MECHANISM OF EUCARYOTIC GENE REGULATION

我们正试图阐明基本的分子机制 植物体内磷代谢的调控 酿酒酵母。我们的方法是坐标方法。 利用生化和基因技术来研究 酸性磷酸酶(APase)基因转录调控。我们 预计这些研究的结果将提供新的 对与基因相关的各种相互关联的方面的见解 对高等真核生物的调控，并将进一步发展我们的 了解磷系统的分子生物学， 尤其是关于调控基因肌动蛋白的机制 和转录调控。 酵母中的磷代谢系统由一个分散的 众多正负调控因子的基因家族 参与磷代谢的结构基因。在以前的 我们克隆并鉴定了APase的表达 基因(PHO5、PH03、PHO10和PH011)，这个家族的成员。 Ph05和ph03基因紧密相连，形成串联 复制，并相互调控：PH05是表达的 只有在低等电点生长的细胞中才有，只有在PH05被抑制时才有ph03。 我们已经开始在物理上孤立并描述 调控基因及其产物通过重组DNA和 遗传学策略。PH05、PH04和 Ph02，已被克隆，并鉴定了它们的产物 受到调查。PH081、PH085和PH080 目前正在分离和分析调控基因 类似的方法。我们计划通过以下方式继续进行这些研究 几个具体的问题。我们将评估该结构并 PH04蛋白结构域在位点特异性中的作用 诱变。这将包括识别蜂窝 负责相互作用的转录控制的组件 使用PH04。我们还将定义交互组件和 通过分离和鉴定参与PH05调节 无人驾驶飞机的第二位抑制因子。这将是 包括化学合成最小的UAS序列，然后 在序列中引入碱基替换和缺失。 最后，通过对PH05和PH03进行元生成分析，我们 将分析PH05介导的PH03的相互调节， 从基因上鉴定和表征其他反式作用 Ph03涉及的调节因素。

We are attempting to elucidate basic molecular mechanisms underlying the regulation of phosphorus metabolism in Saccharomyces cerevisiae. Our approach is a coordinate one utilizing both biochemical and genetic techniques to study the transcriptional control of the acid phosphatase (APase) genes. We anticipate that results from these studies will provide new insights into a variety of interrelated aspects of relevance to gene regulation in higher eucaryotes, and will furher develop our understanding of the molecular biology of the phosphorus system, particularly with respect to mechanisms of regulatory gene actin and transcriptional regulation. The phosphorus metabolism system in yeast consists of a dispersed gene family of positive and negative regulators of numerous structural genes involved in phosphorus metabolism. In previous studies we cloned and characterized the expression of the APase genes (PHO5, PH03, PHO10, and PH011), members of this family. The PH05 and PH03 genes are tightly linked as a tandem duplication, and are reciprocally regulated: PH05 is expressed only in low-Pi grown cells, and PH03 only when PH05 is repressed. We have begun to physically isolated and characterize the regulatory genes and their products by recombinant DNA and genetics strategies. Positive regulators of PH05, PH04, and PH02, have been cloned and their products identified and subjected to investigation. The PH081, PH085, and PH080 regulatory genes are currently being isolated and analyzed by similar methods. We plan to continue these studies by addressing several specific issues. We will evaluate the structure and function of the protien domains of PH04 by site-specific mutagenesis. This will include identificatin of the cellular components responsible for transcriptional control which interact with PH04. We will also define components that interact and participate in PH05 regulation by isolation and characterization of second-site suppressors of UAS rgulatory matations. This will involve chemically synthesizing a minimal UAS sequence and then introducing base substitution and deletions into the sequence. Finally, by subjecting PH05 and PH03 to metagenic analysis, we will analyze the PH05 mediated reciprocal regulation of PH03, genetically identifying and characterizing other trans-acting PH03 regulators factors involved.