SIGNALING PROCESSES IN DICTYOSTELIUM

盘基网菌中的信号传导过程

• 批准号: 2178999
• 负责人: RICHARD A FIRTEL
• 金额: $ 42.7万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-07-01 至 1995-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2178999
• 关键词: DNA footprinting; Dictyostelium; Escherichia coli; G protein; adenylate cyclase; affinity chromatography; antibody; biological signal transduction; cell aggregation; cell differentiation; cell motility; cell transformation; chemotaxis; chimeric proteins; cyclic AMP; cyclic AMP receptors; developmental genetics; electroporation; epitope mapping; fusion gene; gene expression; gene mutation; genetic library; genetic regulatory element; genetic transcription; guanylate cyclase; histogenesis; hydrolase; immunofluorescence technique; laboratory mouse; laboratory rabbit; microorganism genetics; molecular cloning; molecular genetics; oncogenes; phospholipase C; phosphorylation; phosphotransferases; posttranslational modifications; protein purification; protein structure function; temperature sensitive mutant; transcription factor; western blottings

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.

细胞粘菌盘基网柄菌的发育是 部分受到与细胞相互作用的细胞外分子的调节 表面受体。 该应用程序旨在了解 信号转导的分子和生化机制 途径控制细胞形态发生和基因表达 开发的预聚合和聚合阶段。使用 分子技术，我们已经鉴定并克隆了其产物的基因 对于控制这个阶段和其他发展阶段至关重要。这些 包括编码四个 G α 蛋白亚基的基因，每个亚基都有一个 独特的表达发育模式，发育调节 磷酸酪氨酸磷酸酶，以及发育调节的假定 丝氨酸/苏氨酸激酶。 基因破坏和过度表达研究 已经表明这些基因编码的蛋白质发挥作用 发展中的重要作用。 我们的分析表明 G α2 与 cAMP 受体 cARI 偶联并调节多种信号 聚集过程中介导趋化性的转导途径 脉冲诱导基因的诱导。 在此应用中，我们建议剖析信号转导 使用分子技术通过这些蛋白质调节的途径 产生不表达蛋白质或不表达蛋白质的适当菌株 表达修饰的蛋白质。 体外分析将用于 在生化水平上表征突变的影响。 使用 分子互补，我们建议分离额外的基因 先前已通过体内鉴定的信号通路 突变。 我们还将继续描述其功能 信号转导途径内的细胞类型特异性 ras 基因。 在 为了了解受体介导的控制的分子基础 聚集过程中的基因表达，我们计划进一步鉴定 由 cAMP 脉冲调节的基因的顺式作用调节区 纯化介导反应的监管反式作用因子 转录水平。