Inflammation Alters Transcription by ER in Breast Cancer

Institution: University of California, San Diego
Investigator(s): Eliot Bourk, BA -
Award Cycle: 2006 (Cycle 12) Grant #: 12GB-0015 Award: $75,051
Award Type: Dissertation Award
Research Priorities
Biology of the Breast Cell>Pathogenesis: understanding the disease



Initial Award Abstract (2006)

Estrogen acts through the estrogen receptor (ER), a powerful regulator of cell behavior that can switch specific genes either on or off. While most of the known genes regulated by ER are activated in response to estrogen, we have recently discovered a mechanism by which estrogen can act through ER to specifically decrease the expression of other genes as well. These genes, which are normally repressed by estrogen, can be reactivated if the breast cells are subsequently exposed to inflammation.

The body’s immune system is ideally adapted to fighting off pathogenic microorganisms like bacteria and viruses. However, inflammation in the wrong context can harm us by driving the progression of diseases including breast cancer. Inflammation has been demonstrated by many breast cancer studies to correlate with increased aggressive tumor behavior, including angiogenesis and metastasis. Tumor-associated macrophages produce pro-inflammatory cytokines such as IL-1beta that influence the behavior of breast cancer cells. In previous work in my mentor’s (Dr. Michael Rosenfeld) lab we identified a specific macrophage/cancer cell interaction that causes de-repression of ER target genes.

This project is based on preliminary data obtained using an exclusive technology developed in our laboratory (ChIP-DSL) that enabled us to rapidly scan the genome for genes regulated by the estrogen receptor. Our pilot experiments have enabled us to narrow down the list of potential negative ER target genes from ~20,000 down to 18 high probability targets. This selection process allows us to focus on the ER-mediated gene repression machinery within breast cancer cells and determine how certain genes are reactivated under the influence of localized inflammatory conditions.

The overall questions addressed by this research are:
• Which genes are directly shut off by ER in response to estrogen?
• How are these genes reactivated by inflammation?

Our experiments will involve techniques for investigating gene expression, using the polymerase chain reaction (PCR) method to measure the expression levels of the ER target genes, the chromatin immunoprecipitation (ChIP) method to reveal the machinery that regulates these genes, and RNA interference (RNAi) to block the expression of specific proteins that may regulate these genes.

This project will also contribute the first description of this new class of estrogen regulated genes that are subject to control by inflammation, and will provide insight into how breast cancer cells can reactivate these pathogenic genes in response to inflammation.




Final Report (2008)
Estrogen acts in its target tissues through the estrogen receptor (ER), a powerful regulator of cell behavior that can switch specific genes either on or off While most research on ER function has focused on its ability to activate genes, we and others have noted that several genes can be repressed in response to estrogen. The overall scientific goal of this research project was to determine which genes are directly shut off by ER in response to estrogen, and the mechanism by which some of these genes are reactivated by inflammation.

Our first aim was to define the genome-wide program of negative estrogen receptor targets. I compared data from gene expression profiling (microarray) experiments and genome wide location analyses and identified a sub-set of estrogen-repressed genes that are located near ER binding sites in the genome, and confirmed their estrogen regulation in MCF7 breast cancer cells. Some of these genes are ABCG2, BCL3, BHLHB2, CORO2A, ESR1, FOXA1, HBP1, INHBB, NCOA3, RBL2, SUV420H1, and ZNF217. Most of these genes encode transcription factors and cofactors, supporting the idea that ER regulates gene expression networks at multiple levels: directly, by binding to its sites in the genome and assembling cofactor complexes that increase or decrease expression of nearby genes; and indirectly, by regulating expression of other transcription factors and cofactors, thereby affecting each of their individual gene expression programs (including other transcription factors and cofactors). Part of the ER repression program uncovered in this project appears to be a negative feedback loop, decreasing the expression of ER itself as well as other genes that promote ER activity. For example, FOXA1 is a transcription factor known to bind near ER sites in the genome and improve DNA binding of ER at these sites.

Our second aim was to uncover the mechanism for de-repression by inflammatory signals, such as IL (interleukin)-1beta. Our focus was “bone morphogenic protein 7” (BMP7), a member of the TGF-ß superfamily. I have identified certain factors downstream of the IL-1 receptor that are essential for de-repression of BMP7 by IL-1beta; when the expression of any of these proteins is knocked down by siRNA (RNA interference) or their activity is interfered with by expression of a dominant negative, the BMP7 gene stays repressed instead of being reactivated by IL-1beta.

A novel discovery in this project was a previously unknown difference between the two human estrogen receptors, ER-alpha and ER-beta. These two receptors are expressed to varying degrees in the different tissues f the body, with some cells containing predominately one receptor or the other, and other cells expressing both receptors to a similar degree. We found that repression of certain genes by estrogen can be reversed in ER-alpha expressing cells in the presence of inflammation, these genes will remain repressed in ER-beta expressing cells despite inflammation. Further, I have proposed a simple biochemical explanation for this difference between the two estrogen receptors. TAB2 is a signaling component downstream of the IL-1 receptor, and is a key molecule required for de-repression of ER-alpha-repressed genes by inflammation.

In the future, I will work on optimizing the genome-wide localization studies of ER co-regulators and signal transduction proteins involved in the de-repression of certain ER-repressed genes by inflammatory signaling. The results of these studies should help further our understanding of how ER binding at different sites in the genome leads to divergent effects on gene expression.


Symposium Abstract (2007)
Estrogen acts through the estrogen receptor (ER), a powerful regulator of cell behavior that can switch specific genes either “on” or “off.” While most of the known genes regulated by ER are activated in response to estrogen, we have recently discovered a mechanism by which estrogen can act through ER to specifically decrease the expression of other genes as well. These genes, which are normally repressed by estrogen, can be reactivated if the breast cells are subsequently exposed to inflammation. The overall questions addressed by this research are: