SEX-SPECIFIC REGULATION OF TRANSCRIPTION IN DROSOPHILA

果蝇性别特异性转录调控

• 批准号: 3305630
• 负责人: PIETER C WENSINK
• 金额: $ 14.57万
• 依托单位: BRANDEIS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-07-01 至 1995-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3305630
• 关键词: DNA binding protein; DNA footprinting; Drosophilidae; affinity chromatography; chemical binding; developmental genetics; gel filtration chromatography; gene expression; genetic enhancer element; genetic mapping; genetic promoter element; genetic regulatory element; genetic transcription; ion exchange chromatography; molecular genetics; nucleic acid sequence; polymerase chain reaction; protein purification; protein structure function; regulatory gene; sex determination; stoichiometry

This project will investigate the molecular mechanism by which a major, well-characterized developmental regulatory hierarchy of Drosophila melanogaster directs the sex-specificity of target gene transcription. The significance of the project lies in its advanced analysis of the structure and function of a cellular enhancer from a higher organism and in its study of the direct molecular connection between a developmental regulatory hierarchy and target genes. Understanding developmental regulation and cellular enhancer function is crucial to understanding defects in gene expression that lead to human disease and human developmental abnormalities. The project will investigate a particular enhancer that conveys the male- specific repression signal from the hierarchy to a pair of target genes. We will determine how this repression signal is received by the enhancer and then sent to the promoters. A gene at the bottom of the regulatory hierarchy encodes a male-specific repressor of female genes and a female- specific repressor of male genes. In vitro both male and female repressors bind to the male-specific regulatory elements of the enhancer. We will examine the mechanism operating in vivo to determine whether the male repressor is sufficient to cause repression in males and whether the female repressor does not bind these elements, or does bind them but does not repress. This will be done by using in vitro mutagenesis/germline transformation methods to identify the functional elements within the enhancer and to determine whether the bound repressor acts on other enhancer elements or directly on the promoter. These studies will show whether this repressor action occurs by steric hindrance or DNA looping mechanisms. We will also investigate the biochemistry of the interactions between the DNA sites and the purified, expressed, repressors. This biochemical portion of the project aims at an explanation for the male and female repressor activities at this male-specific enhancer. We will then use a transcription system derived from an appropriate purified tissue in order to reproduce this sex-specific repression in vitro and investigate the function of proteins that operate in the repression mechanism.

该项目将研究主要的分子机制， 果蝇特征明确的发育调控层次 黑腹果蝇指导靶基因转录的性别特异性。 这 该项目的意义在于其对结构的高级分析 高等生物细胞增强剂的功能及其研究 发育调节之间的直接分子联系 层次结构和目标基因。 了解发育调节和 细胞增强子功能对于理解基因缺陷至关重要 导致人类疾病和人类发育的表达 异常。 该项目将研究一种特定的增强子，它可以传递男性的 从层次结构到一对靶基因的特定抑制信号。 我们将确定增强子如何接收该抑制信号 然后发送给发起人。 处于监管底层的基因 等级制度编码了女性基因的男性特异性抑制因子和女性基因 男性基因的特异性抑制因子。 体外雄性和雌性抑制子 与增强子的雄性特异性调节元件结合。 我们将 检查体内运作机制以确定男性是否 抑制因子是否足以对男性造成抑制，而女性是否会受到抑制 阻遏蛋白不结合这些元件，或者结合它们但不结合 抑。 这将通过使用体外诱变/种系来完成 转换方法来识别内部的功能元素 增强子并确定结合的阻遏物是否作用于其他 增强子元件或直接位于启动子上。 这些研究将表明 这种抑制作用是通过空间位阻还是 DNA 环发生的 机制。 我们还将研究相互作用的生物化学 DNA 位点和纯化的、表达的阻遏物之间。 这 该项目的生化部分旨在解释男性和女性 女性在这种男性特异性增强子上的抑制活动。 我们随后将 使用源自适当纯化组织的转录系统 为了在体外重现这种性别特异性抑制并进行研究 在抑制机制中起作用的蛋白质的功能。