STRUCTURE/FUNCTION OF GROWTH HORMONE RELATED GENE

生长激素相关基因的结构/功能

• 批准号: 2141634
• 负责人: NORMAN L EBERHARDT
• 金额: $ 20.14万
• 依托单位: MAYO CLINIC ROCHESTER
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-07-01 至 1998-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2141634
• 关键词: conformation; estrogens; gel mobility shift assay; genetic promoter element; genetic regulatory element; genetic transcription; hormone receptor; hormone regulation /control mechanism; molecular cloning; mutant; nuclear runoff assay; nucleic acid sequence; nucleic acid structure; posttranscriptional RNA processing; reporter genes; retinoate; site directed mutagenesis; somatomammotropin; somatotropin; thyroid hormones; tissue /cell culture; transcription factor; transcription termination; transfection; triiodothyronine

Human (h) growth hormone (GH), chorionic somatomammotropin (CS) and prolactin (PRL) comprise a family of genes that are essential for normal human growth and development. Altered GH production has been implicated in various pathological states, including osteoporosis and aging. Knowledge of the regulation of these genes is important for our understanding of their physiologic and pathophysiologic functions. The proposed studies extend previous investigations on the hormonal and tissue-specific factors that mediate transcript ion of hGH/hCS genes. Aim 1 focuses on the functional significance of DNA bending induced by the thyroid hormone receptor (TR), cell-specific factor GHF1 and Sp1. The contributions of intrinsic and transcription factor-induced DNA bending will be established by circular permutation and phasing analyses. The influence of DNA phasing-induced changes on overall promoter conformation will be correlated with promoter activities using transient transfection assays and in vitro transcription systems. TR mutants that exhibit coupled and uncoupled DNA binding and transcriptional activation responses will be examined by bending analyses to determine whether DNA bending is coupled to DNA binding or transactivation functions. Aim 2 will examine the structure and mechanism of action of a unique, promoter-dependent negative thyroid hormone response element (nTRE) located in the 3'-flanking DNA of the hGH gene. The precise structure of the nTRE will be established utilizing mutagenesis and analysis of functional activity with transfection assays. The hGH promoter elements that are required for nTRE function will be established by mutagenesis of the hGH 5'-flanking region/promoter and functional analysis using transient transfection assays. The mechanism of nTRE action will be examined by a combination of run-on transcription, in vitro transcription and in vitro polyadenylation and/or splicing experiments to establish whether nTRE action is coupled to transcript ion initiation, elongation or termination/3'-end formation. The physiologic relevance of nTRE function on hGH gene expression will be examined by the ongoing studies of human acromegalic tumor cells. These studies will extend our understanding of the factors that regulate the hormonal and tissue-specific regulation of hGH/hCS gene expression.

人（H）生长激素（GH），绒毛膜瘤瘤（CS）和 催乳素（PRL）包括一个对正常必不可少的基因家族 人类的成长与发展。 GH生产的改变已被牵涉到 在各种病理状态，包括骨质疏松和衰老。 了解这些基因的调节对我们很重要 了解其生理和病理生理功能。这 拟议的研究扩展了先前对激素和激素的研究 介导HGH/HCS基因转录离子的组织特异性因子。 目的 1侧重于由DNA弯曲的功能意义 甲状腺激素受体（TR），细胞特异性因子GHF1和SP1。 这 内在和转录因子诱导的DNA弯曲的贡献 将通过循环排列和分阶段分析来建立。 这 DNA相位诱导的变化对总体启动子构象的影响 将使用瞬态转染与启动子活动相关 测定和体外转录系统。表现出耦合的TR突变体 和未耦合的DNA结合和转录激活反应将是 通过弯曲分析检查以确定是否耦合DNA弯曲 DNA结合或反式激活功能。 AIM 2将检查 独特的启动子依赖性负面的结构和作用机理 甲状腺激素反应元件（NTRE） HGH基因。 NTRE的确切结构将建立 利用诱变和功能活性分析 转染测定法。 NTRE所需的HGH启动子元素 功能将通过HGH 5'FARKing的诱变来确定 使用瞬态转染的区域/启动子和功能分析 测定。 NTRE动作的机制将通过结合 跑步转录，体外转录和体外聚腺苷酸化 和/或剪接实验，以确定NTRE动作是否耦合到 成绩单离子启动，伸长或终止/3'末端形成。 NTRE功能在HGH基因表达上的生理相关性将是 通过正在进行的人类肿瘤肿瘤细胞的研究进行了检查。 这些 研究将扩展我们对调节的因素的理解 HGH/HCS基因表达的激素和组织特异性调节。