summarize the in vitro studies that are the basis of our current understanding of the mechanisms of action of ERα as a nuclear receptor and the key roles played by its two activation functions (AFs) in its transcriptional activities. We then depict the consequences of the selective inactivation of these AFs in mouse models, focusing on the prominent roles played by ERα in the reproductive tract and in the vascular system. Evidence has accumulated over the two last decades that ERα is also associated with the plasma membrane and activates non-nuclear signaling from this site. These rapid/nongenomic/membrane-initiated steroid signals (MISS) have been characterized in a variety of cell lines, and in particular in endothelial cells. The development of selective pharmacological tools that specifically activate MISS and the generation of mice expressing an ERα protein impeded for membrane localization have begun to unravel the physiological role of MISS in vivo. Finally, we discuss novel perspectives for the design of tissue-selective ER modulators based on the integration of the physiological and pathophysiological roles of MISS actions of estrogens.
Physiological Reviews

hal-01533249
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01533249
DOI : 10.1152/physrev.00024.2016
PUBMED : 28539435

Éditeur(s) : HAL CCSD Elsevier
Résumé : International audience
Estrogen receptor alpha (ERα) is generally considered to be a good prognostic marker because almost 70% of ERα-positive tumors respond to anti-hormone therapies. Unfortunately, during cancer progression, mammary tumors can escape from estrogen control, resulting in resistance to treatment. In this study, we demonstrate that activation of the actin/megakaryoblastic leukemia 1 (MKL1) signaling pathway promotes the hormonal escape of estrogen-sensitive breast cancer cell lines. The actin/MKL1 signaling pathway is silenced in differentiated ERα-positive breast cancer MCF-7 and T47D cell lines and active in ERα-negative HMT-3522 T4-2 and MDA-MB-231 breast cancer cells, which have undergone epithelial-mesenchymal transition. We showed that MKL1 activation in MCF-7 cells, either by modulating actin dynamics or using MKL1 mutants, down-regulates ERα expression and abolishes E2-dependent cell growth. Interestingly, the constitutively active form of MKL1 represses PR and HER2 expression in these cells and increases the expression of HB-EGF, TGFβ, and amphiregulin growth factors in an E2-independent manner. The resulting expression profile (ER-, PR-, HER2-) typically corresponds to the triple-negative breast cancer expression profile.
ISSN: 0303-7207

hal-01061363
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01061363
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01061363/document
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01061363/file/Activationsur the_MKL1actin_Kerdivel_et_al._text_Figures.pdf
DOI : 10.1016/j.mce.2014.03.009

Éditeur(s) : HAL CCSD IOS Press
Résumé : International audience
Cancer is generally conceived as a dedifferentiation process in which quiescent post-mitotic differentiated cells acquire stem-like properties and the capacity to proliferate. This view holds for the initial stages of carcinogenesis but is more questionable for advanced stages when the cells can transdifferentiate into the contractile phenotype associated to migration and metastasis. Singularly from this perspective, the hallmark of the most aggressive cancers would correspond to a genuine differentiation status, even if it is different from the original one. This seeming paradox could help reconciling discrepancies in the literature about the pro-or anti-tumoral functions of candidate molecules involved in cancer and whose actual effects depend on the tumoral grade. These ambiguities which are likely to concern a myriad of molecules and pathways, are illustrated here with the selected examples of chromatin epigenetics and myocardin-related transcription factors, using the human MCF10A and MCF7 breast cancer cells. Self-renewing stem like cells are characterized by a loose chromatin with low levels of the H3K9 trimetylation, but high levels of this mark can also appear in cancer cells acquiring a contractile-type differentiation state associated to metastasis. Similarly, the myocardin-related transcription factor MRTF-A is involved in metastasis and epithelial-mesenchymal transition, whereas this factor is naturally enriched in the quiescent cells which are precisely the most resistant to cancer: cardiomyocytes. These seeming paradoxes reflect the bistable epigenetic landscape of cancer in which dedifferentiated self-renewing and differentiated migrating states are incompatible at the single cell level, though coexisting at the population level. © 2016-IOS Press and the authors. All rights reserved.
ISSN: 0888-6008

hal-01367146
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01367146
DOI : 10.3233/BD-150210
PUBMED : 27177343

Éditeur(s) : HAL CCSD Springer Verlag
Résumé : International audience
A hallmark of living systems is the management and the storage of information through genetic and epigenetic mechanisms. Although the notion of epigenetics was originally given to any regulation beyond DNA sequence, it has often been restricted to chromatin modifications, supposed to behave as cis-markers, specifying the sets of genes to be expressed or repressed. This definition does not take into account the initial view of epigenetics, based on nonlinear interaction networks whose "attractors" can remain stable without need for any chromatin mark. In addition, most chromatin modifications are the steady state resultants of highly dynamic modification and de-modification activities and, as such, seem poorly appropriate to work as long-term memory keepers. Instead, the basic support of epigenetic memory could remain the attractors, to which chromatin modifications belong as do many other components. The influence of chromatin modifications in memory is highly questionable when envisioned as static structural marks, but can be recovered under the dynamic circuitry perspective, thanks to their self-templating properties. Beside their standard repressive or permissive functions, chromatin modifications can also influence transcription in multiple ways such as: (1) by randomizing or inversely stabilizing gene expression, (2) by mediating cooperativity between pioneer and secondary transcription factors, and (3) in the hysteresis and the ultrasensitivity of gene expression switches, allowing the cells to take unambiguous transcriptional decisions.
ISSN: 1420-682X

hal-01063617
https://hal.archives-ouvertes.fr/hal-01063617
DOI : 10.1007/s00018-012-0923-7
PUBMED : 22331281

Éditeur(s) : HAL CCSD
Résumé : International audience
In addition to soluble factors, mechanical constraints and extracellular matrix stiffness are important regulators of cell fate that are mediated by cytoskeletal modifications. The EMT (epithelial-mesenchymal transition) that occurs during normal development and malignant progression is a typical example of the phenotypic switch associated with profound actin remodelling and changes in gene expression. For instance, actin dynamics control motile cell functions in EMT, in part, through regulating the subcellular localization of the myocardin-related transcription factor MKL1 (megakaryoblastic leukaemia translocation 1), a co-activator of SRF (serum-responsive factor). In the present paper, we show that MKL1 participates also to the control of the cellular switch between growth and quiescence. Experimental disconnection between MKL1 and G-actin (globular actin), by using an MKL1 mutant or enhancing the F (filamentous)-/G-actin ratio, generates a widely open chromatin state and a global increase in biosynthetic activity, classically associated with cell growth. Conversely, G-actin accumulation favours nuclear condensation and cell quiescence. These large-scale chromatin changes rely upon extensive histone modifications, exemplified by that of H3K9 (H3 Lys9) shifting from trimethylation, a heterochromatin mark, to acetylation, a mark of euchromatin. The present study provides the first evidence for a global reversible hetero/euchromatinization phenomenon triggered by the actin/MKL1 signalling pathway.
The Biochemical Journal

hal-01063933
https://hal.archives-ouvertes.fr/hal-01063933
DOI : 10.1042/BJ20131240
PUBMED : 24762104

Éditeur(s) : HAL CCSD BioMed Central
Résumé : International audience
BackgroundTo date, all studies conducted on breast cancer diagnosis have focused on the expression of the full-length 66-kDa estrogen receptor alpha (ERα66). However, much less attention has been paid to a shorter 46-kDa isoform (ERα46), devoid of the N-terminal region containing the transactivation function AF-1. Here, we investigated the expression levels of ERα46 in breast tumors in relation to tumor grade and size, and examined the mechanism of its generation and its specificities of coregulatory binding and its functional activities.MethodsUsing approaches combining immunohistochemistry, Western blotting, and proteomics, antibodies allowing ERα46 detection were identified and the expression levels of ERα46 were quantified in 116 ERα-positive human breast tumors. ERα46 expression upon cellular stress was studied, and coregulator bindings, transcriptional, and proliferative response were determined to both ERα isoforms.ResultsERα46 was expressed in over 70% of breast tumors at variable levels which sometimes were more abundant than ERα66, especially in differentiated, lower-grade, and smaller-sized tumors. We also found that ERα46 can be generated via internal ribosome entry site-mediated translation in the context of endoplasmic reticulum stress. The binding affinities of both unliganded and fully-activated receptors towards co-regulator peptides revealed that the respective potencies of ERα46 and ERα66 differ significantly, contributing to the differential transcriptional activity of target genes to 17β estradiol (E2). Finally, increasing amounts of ERα46 decrease the proliferation rate of MCF7 tumor cells in response to E2.ConclusionsWe found that, besides the full-length ERα66, the overlooked ERα46 isoform is also expressed in a majority of breast tumors. This finding highlights the importance of the choice of antibodies used for the diagnosis of breast cancer, which are able or not to detect the ERα46 isoform. In addition, since the function of both ERα isoforms differs, this work underlines the need to develop new technologies in order to discriminate ERα66 and ERα46 expression in breast cancer diagnosis which could have potential clinical relevance.
ISSN: 1465-5411

inserm-01412032
http://www.hal.inserm.fr/inserm-01412032
http://www.hal.inserm.fr/inserm-01412032/document
http://www.hal.inserm.fr/inserm-01412032/file/13058_2016_Article_780.pdf
DOI : 10.1186/s13058-016-0780-7

Éditeur(s) : HAL CCSD Public Library of Science
Résumé : International audience
Many studies have reported proliferative, differentiating or protective effects of estradiol, notably through estrogen receptor alpha (ERα). On the contrary, the ligand-independent action of ERα is currently poorly documented notably in cell protection. The stable transfection of wild type, substituted or truncated form of ERα in PC12 cells (ERα negative cell line) lead the specific study of its ligand-independent action. Hence, we demonstrate here that, in the absence of E2, the expression of ERα prevents cells from apoptosis induced by serum deprivation. This protection is not due to an ERE-mediated transcription and does not require either AF-1 or AF-2 transactivation functions. It is afforded to the Y537 residue of ERα and activation of c-Src/Stat3 signaling pathway.
ISSN: 1932-6203

inserm-00865332
http://www.hal.inserm.fr/inserm-00865332
http://www.hal.inserm.fr/inserm-00865332/document
http://www.hal.inserm.fr/inserm-00865332/file/PMID_23825704.pdf
DOI : 10.1371/journal.pone.0069081

Éditeur(s) : HAL CCSD Elsevier
Résumé : International audience
Seventy percent of breast tumors express the estrogen receptor (ER), which is generally considered to predict a better outcome relative to ER-negative tumors, as they often respond to antiestrogen therapies. During cancer progression, mammary tumors can escape from estrogen control, resulting in the acquisition of invasive properties and resistance to treatment. ER expression is a dynamic phenomenon and is finely regulated at numerous levels, including the gene, mRNA, and protein levels. As a consequence, many molecular mechanisms have been implicated in modulating ER activity and estrogen signaling in mammary cancer. In fact, one-third of ER-positive breast cancer cells do not respond to first-line endocrine therapies, and a large subset of relapsing tumors retain ER expression. Increased knowledge of these mechanisms has led to the development of better prognostic methods and targeted therapies for patients; however, additional research is still needed to improve patient survival. In this chapter, we focus on the signaling pathways leading to changes in or loss of ER activity in breast cancer progression.
ISSN: 0083-6729

hal-00874313
https://hal.archives-ouvertes.fr/hal-00874313
DOI : 10.1016/B978-0-12-416673-8.00004-6
PUBMED : 23810005

Éditeur(s) : HAL CCSD BioMed Central
Résumé : International audience
BACKGROUND: The orphan receptors COUP-TF (chicken ovalbumin upstream promoter transcription factor) I and II are members of the nuclear receptor superfamily that play distinct and critical roles in vertebrate organogenesis. The involvement of COUP-TFs in cancer development has recently been suggested by several studies but remains poorly understood. METHODS: MCF-7 breast cancer cells overexpressing COUP-TFI and human breast tumors were used to investigate the role of COUP-TFI in the regulation of CXCL12/CXCR4 signaling axis in relation to cell growth and migration. We used Immunofluorescence, western-blot, RT-PCR, Formaldehyde-assisted Isolation of Regulatory Elements (FAIRE) assays, as well as cell proliferation and migration assays. RESULTS: Previously, we showed that COUP-TFI expression is enhanced in breast cancer compared to normal tissue. Here, we report that the CXCL12/CXCR4 signaling pathway, a crucial pathway in cell growth and migration, is an endogenous target of COUP-TFI in breast cancer cells. The overexpression of COUP-TFI in MCF-7 cells inhibits the expression of the chemokine CXCL12 and markedly enhances the expression of its receptor, CXCR4. Our results demonstrate that the modification of CXCL12/CXCR4 expression by COUP-TFI is mediated by the activation of epithelial growth factor (EGF) and the EGF receptor. Furthermore, we provide evidence that these effects of COUP-TFI increase the growth and motility of MCF-7 cells in response to CXCL12. Cell migration toward a CXCL12 gradient was inhibited by AMD3100, a specific antagonist of CXCR4, or in the presence of excess CXCL12 in the cell culture medium. The expression profiles of CXCR4, CXCR7, CXCL12, and COUP-TFI mRNA in 82 breast tumors and control non-tumor samples were measured using real-time PCR. CXCR4 expression was found to be significantly increased in the tumors and correlated with the tumor grade, whereas the expression of CXCL12 was significantly decreased in the tumors compared with the healthy samples. Significantly higher COUP-TFI mRNA expression was also detected in grade 1 tumors. CONCLUSIONS: Together, our mechanistic in vitro assays and in vivo results suggest that a reduction in chemokine CXCL12 expression, with an enhancement of CXCR4 expression, provoked by COUP-TFI, could be associated with an increase in the invasive potential of breast cancer cells.
ISSN: 1471-2407

hal-01024753
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01024753
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01024753/document
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01024753/file/1471-2407-14-407.pdf
DOI : 10.1186/1471-2407-14-407

Éditeur(s) : HAL CCSD Springer New York
Résumé : International audience
Estrogen receptor α (ERα) exerts regulatory actions through genomic mechanisms. In the classical pathway, ligand-activated ERα binds directly to DNA through estrogen response elements (ERE) located in the promoter of target genes. ERα can also exert indirect regulation of transcription via protein-protein interaction with other transcription factors such as AP-1. Several ERα synonymous polymorphisms have been identified and efforts to understand their implications have been made. Nevertheless effects of synonymous polymorphisms are still neglected. This chapter focuses on the experimental procedure employed in order to characterize the transcriptional activity of a synonymous polymorphism of the ERα (rs746432) called Alanine 87 (Ala87). Activity of both WT and Ala87 ERα isoforms on transcriptional pathways can be analyzed in transiently transfected cells using different reporter constructs. ERα efficiency on the classical genomic pathway can be analyzed by determining its transactivation activity on an ERE-driven thymidine kinase (TK) promoter controlling the expression of the luciferase reporter gene. Transcriptional activity through the indirect genomic pathway can be analyzed by employing an AP-1 DNA response element-driven promoter also controlling the expression of luciferase reporter gene
Estrogen Receptors

ISBN : 978-1-4939-3126-2
hal-01237073
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01237073
DOI : 10.1007/978-1-4939-3127-9_22

Éditeur(s) : HAL CCSD Wiley Open Access
Résumé : International audience
Estetrol (E4) is a natural estrogen with a long half-life produced only by the human fetal liver during pregnancy. The crystal structures of the estrogen receptor α (ERα) ligand-binding domain bound to 17β-estradiol (E2) and E4 are very similar, as well as their capacity to activate the two activation functions AF-1 and AF-2 and to recruit the coactivator SRC3. In vivo administration of high doses of E4 stimulated uterine gene expression, epithelial proliferation, and prevented atheroma, three recognized nuclear ERα actions. However, E4 failed to promote endothelial NO synthase activation and acceleration of endothelial healing, two processes clearly dependent on membrane-initiated steroid signaling (MISS). Furthermore, E4 antagonized E2 MISS-dependent effects in endothelium but also in MCF-7 breast cancer cell line. This profile of ERα activation by E4, uncoupling nuclear and membrane activation, characterizes E4 as a selective ER modulator which could have medical applications that should now be considered further.
ISSN: 1757-4676

hal-01118028
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01118028
PUBMED : 25214462
PUBMEDCENTRAL : PMC4287935
DOI : 10.15252/emmm.201404112