Molecular Analysis Transcription Factors Regulating phyA Gene Expression

9513590 Quail这项提议的重点是先前克隆的转录因子GT-2。具体目标是：(A)对GT-2的功能活性进行进一步的详细分析，包括使用转染法进行转录激活分析以确定参与反式激活的GT-2结构域，以及一系列生化分析以评估多聚体的能力并绘制涉及多聚体的任何结构域的图谱；(B)使用高分辨率核磁共振波谱对先前定义的GT-2的孪生最小DNA结合结构域进行结构分析，以便开始了解这类新的转录因子识别序列特异性DNA的分子基础；以及(C)利用体外相互作用克隆和酵母双杂交系统筛选与GT-2相互作用的其他蛋白因子的cDNA文库，以探讨GT-2增强转录活性的机制。植物对光的反应是通过激活某些控制生长和分化的基因来实现的。这一机制涉及细胞中的一个光激活受体，它向特定的光激活基因发送化学信号，包括受体本身。一种与含有受体密码的DNA紧密结合的蛋白质已经被发现。这种蛋白质是植物所特有的，在两端都有DNA结合区。这种蛋白质通过与DNA结合以及与其他有助于产生信息的蛋白质结合来加速受体的产生。这项研究将确定蛋白质的哪些部分负责其加速信息产生的能力。此外，还将分析蛋白质的结构，以确定分子的每个片段是如何排列成分子蓝图的。***

9513590 Quail This proposal is focused on the previously cloned transcription factor, GT-2. The specific objectives are: (a) to perform further detailed analyses of the functional activities of GT-2, including transcriptional activation analysis using a transfection assay to define domains of GT-2 involved in transactivation, and a series of biochemical assays to assess capacity for, and map any domains involved in, multimerization; (b) to undertake a structural analysis of the previously defined twin minimal DNA-binding domains of GT-2 using high resolution nuclear magnetic resonance spectroscopy for the purpose of beginning to understand the molecular basis of sequence-specific DNA recognition by this novel class of transciption factor; and (c) to screen cDNA libraries for other protein factors that interact with GT-2 using in vitro interaction cloning and the yeast two hybrid system for the purpose of approaching the question of the mechanism by which GT-2 enhances transcriptional activity. %%% Plants respond to light by activating certain genes that govern growth and differentiation. The mechanism involves a light-activated receptor in the cell that sends a chemical signal to certain genes that are specifically light activated including the receptor itself. A protein that binds close to the DNA that contains the code for the receptor has been discovered. This protein is unique to plants and has DNA binding regions at both ends. This protein accelerates the production of the receptor by binding to the DNA and by binding to other proteins that help produce the message. The research will determine what parts of the protein are responsible for its ability to accelerate the production of the message. Also, the structure of the protein will be analyzed to determine how each segment of the molecule is arranged into a molecular blueprint. ***