REGULATION OF THE GAL GENES IN YEAST

酵母中 gal 基因的调控

• 批准号: 3281021
• 负责人: MARK S PTASHNE
• 金额: $ 38.63万
• 依托单位: HARVARD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1983
• 资助国家: 美国
• 起止时间: 1983-07-01 至 1993-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3281021
• 关键词: DNA binding protein; DNA directed RNA polymerase; Drosophilidae; Saccharomyces; chromosome deletion; eukaryote; fungal genetics; galactose; gene deletion mutation; gene expression; genetic promoter element; genetic transcription; mutant; nucleic acid sequence; nucleic acid structure; point mutation; protein structure

The longterm objective of our work is to understand how, in eukaryotes, regulatory proteins bind to specific DNA sequences and turn genes on and off. Our experiments focus on the yeast activator GAL4 that binds to the so called galactose upstream sequence (UASG) and turns on transcription of flanking genes at distance spanning many hundreds of base pairs. This activator also turns on gene expression in mouse cells in tissue culture, and understanding the mechanism of action of GAL4 may provide us with very general insights into the molecular basis of gene regulation. We will continue our characterization of the activating surfaces of yeast transcriptional activators: we will isolate mutants affecting the activating region(s) of GAL4, design synthetic activating regions and attempt to isolate new classes of activating regions that stimulate genes not activated by GAL4. We will study the effects of yeast activatorS in mammalian and Drosophila cells. we will study the basis of the DNA binding specificity of GAL4: we will isolate mutants with altered or relaxed specificity for various defined mutant operators and test whether the "cysteine fIngers of GAL4 and PPRl are responsible for sequence recognition by studying hybrids of these two molecules. We will define the part(s) of GAL4 that mediate(s) cooperative binding of GAL4 to multiple sites in vivo. We will attempt to reproduce cooperative binding of GAL4 to reiterated sites in vitro. We will study the activities of various GAL4-derived activators on constructs that vary the distance of the GAL4 binding site from the transcriptional start site. We will identify extragenic suppressors of GAL4 mutants deficient in the activation function; these suppressors might define the transcription factor(s) (e.g. RNA polymerase?) that interact with the activation function of GAL4. We will characterize the mechanism(s) by which growth in glucose blocks GAL4's stimulatory activity. We will study negative control at a distance by placing the GAL1 promoter under control of the HMR silencer and analyzing the effect on GAL4 binding and activity in vivo.

我们工作的长期目标是了解如何 真核生物，调节蛋白与特定的DNA序列和 打开和关闭基因。 我们的实验专注于酵母 激活剂gal4与上游的所谓半乳糖结合 序列（UASG）并打开侧面基因的转录 跨越数百个基对的距离。 此激活剂也是如此 在组织培养中的小鼠细胞中打开基因表达，并 了解GAL4的作用机理可能会为我们提供 对基因调节的分子基础的非常一般的见解。 我们将继续对激活表面的表征 酵母转录激活剂：我们将分离突变体 影响GAL4的激活区域，设计合成 激活区域并尝试隔离新类别的激活 刺激未被GAL4激活的基因的区域。 我们将学习 酵母活化剂在哺乳动物和果蝇细胞中的影响。 我们将研究GAL4的DNA结合特异性的基础： 我们将以改变或放松的特异性分离突变体 各种定义的突变算子，并测试“半胱氨酸 GAL4和PPRL的手指负责序列识别 通过研究这两个分子的杂种。 我们将定义 GAL4的一部分介导GAL4与 体内多个站点。 我们将尝试重现合作社 GAL4与体外重复位点的结合。 我们将研究 各种GAL4衍生的激活剂的活动 改变GAL4结合位点与转录的距离 启动站点。 我们将确定GAL4的外部抑制器 突变体缺乏激活函数；这些抑制器 可以定义转录因子（例如RNA聚合酶？） 与GAL4的激活函数相互作用。 我们将 表征葡萄糖块中生长的机制 GAL4的刺激活动。 我们将研究一个负面对照 通过将GAL1启动子置于HMR的控制下，距离 消音和分析对GAL4结合和活性的影响 体内。