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MECHANISM OF EUCARYOTIC GENE REGULATION

真核基因调控机制

基本信息

批准号：
3281387
负责人：
KEITH A BOSTIAN
金额：
$ 15.22万
依托单位：
BROWN UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1984
资助国家：
美国
起止时间：
1984-04-01 至 1989-03-31
项目状态：
已结题

项目摘要

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.

我们试图阐明基本的分子机制磷代谢的调节机制酿酒酵母我们的方法是协调一致的利用生物化学和遗传学技术来研究酸性磷酸酶（AP酶）基因的转录控制。我们预计这些研究的结果将提供新的深入了解与基因相关的各种相互关联的方面，调节高等真核生物，并将进一步发展我们的了解磷系统的分子生物学，特别是关于调节基因肌动蛋白的机制，和转录调控。酵母中的磷代谢系统由分散的基因家族的积极和消极的调节许多参与磷代谢的结构基因。前几我们克隆了APase的表达并对其进行了表征，基因（PHO5，PH03，PHO10和PH011），该家族的成员。 PH05和PH03基因紧密连锁为串联重复，并受到严格的调控：PH05表达仅在低Pi生长的细胞中，并且仅当PH05被抑制时PH03。我们已经开始在物理上隔离和描述调控基因及其产物，遗传学策略 PH 05、PH 04和PH 06的正调节剂 PH02，已被克隆，并鉴定了它们的产物，接受调查。 PH081、PH085和PH080 调控基因目前正在分离和分析，类似的方法。我们计划继续进行这些研究，几个具体问题。我们将评估结构， PH04蛋白质结构域的功能，通过位点特异性诱变这将包括识别负责转录控制的成分，在PH04 我们还将定义交互的组件，通过分离和表征参与PH05调节 UAS调节性交配的第二位点抑制因子。这将包括化学合成最小的UAS序列，在序列中引入碱基取代和缺失。最后，通过对PH05和PH03进行元基因分析，将分析PH 05介导的PH 03的相互调节，基因识别和表征其他反式作用 PH 03涉及监管因素。