Dose-Dependent Effects of Hormone on the Activity of Estrogen Receptor Enhancers and Target Genes

Ut Southwestern Medical Center, Dallas TX

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Abstract

Project Summary/Abstract Cellular signaling by estrogens plays a critical role in the development and physiology of a wide variety of tissues, including the reproductive organs, mammary glands, bone, heart, vasculature, adipose, liver, and central nervous system, as well as diseases of the same tissues. Within an individual, estrogen levels may fluctuate dramatically and acutely across multiple time scales: hours, days, months, years, and decades. The circulating levels of 17b- estradiol (E2), the predominant naturally-occurring estrogen, fluctuate dramatically in women from lows of ~70 pM to highs of ~1.8 nM in the non-pregnant state. In contrast, most molecular, cellular, and genomic studies use saturating supraphysiological levels of E2 (e.g., 10-100 nM). Although the importance of dose, and the frequency and duration of administration, have long been recognized as important aspects of the therapeutic use of steroid hormones, the effects of physiological doses of E2 on molecular, cellular, and genomic endpoints have not been well characterized. Our proposed studies will address these key gaps in knowledge. The molecular actions of E2 are mediated through estrogen receptor proteins (e.g., ERa), which function primarily as ligand-regulated transcription factors that drive cell type-specific patterns of gene expression by promoting the coordinated assembly of transcriptional enhancers at ERa binding sites. There is a growing interest in the mechanisms and functions of signal-regulated enhancers, but many questions remain. For example, we do not have a detailed understanding of (1) how different doses of E2 may differentially regulate the assembly and function of ERa enhancers across the genome and (2) how different dose sensitivities of ERa enhancers may lead drive distinct biological outcomes in different estrogen-responsive biological systems. The overarching goal of these studies is to gain a better understand the molecular mechanisms by which E2 controls ERa enhancers to regulate gene expression and downstream biological outcomes. Our broad hypothesis, which is supported by considerable preliminary data, is that different ERa enhancers and target genes exhibit different sensitivities to the dose of E2, specifying different classes of enhancers with different mechanisms of action. We will test this hypothesis in two ERa-positive cell types (i.e., human breast cancer cells and primary mouse myometrial cells) using an integrated set of experimental approaches. Our specific aims are to: (1) Characterize dose-effects of E2 on molecular and genomic endpoints of estrogen signaling (Aim 1); (2) Determine the molecular mechanisms by which different doses of E2 promote enhancer activity (Aim 2); and (3) Explore genetic determinants underlying dose-effects of E2 on biological endpoints (Aim 3). Our studies on the molecular mechanisms and functions of ERa enhancers seek to challenge and expand current approaches and biological models for studying the mechanisms of steroid hormone signaling. These studies will yield new knowledge about the molecular mechanisms of E2 signaling; a diverse collection of genomic data; and new concepts about E2 signaling that may suggest new ways to target ERa therapeutically.