Signalling Processes in Dictyostelium

盘基网柄菌的信号传导过程

• 批准号: 7988495
• 负责人: RICHARD A FIRTEL
• 金额: $ 9.74万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-12-17 至 2010-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7988495
• 关键词: Adaptor Signaling Protein; Adhesions; Affect; Back; Biochemical; Biochemical Genetics; Biological; Biological Assay; Biological Models; Biological Process; Cell Adhesion; Cell Polarity; Cell-Matrix Junction; Cells; Chemotactic Factors; Chemotaxis; Complex; Cytoskeleton; Defect; Development; Dictyostelium; Eukaryotic Cell; Evolution; Family; GTPase-Activating Proteins; Goals; Guanine Nucleotide Exchange Factors; Guanosine Triphosphate Phosphohydrolases; Immune system; Individual; Insertional Mutagenesis; Lateral; Ligand Binding; Ligands; Mediating; Methods; Monomeric GTP-Binding Proteins; Movement; Natural Immunity; Pathway interactions; Phosphorylation; Phosphotransferases; Physiological; Play; Positioning Attribute; Process; Protein Phosphatase 2A Regulatory Subunit PR53; Proteins; Reaction; Regulation; Reporter; Role; Side; Signal Pathway; Signal Transduction; Surface; System; Time; Translating; Work; base; blastomere structure; cancer cell; cell motility; cell type; computerized data processing; directional cell; extracellular; human disease; insight; macrophage; man; migration; myosin heavy-chain kinase A; neutrophil; null mutation; ras GTPase-Activating Proteins; ras Guanine Nucleotide Exchange Factors; ras Proteins; ras-Related G-Proteins; response; small molecule; yeast two hybrid system

DESCRIPTION (provided by applicant): Chemotaxis, or directed cell movement toward a small molecule ligand, plays a key role in many cellular and physiological responses, including metastasis of cancer cells, movement of neutrophils and macrophages in immune system reactions, migration of embryonic cells during development, and aggregation of Dictyostelium during development. In each of these varied cell types and processes, the responding cells are able to amplify a shallow extracellular chemoattractant gradient into a very steep intracellular gradient and thus translate the directional signal into directional cell movement. Fulfillment of this requirement occurs through an integrated circuit of signaling pathways that are highly conserved through evolution between Dictyostelium and man. Recent findings establish that the Ras and the related GTPase Rap1 are important for directional sensing, pseudopod formation, cell polarization, and cell attachment. Both Ras and Rap1 are preferentially activated at the leading edge of chemotaxing Dictyostelium cells and abrogation of their function impairs this process. This proposal focuses on the further examination of the roles of Ras and Rap1 of using Dictyostelium cells, which are amenable to biochemical, genetic, and cell biological approaches. We propose to identify the mechanisms by which the activations of Ras and Rap1 are regulated and spatially restricted to the leading edge of chemotaxing cells. This will be achieved through the analysis of a RasGEF complex and by examining defects in chemotaxis resulting from disruptions of specific GTPase activating proteins (GAPs) for Ras and for Rap1. Through the examination of the spatial and temporal regulation of activated Ras and Rap1 strains in which normal Ras and Rap1 activity is altered, we will elucidate the mechanisms by which cells orient themselves in a chemoattractant gradient. We propose to determine how Ras and Rap1 help mediate chemotaxis through the identification of downstream effectors. The function of these will be examined through the analysis of their null mutations with the aid of real-type fluorescent reporters and biochemical assays that reveal different components of directional sensing, pseudopod formation, and cell polarization. The work proposed in this application should provide new and important insights into mechanisms that control this highly evolutionarily conserved cell biological process, and thus provide the needed background to elucidate the cellular basis underlying a variety of human diseases, including those affecting innate immunity and metastasis of cancer cells.

描述（由申请人提供）：趋化性或定向细胞向小分子配体的运动在许多细胞和生理反应中起着关键作用，包括癌细胞的转移、免疫系统反应中中性粒细胞和巨噬细胞的运动、发育过程中胚胎细胞的迁移以及发育过程中盘基网柄菌的聚集。在这些不同的细胞类型和过程中，响应细胞能够将浅的细胞外趋化剂梯度放大为非常陡的细胞内梯度，从而将定向信号转化为定向细胞运动。这一要求的实现是通过信号通路的集成电路实现的，这些信号通路在盘基网柄菌和人类之间的进化过程中高度保守。最近的研究结果表明，Ras 和相关的 GTPase Rap1 对于定向传感、伪足形成、细胞极化和细胞附着非常重要。 Ras 和 Rap1 均优先在趋化盘基网柄菌细胞的前缘被激活，并且其功能的废除会损害该过程。 该提案的重点是进一步检查 Ras 和 Rap1 在使用盘基网柄菌细胞中的作用，这些细胞适合生化、遗传和细胞生物学方法。我们建议确定 Ras 和 Rap1 的激活被调节并在空间上限制在趋化细胞前缘的机制。这将通过分析 RasGEF 复合物并检查因 Ras 和 Rap1 的特定 GTP 酶激活蛋白 (GAP) 破坏而导致的趋化缺陷来实现。通过检查激活的 Ras 和 Rap1 菌株的空间和时间调节（其中正常 Ras 和 Rap1 活性发生改变），我们将阐明细胞在趋化梯度中定向自身的机制。我们建议通过鉴定下游效应子来确定 Ras 和 Rap1 如何帮助介导趋化性。这些的功能将通过借助真实型荧光报告基因和生化分析来分析它们的无效突变来检查，这些分析揭示了方向传感、伪足形成和细胞极化的不同组成部分。本申请中提出的工作应该为控制这种高度进化保守的细胞生物过程的机制提供新的重要见解，从而提供所需的背景来阐明各种人类疾病的细胞基础，包括影响先天免疫和癌细胞转移的疾病。