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Type: Thesis
Title: Steroid receptor crosstalk in breast cancer cells.
Author: Swinstead, Erin Elizabeth
Issue Date: 2014
School/Discipline: School of Medicine
Abstract: Breast cancer is the leading cause of cancer related death in women, and approximately 1 in 11 women will develop breast cancer before the age of 75. In 2003, breast cancer was responsible for 16% of cancer related deaths in Australian women. This demonstrates that throughout the life span of the female, this organ has a high risk of developing cancer. The growth and survival of normal breast epithelial cells and breast cancer cells is promoted by estrogens and progesterone and both estrogen receptor (ER) and progesterone receptor (PR) have been shown to play prominent roles in breast cancer progression. It has also been demonstrated that co-treatment of breast cancer cells with corticosteroids and 17β-estadiol (E2) can have opposing effects on the proliferation of breast cancer cells compared with the single treatment. In addition, glucocorticoid receptor (GR) levels have been shown to have clinical implications for breast cancer cell survival. This suggests a possible role for activated GR in breast cancer development. Forkhead box protein 1 (FoxA1), a member of the forkhead class of DNA-binding proteins, has also been shown to be an important factor in breast cancer development. FoxA1 has been shown to dictate ER binding in breast cancer cells and has been deemed responsible for the rapid reprogramming of ER signalling seen in breast cancers with poor outcomes and treatment resistance. However, the effects of ER on the function of FoxA1 have been controversial. The aim of this thesis is to further investigate and characterise GR, ER, and FoxA1 crosstalk in three estrogenic breast cancer cell lines, MCF-7, ZR-75-1, and T-47D cells. It has been determined that the combination of dexamethasone (Dex) and E2 have an altered affect on the cell proliferation of breast cancer cells, compared to the single treatment, suggesting GR can modulate the ER response. In an artificial cell model it has been demonstrated by genome-wide investigations, that activated GR and estrogen receptor (ER) can alter the binding of each other at a subset of sites, by a mechanism termed DynaLoad. In addition, it has been shown that Dex and E2 in combination can regulate a unique subset of genes in breast cancer cells. This provides evidence to indicate that Dex can oppose the growth stimulatory effects of E2 signalling, and further, in combination, Dex and E2, can alter the gene transcriptional prolife of MCF-7 breast cancer cells. To understand how the molecular interplay between GR and ER effect breast cancer progression the genome-wide binding events of activated GR and ER have been investigated. These studies show that a GR and ER DynaLoad mechanism also exists in all three breast cancer cell lines utilised; however, there was very little crossover of binding patterns observed. This suggests that while the mechanisms of DynaLoad are present in all three cell lines, the sites altered are cell specific. Most surprisingly is the discovery of an elevated number of GR sites that are lost upon activation of ER in MCF-7 cells. However, in the other breast cancer cell lines, this finding is not as pronounced. Immunblots show that MCF-7 cells have lower GR protein levels than the other cell lines indicating that steroid receptor (SR) levels play a major role in the effect that the dual hormone treatment has on the cell. This suggests that in a highly estrogenic cell line, ER plays a strong role in modulating GR function, which could have important consequences for disease outcome. Furthermore, and contrary to previous findings, this thesis establishes that activated ER and GR have the ability to alter the genomic response of the well-established pioneer factor FoxA1. Genome-wide analysis of FoxA1 binding, upon treatment of E2 or Dex, shows that both ER and GR can recruit FoxA1 to specific binding sites within the genome through a DynaLoad mechanism. These results indicate that there is not a specific set of pioneer factors which bind to closed chromatin and establish the binding landscape for other transcription factors (TFs). Instead this data suggests that every factor has the potential to affect the binding landscape of other TFs, depending on the chromatin context. Overall, the findings from this thesis have provided novel insight into the crosstalk between GR, ER, and FoxA1, further highlighting the ability of activated SRs to alter the response of one another, and other TFs. In addition, it has also been determined that the outcomes of SR crosstalk is cell-specific and that differing estrogenic breast cancer cells can have altered outcomes, which are dependent on SR levels. This can have potential consequences in breast cancer disease outcomes and progression. In addition, the findings in this thesis have begun to shift our classical understanding of pioneer factors in breast cancer, demonstrating that activated GR and ER have the capabilities to recruit and alter the response of FoxA1. This has provided information on a previously unknown complexity to FoxA1 action in breast cancer cells. The studies in this thesis highlight the signalling complexity of TFs in breast cancer cells and provide the basis for further investigations into GR, ER, and FoxA1 mechanisms and the direct consequences of this on breast cancer outcomes.
Advisor: Hager, Gordon
Dissertation Note: Thesis (Ph.D.) -- University of Adelaide, School of Medicine, 2014
Keywords: estrogen receptor; glucocorticoid receptor; progesterone receptor; steroid receptor; forkhead box protein A1; DynaLoad; breast cancer
Provenance: This electronic version is made publicly available by the University of Adelaide in accordance with its open access policy for student theses. Copyright in this thesis remains with the author. This thesis may incorporate third party material which has been used by the author pursuant to Fair Dealing exceptions. If you are the owner of any included third party copyright material you wish to be removed from this electronic version, please complete the take down form located at:
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