The Role of Two Novel cGMP Binding Proteins in Dictyostelium

趋化性是指细胞沿化学梯度向上移动。在盘基网柄藻中，趋化性在生长条件下用来寻找细菌作为食物来源，在饥饿条件下用来促进细胞聚集，从而形成子实体。细胞检测趋化物质的基本机制在Dictyostelius和哺乳动物细胞之间是保守的，驱动细胞迁移的细胞机制也是保守的。梯度上移涉及刺激后第一分钟细胞骨架的不对称重塑，因此需要在时间和空间上严格调控关键细胞骨架成分的活动和定位。该项目的长期目标是了解在趋化等过程中控制细胞骨架重塑的信号通路。网柄苔藓的实验可控性，再加上它的相对简单，使它成为一种可以详细研究这些基本信号转导途径的极好的模式生物。一般的策略是重点观察cGMP的胞内峰的形成对于Dictyostelius细胞的趋化是必不可少的。假说是细胞内关键的cGMP结合蛋白直接或间接激活化学诱导剂处理后的细胞骨架成分。尽管cGMP在趋化作用中的信号转导中具有重要作用，但到目前为止，还没有在网柄基菌中纯化或克隆cGMP结合蛋白。两个候选的cGMP结合蛋白最近通过分析Dictyostants基因组数据库的数据而被鉴定出来。这些cGMP结合蛋白的单基因和双基因中断菌株将通过同源重组得到工程，而它们在趋化作用中发挥核心作用的假设将通过拍摄细胞对充满趋化物质的微管的反应来检验。鉴定网柄苔藓中cGMP结合蛋白并确定它们在趋化作用中的作用，将是理解这一过程中信号转导机制的重大进展。该项目还为本科生提供了参与自主研究的机会。

Chemotaxis is the movement of cells up a chemical gradient. In Dictyostelium discoideum, chemotaxis is used under growth conditions to find bacteria as a food source, and under starvation conditions to promote aggregation of cells so that fruiting bodies may be formed. The basic mechanisms by which cells detect chemoattractants are conserved between Dictyostelium and mammalian cells, and the cellular machinery that drives cell migration is also conserved. Migration up a gradient involves the asymmetric remodeling of the cytoskeleton during the first minute after stimulation, and therefore requires that the activities and localization of key cytoskeletal components be tightly regulated in time and space. The long-term goal of this project is to understand the signaling pathways that control remodeling of the cytoskeleton during processes such as chemotaxis. The experimental tractability of Dictyostelium, coupled with its relative simplicity, make it an excellent model organism in which these fundamental signal transduction pathways can be worked out in detail. The general strategy is to focus on the observation that formation of an intracellular peak of cGMP is essential for chemotaxis in Dictyostelium cells. The hypothesis is that key cGMP-binding proteins in the cell directly or indirectly activate cytoskeletal components after treatment with chemoattractants. Despite the well-established importance of cGMP in signaling in chemotaxis, to date no cGMP-binding proteins have been purified or cloned in Dictyostelium. Two candidate cGMP-binding proteins have recently been identified by analyzing data from the Dictyostelium genomic database. Single and double gene disruption strains for these cGMP binding proteins will be engineered by homologous recombination, and the hypothesis that they play a central role in chemotaxis will be tested by filming cells as they respond to a micropipet filled with chemoattractant. The identification of cGMP binding proteins in Dictyostelium and determination of their role in chemotaxis will be a major advance in understanding how signaling during this process works. The project also provides the opportunity for undergraduate students to participate in independent research.