STEROIDS AND GROWTH

类固醇与生长

• 批准号: 3163127
• 负责人: GEOFFREY L GREENE
• 金额: $ 28.37万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 1977
• 资助国家: 美国
• 起止时间: 1977-04-01 至 1994-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3163127
• 关键词: DNA binding protein; MCF7 cell; breast neoplasms; cell differentiation; cell growth regulation; chemical binding; chemical structure function; complementary DNA; estrogen receptors; female; gene expression; genetic transcription; genetic translation; hormone binding protein; hormone regulation /control mechanism; hormone related neoplasm /cancer; human tissue; laboratory rat; mammary gland; messenger RNA; molecular cloning; molecular oncology; monoclonal antibody; neoplasm /cancer genetics; neoplastic cell; phosphorylation; progesterone receptors; protein engineering; protein sequence; recombinant DNA; tissue /cell culture; uterus

The major goal of this investigation is to elucidate the role of specific intracellular receptor proteins in the estrogenic regulation of cell proliferation and function in hormone-responsive tissues and cancers derived from these tissues. The proposal focuses on the use of biochemical, immunochemical and recombinant DNA techniques to determine the structure, composition and intracellular interactions and dynamics of the estrogen receptor and progesterone receptor from human or rodent sources. We are especially interested in the molecular mechanisms by which activity and expression of these receptors are regulated by estrogens, progestins and their antagonists in breast cancer cell lines. This work is facilitated by the availability in our laboratory of purified, or partially purified receptor proteins, monoclonal antibodies specific for both receptors, and cloned cDNAs that correspond to all of the translated mRNA for human ER from MCF-7 breast cancer cells and 90% of the translated mRNA for human PR from T47D breast cancer cells. Specific aims include: 1) To isolate, sequence and characterize cDNA clones corresponding to the entire mRNA for T47D human PR; 2) to isolate, sequence and characterize genomic clones for human ER and PR, including upstream regulatory sequences; 3) to characterize the regulation of ER and PR expression in breast cancer cells and rat uterus by estrogens, progestins and their antagonists; 4) to locate and characterize ER and any PR binding sites (responses elements) on the chromosomal genes for both receptors; 5) to dissect and characterize the functional domains of both receptors by in vitro mutagenesis, including domain switching between ER and PR, coupled with transient expression assays in heterologous cells containing estrogen- or progestin-responsive reporter genes; 6) to isolate mutant fragments either from transfected cells or from in vitro translations of transcripts and study the steroid-binding and specific DNA-binding properties of these fragments; 7) to overexpress human ER and PR cDNAs in suitable cell lines, purify expressed receptor and carry out detailed structural and biochemical studies on full length receptor or genetically engineered receptor fragments; 8) to determine the amino acids involved in the binding of both agonists and antagonists, and the location, role and hormonal regulation of phosphorylation sites in rat and human ER and PR.

这项调查的主要目的是阐明 雌激素中特定的细胞内受体蛋白 调节激素反应性的细胞增殖和功能 源自这些组织的组织和癌症。 提案 专注于使用生化，免疫化学和重组 DNA技术来确定结构，组成和 雌激素受体的细胞内相互作用和动力学 来自人类或啮齿动物来源的孕激素受体。 我们是 特别对活性的分子机制感兴趣 这些受体的表达受雌激素调节， 孕激素及其在乳腺癌细胞系中的拮抗剂。 这 我们的实验室的可用性促进了工作 纯化或部分纯化的受体蛋白，单克隆 对两种受体的特异性抗体和克隆的cDNA 对应于MCF-7的所有人类ER的翻译mRNA 乳腺癌细胞和90％的人类PR的翻译mRNA 来自T47D乳腺癌细胞。 具体目的包括：1） 分离，序列和表征对应于 T47D人类PR的整个mRNA； 2）分离，序列和 表征人类ER和PR的基因组克隆，包括上游 调节序列； 3）表征ER的调节和 乳腺癌细胞和大鼠子宫中的PR表达，雌激素， 孕激素及其对手； 4）定位和表征ER 以及染色体上的任何PR结合位点（响应元素） 两个受体的基因； 5）剖析和描述 通过体外诱变，两种受体的功能结构域， 包括在ER和PR之间切换的域，与 含有的异源细胞中的瞬态表达测定 雌激素或孕激素反应性报告基因； 6）分离 来自转染细胞或体外的突变片段 转录本的翻译并研究类固醇结合和 这些片段的特定DNA结合特性； 7） 过表达的人ER和PR cDNA在合适的细胞系中，净化 表达受体并执行详细的结构和 关于全长受体或遗传学的生化研究 工程受体碎片； 8）确定氨基酸 参与激动剂和拮抗剂的结合，以及 磷酸化位点的位置，角色和激素调节 老鼠和人类ER和PR。

项目成果

Estrogen receptor determination in endometrial carcinoma: ligand binding assay versus enzyme immunoassay.

子宫内膜癌中雌激素受体的测定：配体结合测定与酶免疫测定。

• 发表时间: 1995
• 期刊: Anticancer research.
• 作者: Nyholm,HC;Nielsen,AL;Lyndrup,J;Greene,GL;Thorpe,SM
• 通讯作者: Thorpe,SM

Immunostaining of estrogen receptor in paraffin sections of breast carcinomas using monoclonal antibody D75P3 gamma: effects of fixation.

使用单克隆抗体 D75P3 γ 对乳腺癌石蜡切片中的雌激素受体进行免疫染色：固定的效果。

• DOI: 10.1097/00000478-198712000-00004
• 发表时间: 1987
• 期刊: The American journal of surgical pathology
• 作者: DeRosa,CM;Ozzello,L;Greene,GL;Habif,DV
• 通讯作者: Habif,DV

The use of monoclonal antibodies to estrogen receptors (ER) for immunoperoxidase detection of ER in paraffin sections of human breast cancer tissue.

使用雌激素受体 (ER) 单克隆抗体通过免疫过氧化物酶检测人乳腺癌组织石蜡切片中的 ER。

• DOI: 10.1177/33.2.2578501
• 发表时间: 1985
• 期刊: The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society
• 作者: Poulsen,HS;Ozzello,L;King,WJ;Greene,GL
• 通讯作者: Greene,GL

Receptors reconsidered: a 20-year perspective.

受体重新思考：20 年的视角。

• DOI: 10.1016/b978-0-12-571138-8.50006-8
• 发表时间: 1982
• 期刊: Recent progress in hormone research
• 作者: Jensen,EV;Greene,GL;Closs,LE;DeSombre,ER;Nadji,M
• 通讯作者: Nadji,M

Immunohistochemical assessment of estrogen and progesterone receptors in stored imprints and cryostat sections of breast carcinomas.

对储存的乳腺癌印记和低温切片中的雌激素和孕激素受体进行免疫组织化学评估。

• DOI: 10.1097/00000658-198908000-00015
• 发表时间: 1989
• 期刊: Annals of surgery
• 影响因子: 9
• 作者: DeRosa,CM;Ozzello,L;Habif,DV;Konrath,JG;Greene,GL
• 通讯作者: Greene,GL