TISSUE-SPECIFIC EXPRESSION OF CRH TRANSGENES

CRH 转基因的组织特异性表达

• 批准号: 3429350
• 负责人: Sally A. Camper
• 金额: $ 7.52万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-03-01 至 1992-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3429350
• 关键词: corticotropin releasing factor; fusion gene; gene expression; genetic manipulation; genetic transcription; genetically modified animals; histochemistry /cytochemistry; in situ hybridization; laboratory mouse; molecular cloning; neuroendocrine system; oncogenes; paraventricular nucleus; pituitary adrenal axis; stress; tissue /cell culture; transcription factor

Glucocorticoids (GC) produced by the adrenal cortex mediate the body's adaptive response to stress. The synthesis and secretion of these steroids is regulated by the hypothalamic-anterior pituitary system. In response to stressful stimuli, the hypothalamus is induced to secrete corticotrophin releasing hormone (CRH) . CRH is transported via the blood to the anterior pituitary where it stimulates the secretion of the neurohormone ACTH, which then stimulates the adrenal cortex to synthesize and secrete GC. Feedback inhibition by GC on both the anterior pituitary and the hypothalamus completes the regulatory loop. Abnormalities in hypothalamic-pituitary-adrenal (HPA) axis function can result in a number of clinical disorders including depression. Depressed patients show hypersecretion of CRH, consistent with the findings that CRH influences motor activity, feeding, and sexual behaviors, all functions which are altered by depression. Knowledge of the regulation of CRH gene expression will provide the groundwork for understanding both normal and abnormal responses to stress. The first step will be to establish the basis for tissue-specific activation of the CRH gene in the paraventricular nucleus of the hypothalamus and other localized regions of the brain and periphery. Transgenic mice bearing the rat CRH gene and varied amounts of 5' flanking DNA will be constructed and tissues analyzed by in situ hybridization in order to locate the cis-acting DNA sequences necessary and sufficient for appropriate expression of the transgene. Immortalized cell lines producing gene products characteristic of differentiated hypothalamic cells would be invaluable for studying CRH gene regulation. To achieve this, tissue-specific elements from the CRH gene will be engineered onto a gene encoding a selectable marker (neomycin) as well as an oncogene (temperature sensitive SV40 T-antigen) and the hybrid genes used to produce transgenic mice. Cell lines will be derived from these animals at a young age using the selective agent and expression studies done at the nonpermissive temperature. CRH transgenic mouse studies will shed light on the intricacies of cell-specific expression in the brain and hypothalamus, develop cell lines necessary to study regulation of gene expression, and thus provide the basis for understanding clinically important syndromes such as depression.

肾上腺皮质产生的糖皮质激素 (GC) 介导身体的 对压力的适应性反应。这些类固醇的合成和分泌 受下丘脑-垂体前叶系统调节。作为回应 应激刺激时，诱导下丘脑分泌促肾上腺皮质激素 释放激素（CRH）。 CRH通过血液输送到前部 垂体刺激神经激素 ACTH 的分泌， 然后刺激肾上腺皮质合成并分泌GC。反馈 GC 对垂体前叶和下丘脑的抑制作用 完成监管循环。异常情况 下丘脑-垂体-肾上腺 (HPA) 轴功能可导致许多 包括抑郁症在内的临床疾病。抑郁症患者表现 CRH 分泌过多，与 CRH 影响的发现一致 运动活动、进食和性行为，所有这些功能 因抑郁而改变。 CRH基因表达调控的知识 将为理解正常和异常提供基础 对压力的反应。第一步将是建立基础 室旁核中 CRH 基因的组织特异性激活 下丘脑以及大脑和外周的其他局部区域。 携带大鼠 CRH 基因和不同数量的 5' 侧翼的转基因小鼠 将通过原位杂交构建 DNA 并分析组织 为了找到必要且充分的顺式作用DNA序列 转基因的适当表达。永生化细胞系生产 分化的下丘脑细胞的基因产物特征是 对于研究 CRH 基因调控具有不可估量的价值。为了实现这一目标， 来自 CRH 基因的组织特异性元件将被工程化到基因上 编码可选择标记（新霉素）以及癌基因（温度 敏感的 SV40 T 抗原）和用于产生转基因的杂交基因 老鼠。细胞系将在这些动物年轻时使用 在非许可条件下进行的选择剂和表达研究 温度。 CRH 转基因小鼠研究将揭示 大脑和下丘脑细胞特异性表达的复杂性， 开发研究基因表达调控所需的细胞系，以及 从而为理解临床上重要的综合征提供基础 比如抑郁症。