Signal Integration in Neutrophil Chemotaxis

描述（由申请人提供）：定向细胞迁移是单细胞生物狩猎和交配所必需的，使先天免疫细胞能够寻找和摧毁病原体，对多细胞生物的形态发生至关重要。细胞迁移的错误调控与动脉粥样硬化和心脏发育缺陷密切相关。虽然我们开始了解细胞迁移的一些关键组成部分，但我们不了解这些组成部分如何共同作用来组织细胞的形状和运动。为了解决这个问题，我们分析了一个关键的肌动蛋白调节因子Scar/WAVE复合物的空间动力学，这是后生动物和植物形态发生所必需的。我们最近发现，Scar/WAVE复合体的Hem-1成分定位于似乎组织运动免疫细胞（人类中性粒细胞）前沿的传播波。奇怪的是，肌动蛋白既是Scar/WAVE复合物的输出又是输入：复合物刺激肌动蛋白组装，肌动蛋白聚合物也需要将复合物从膜上移除。这些相互作用似乎产生肌动蛋白成核的传播波，体现了许多运动细胞形态发生的特性，如细胞绕屏障流动的能力和前沿突起的复杂空间组织。我们的中心假设是，在细胞迁移过程中，Hem-1波发生器和其他信号信号之间的相互作用在空间上组织了肌动蛋白聚合。在这一建议中，我们将剖析组织Hem-1波动力学的信号，并研究它们与细胞形态发生和定向运动的关系。具体而言，我们将：

DESCRIPTION (provided by applicant): Directed cell migration is required for single-celled organisms to hunt and mate, enables innate immune cells to seek and destroy pathogens, and is essential for the morphogenesis of multicellular organisms. Misregulation of cell migration is intimately involved in atherosclerosis and defective cardiac development. Though we are beginning to understand some of the key components involved in cell migration, we do not understand how these components act together to organize cell shape and movement. To address this question, we have analyzed the spatial dynamics of a key actin regulator the Scar/WAVE complex, which is required for morphogenesis in both metazoans and plants. We have recently discovered that the Hem-1 component of the Scar/WAVE complex localizes to propagating waves that appear to organize the leading edge of a motile immune cell, the human neutrophil. Curiously, actin is both an output and input to the Scar/WAVE complex: the complex stimulates actin assembly, and actin polymer is also required to remove the complex from the membrane. These reciprocal interactions appear to generate propagated waves of actin nucleation that embody many of the properties of morphogenesis in motile cells such as the ability of cells to flow around barriers and the intricate spatial organization of protrusion at the leading edge. Our central hypothesis is that the interaction between the Hem-1 wave generator and other signaling cues spatially organizes actin polymerization during cell migration. In this proposal, we will dissect the signals that organize Hem-1 wave dynamics and study their relationship to cell morphogenesis and directed motility. Specifically, we will: Quantitate the effect of external gradients on Hem-1 wave dynamics. We will quantitatively analyze Hem-1 wave dynamics during chemotaxis to test two competing hypotheses in the field-- whether generation of new protrusions or selection among existing ones is responsible for directional migration. 2. Dissect the reciprocal interactions between Rac and Hem-1. We are using both micropatterning and small molecule dimerizers to control the spatial and temporal dynamics of Rac and Hem-1 localization in living cells to dissect how these signals interact with one another. 3. Elucidate the role of the actin cytoskeleton in Hem-1 wave propagation. We will use a combination of actin perturbing drugs and targeted mislocalization of actin nucleation factors to investigate how actin polymer interfaces with Hem-1 wave dynamics. 4. Test role of Hem-1 in front/back crosstalk. We are using microfluidics-based drug perfusion and small-molecule based dimerization to spatially manipulate the signals involved in front (Rac/Hem-1) and back (Rho/myosin) organization to determine how these regulators of polarity communicate. PUBLIC HEALTH REVELANCE: Misregulation of actin polymerization and leukocyte migration are causative factors in heart disease. Leukocytes play a central role in atherosclerosis, and the motility circuit that we study is essential for angiogenesis and cardiovascular development. The ability to control cell migration would be a valuable tool for combating atherosclerosis and other pathological processes that occur upon disruption of cellular guidance mechanisms.