THYROTROPIN-BETA SUBUNIT GENE TRANSCRIPTION FACTORS

促甲状腺素-β亚基基因转录因子

• 批准号: 2139893
• 负责人: Eli CHESTER RIDGWAY
• 金额: $ 24.27万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 1985
• 资助国家: 美国
• 起止时间: 1985-09-01 至 1998-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2139893
• 关键词: DNA binding protein; DNA footprinting; DNA methylation; gel mobility shift assay; gene expression; genetic promoter element; genetic regulation; genetic regulatory element; genetic transcription; hormone regulation /control mechanism; laboratory mouse; molecular cloning; mutant; nucleic acid sequence; protein isoforms; protein purification; protein reconstitution; protein structure function; site directed mutagenesis; thyrotropin; tissue /cell culture; transcription factor; transfection

Thyrotropin (TSH) is the pituitary glycoprotein hormone which controls the growth and function of the thyroid gland. It is produced solely by thyrotrope cells, one of five terminally differentiated pituitary cell types. The TSH molecule is composed of two nonidentical glycosylated subunits, alpha and TSHbeta, which are noncovalently associated. These two subunits arise independently from separate genes located on different chromosomes. The alpha-subunit gene is expressed not only in TSH cells but also pituitary gonadotropes and placental chorionic gonadotropes. In contrast, TSH-beta gene expression is restricted only to thyrotropes. The studies outlined in this grant proposal are firmly based on previous studies from this laboratory in which the gene for the murine TSH-beta subunit has been isolated and sequenced, its promoter-region characterized, the important functional cis-acting DNA elements identified, and the putative areas of protein-DNA interactions on the promoter determined. Our proposed investigations will be to first define the DNA sequences contained in the functionally most important footprint of the TSH-beta promoter which are responsible for thyrotrope-specific expression of this gene. We will then analyze the role of our newly discovered thyrotrope-specific variant isoform of the Pit-1 protein. The studies are designed to understand precisely the role of this novel factor on TSH-beta promoter activation. We will then attempt to reconstitute TSH- beta promoter activation in stably transfected cells with the unique Pit-1 isoforms. For these studies, we will utilize another unique reagent developed in this laboratory, an isogenic alpha-TSH cell line which has lost the ability to express the TSH-beta subunit gene. Finally, we will utilize our precise understanding of the cis-acting determinants of the TSH-beta promoter to identify and clone other novel DNA-binding factors in thyrotrope cells which activate the TSH-beta promoter. These projects will utilize the most modern techniques of cell biology and molecular endocrinology. Recombinant DNA technology will be coupled with novel methods analyzing gene expression in unique cell lines. These studies will provide fundamental information on pituitary cell differentiation and cell-specific gene expression which is directly applicable to human thyroid-pituitary disease.

甲状腺激素（TSH）是垂体糖蛋白激素，它控制 甲状腺的生长和功能。它仅由 甲状腺细胞，五个末端分化的垂体细胞之一 类型。 TSH分子由两个非相同的糖基化组成 亚基，α和tshbeta，它们无共价相关。这两个 亚基独立于不同的基因 染色体。 α-亚基基因不仅在TSH细胞中表达，还表达 还有垂体性腺体和胎盘绒毛膜促性腺激素。在 对比，TSH-β基因表达仅限于甲状腺。 该赠款提案中概述的研究牢固地基于以前的 该实验室的研究，其中鼠TSH-Beta的基因 亚基已被隔离和测序，其启动子区域 表征，重要的功能性顺式作用DNA元件 确定，以及蛋白-DNA相互作用的推定区域 启动子确定。我们提议的调查将首先定义 功能上最重要的足迹中包含的DNA序列 TSH-BETA启动子的促甲状腺特异性 该基因的表达。然后，我们将分析我们新的角色 发现的PIT-1蛋白的甲状腺特异性变体同工型。这 研究旨在准确理解这一新因素的作用 在TSH-BETA启动子激活上。然后，我们将尝试重新建立TSH- 具有独特的PIT-1的稳定转染细胞中的β启动子激活 同工型。对于这些研究，我们将利用另一种独特的试剂 在该实验室开发的，是一种ISEGENICα-TSH细胞系，具有 失去表达TSH-β亚基基因的能力。最后，我们会的 利用我们对顺式作用决定因素的精确理解 tsh-beta启动子以识别和克隆其他新型的DNA结合因子 激活TSH-β启动子的甲状腺细胞。 这些项目将利用细胞生物学的最现代技术和 分子内分泌学。重组DNA技术将与 新方法分析独特细胞系中的基因表达。这些 研究将提供有关垂体细胞的基本信息 分化和细胞特异性基因表达直接 适用于人类甲状腺垂体疾病。