Regulation of Estrogen-Responsive Genes

雌激素反应基因的调控

• 批准号: 7163495
• 负责人: Ann Nardulli
• 金额: $ 32.22万
• 依托单位: UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-01-01 至 2008-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7163495
• 关键词: 4-Hydroxy-Tamoxifen; Agar Gel Electrophoresis; Binding; Binding Sites; Biochemical; Biological Assay; Breast; Breast Cancer Cell; Cardiovascular system; Cell Line; Cells; Chromatin; Complex; DNA-Protein Interaction; Development; Elements; Endometrial; Environment; Estrogens; Ethanol; Event; Female; Fertility; Gene Expression; Gene Targeting; Genes; Genetic Transcription; Hormones; Human; In Vitro; Laboratories; MCF7 cell; Maintenance; Mammary Neoplasms; Mammary gland; Mass Spectrum Analysis; Mediating; Methodology; Methods; Modeling; Molecular; Monitor; Neurophysiology - biologic function; Normal tissue morphology; Nuclear Protein; Nuclear Proteins; Numbers; Personal Satisfaction; Pharmacologic Substance; Play; Postmenopause; Process; Progesterone; Progesterone Receptors; Protein Binding; Proteins; Raloxifene; Receptor Gene; Recruitment Activity; Regulation; Response Elements; Role; Selective Estrogen Receptor Modulators; Site; Skeletal system; Symptoms; Techniques; Time; Tissues; Trans-Activators; Transcription Factor AP-1; Transcriptional Activation; Transcriptional Regulation; Transfection; Uterus; chromatin immunoprecipitation; in vivo; insight; male; malignant breast neoplasm; neoplastic cell; novel; prevent; promoter; receptor; receptor expression; reproductive; research study; sperm cell; steroid hormone; transcription factor; tumor

DESCRIPTION (provided by applicant): The steroid hormone estrogen plays a critical role in development and maintenance of female reproductive and mammary tissues, but is also involved in maintenance of cardiovascular, skeletal, and neural function, proliferation of breast cancer cells, and concentration of sperm in the male reproductive tract. Moreover, estrogens and selective estrogen receptor modulators (SERMs) are widely used in regulating fertility, alleviating postmenopausal symptoms, and preventing and treating breast cancer. Although it is widely accepted that the estrogen-occupied receptor mediates its effects by interacting with estrogen response elements (EREs) residing in target genes, a number of estrogen-responsive genes contain no identifiable ERE. To understand how estrogen-responsive genes containing AP-1 and Sp1 sites but lacking EREs respond to hormone treatment, we will study the regulation of the endogenous progesterone receptor (PR) gene. The PR plays a crucial role in reproductive and mammary cell development and fertility and has been used clinically as an indicator of estrogen-responsiveness in human breast cancer tumors. Recent studies from our laboratory indicate that the human PR gene, which lacks an ERE, is controlled by multiple cis elements including an AP-1 site and two Sp 1 sites adjacent to an ERE half site. Thus, the PR gene provides an excellent model to define mechanisms by which target genes lacking an ERE can be regulated by hormone. We will monitor the ordered recruitment of transcription factors and coregulatory proteins to these AP-1 and Sp 1 sites in the endogenous PR gene to delineate the temporal sequence of events required for maintaining basal PR expression and initiating and sustaining high levels of transcription using highly sensitive chromatin immunoprecipitation assays. The cis elements and coregutatory proteins involved in mediating the effects of estrogen and the SERMs 4-hydroxytamoxifen and raloxifene will be determined using in vivo footprinting and chromatin immunoprecipitation experiments. Both mammary and uterine cell lines will be examined to define mechanisms by which these clinically important SERMs mediate their tissue-specific effects. To define mechanisms by which estrogen-responsive genes containing AP-1 and Sp 1 sites are regulated, we will isolate and identify novel coregulatory proteins that associate with AP-1 and Sp 1 sites using agarose gel electrophoresis and mass spectrometry analysis and characterize the effects of these proteins on expression of genes containing AP-1 and Sp 1 sites. The insights gained in these combined studies will help to delineate how estrogens and SERMs regulate expression of genes containing AP-1 and Sp1 sites and enhance our understanding of cellular responsiveness of mammary tumor cells as well as normal tissues such as the uterus and breast to these clinically important pharmaceutical agents.

描述（由申请人提供）：类固醇激素雌激素在女性生殖和乳腺组织的发育和维持中起关键作用，但也参与心血管、骨骼和神经功能的维持、乳腺癌细胞的增殖以及男性生殖道中精子的浓缩。此外，雌激素和选择性雌激素受体调节剂（SERM）被广泛用于调节生育能力，缓解绝经后症状，预防和治疗乳腺癌。虽然人们普遍认为雌激素受体通过与靶基因中的雌激素反应元件（ERE）相互作用来介导其效应，但许多雌激素反应基因不含可识别的ERE。为了了解含有AP-1和Sp1位点但缺乏ERE的雌激素反应基因如何对激素治疗作出反应，我们将研究内源性孕酮受体（PR）基因的调节。PR在生殖和乳腺细胞发育和生育力中起着至关重要的作用，并已在临床上用作人类乳腺癌肿瘤中雌激素反应性的指标。本实验室最近的研究表明，人类PR基因是由多个顺式元件控制的，其中包括一个AP-1位点和两个与ERE半位点相邻的Sp-1位点。因此，PR基因提供了一个很好的模型来定义缺乏ERE的靶基因可以被激素调节的机制。我们将监测这些AP-1和SP 1网站在内源性PR基因的转录因子和共调节蛋白的有序招聘描绘的时间序列的事件所需的维持基础PR表达和启动和维持高水平的转录，使用高灵敏度染色质免疫沉淀试验。将使用体内足迹法和染色质免疫沉淀实验确定参与介导雌激素和SERM 4-羟基他莫昔芬和雷洛昔芬作用的顺式元件和coregutatory蛋白。将检查乳腺和子宫细胞系，以确定这些临床上重要的SERM介导其组织特异性作用的机制。要定义的机制，雌激素反应基因含有AP-1和Sp 1网站的监管，我们将分离和鉴定新的共调节蛋白，与AP-1和Sp 1网站使用琼脂糖凝胶电泳和质谱分析，并表征这些蛋白对表达的影响，含有AP-1和Sp 1网站的基因。在这些综合研究中获得的见解将有助于阐明雌激素和SERM如何调节含有AP-1和Sp1位点的基因表达，并增强我们对乳腺肿瘤细胞以及正常组织（如子宫和乳腺）对这些临床重要药物的细胞反应性的理解。