ANALYSIS OF SIGNALLING PROCESSES IN DICTYOSTELIUM

细胞黏菌盘基网柄菌的发育 部分由与细胞相互作用的细胞外分子调节 表面受体 本申请旨在理解 信号转导的分子和生化机制 细胞周期中的细胞形态发生和基因表达 聚合前和聚合发展阶段。使用 分子技术，我们已经确定和克隆的基因，其产品 是控制这个和其他发展阶段的关键。这些 包括编码四个G α蛋白亚基的基因，每个亚基具有 独特的表达发展模式，一种发育调节的 磷酸酪氨酸磷酸酶和发育调节的推定的 丝氨酸/苏氨酸激酶。 基因破坏和过表达研究 已经表明这些基因编码的蛋白质 在发展中的重要作用。 我们的分析表明G α 2 与cAMP受体cARI偶联，调节多种信号 在聚集过程中介导趋化性的转导途径， 脉冲诱导基因的诱导。 在这个应用中，我们建议剖析信号转导 这些蛋白质调控的途径， 产生不表达蛋白质或 表达修饰的蛋白质。 体外分析将用于 在生化水平上描述突变的影响。 使用 分子互补，我们建议分离额外的基因， 先前已经通过体内实验确定的信号通路 突变。 我们还将继续研究 细胞类型特异性ras基因的信号转导途径。 在 为了了解受体介导的控制的分子基础， 在聚集过程中的基因表达，我们计划进一步确定 由cAMP脉冲调节的基因的顺式作用调节区， 纯化调节反式作用因子，介导反应， 转录水平。

Development in the cellular slime mold Dictyostelium discoideum is regulated in part by extracellular molecules that interact with cell surface receptors. This application is directed at understanding the molecular and biochemical mechanisms by which signal transduction pathways control both cellular morphogenesis and gene expression during the pre-aggregation and aggregation stages of development. Using molecular techniques, we have identified and cloned genes whose products are essential in controlling this and other stages of development. These include genes encoding four G alpha protein subunits, each with a distinct developmental pattern of expression, a developmentally regulated phosphotyrosine phosphatase, and developmentally regulated putative serine/threonine kinases. Gene disruptions and overexpression studies have indicated that the proteins encoded by each of these genes play essential roles in development. Our analysis indicates that G alpha2 couples to the cAMP receptor cARI and regulates diverse signal transduction pathways that mediate chemotaxis during aggregation and the induction of pulse-induced genes. In this application, we propose to dissect the signal transduction pathways regulated by these proteins using molecular techniques to produce appropriate strains that either do not express the proteins or express modified proteins. In vitro analysis will be used to characterize the effects of the mutations at a biochemical level. Using molecular complementation, we propose to isolate additional genes in the signaling pathways that have been previously identified by in vivo mutations. We will also continue to characterize the function of the cell-type-specific ras gene within the signal transduction pathway. In order to understand the molecular basis of receptor-mediated control of gene expression during aggregation, we plan to further identify the cis-acting regulatory regions of genes regulated by cAMP pulses and to purify the regulatory trans-acting factors that mediate the response at the level of transcription.