TRANS-ACTING FACTORS REGULATING ADH GENE EXPRESSION

这项建议的总体目标是深入了解 DNA结构、反式作用因子、蛋白质之间的关系 核酸相互作用与基因调控 植物中的转录。实验涉及玉米 酒精脱氢酶基因。ADHL和ADH2表现出差异 组织特异性，但厌氧调节坐标 两种ADH基因的转录表达。这提供了一种 评估变装者所扮演的角色的独特机会 激活中的因素和蛋白质-核酸相互作用 两个密切相关的基因，并比较和对比 基因转录的发育和环境激活。 这个相当直白的提案的主要关注点是In 反式作用蛋白的体外鉴定和纯化 影响ADH基因调控的蛋白质因素。 虽然这一总体研究领域在#年一直在积极进行 动物系统、植物的反式作用因素才刚刚开始 接受调查。这项提议有什么特别之处？ 可行和适当的是我们已经知道(至少有一些 参与ADH调控的因素。从我们体内的 足迹研究我们知道至少有四个结合部位 监管因素，我们知道他们的硫酸二甲酯足迹 签名。因此，我们确切地知道我们是什么因素 搜索纯化，而不是简单地寻找蛋白质 (只是)与ADH DNA结合。

The overall goal of this proposal is to gain insight into the relationships among DNA structure, trans-acting factors, protein- nucleic acid interactions, and the regulation of gene transcription in plants. The experiments involve the maize alcohol dehydrogenase genes. Adhl and Adh2 exhibit differential tissue specificity, yet anaerobiosis mediates the coordinate transcriptional expression of both Adh genes. This provides a unique opportunity to evaluate the roles played by trans-acting factors and protein-nucleic acid interactions in the activation of two closely related genes, and to compare and contrast the developmental and environmental activation of gene transcription. The main focus of this rather straightforward proposal is the in vitro characterization and purification of the trans-acting protein factors that affect the regulation of the Adh genes. While this general area of research has been actively pursued in animal systems, plant trans-acting factors are only now beginning to be investigated. What makes this proposal particularly feasible and appropriate is that we already know (at least some of) the factors involved in Adh regulation. From our in vivo footprinting studies we know the binding sites of at least four regulatory factors, and we know their dimethyl sulfate footprint signatures. Therefore we know precisely the factors we are searching to purify and are not simply looking for proteins that (just) bind to Adh DNA.