Signalling Processes in Dictyostelium

描述（由申请人提供）：趋化性，或指向小分子配体的细胞运动，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，胚胎细胞在发育过程中的迁移，以及发育过程中盘基骨柱的聚集。在这些不同的细胞类型和过程中，响应细胞能够将细胞外的浅化学引诱梯度放大到非常陡峭的细胞内梯度，从而将定向信号转化为定向细胞运动。这一需求的实现是通过一个信号通路的集成电路来实现的，这个信号通路在盘基骨龙和人类之间的进化过程中是高度保守的。最近的研究发现，Ras和相关的GTPase Rap1对定向传感、假足形成、细胞极化和细胞附着都很重要。Ras和Rap1都是在盘基骨柱细胞的化学代谢前沿被优先激活的，取消它们的功能会损害这一过程。

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.