MOLECULAR DESIGN OF FILOPODIA, CELL'S SENSORY ORGANELLES

描述(由申请人提供)：这项提案的广泛目标是了解细胞如何以有意义的方式相互交流以及与环境进行交流。为了找到黏附的目标表面或最终的迁移目的地，细胞应该能够探索环境。丝状伪足是细长的细胞突起，被认为是细胞的感官和引导细胞器。它们可以识别环境线索，检测合适的表面，并确定细胞运动的方向。丝状伪足对于正常组织细胞的导航和癌细胞的转移特别重要，但对于特殊结构的初始化也特别重要，例如突触、连接和通讯路径。本项目将致力于研究丝状伪足在其形成、突起和分化为特化结构过程中动态行为的结构基础。其具体目的是通过分析不同细胞类型在不同条件和不同目的下形成的丝状足细胞之间的结构和动态变化，在深度和广度上深入研究丝状细胞骨架与质膜的相互作用。我们将结合结构、动力学、功能和分子遗传学的方法来研究丝状结构的中心成员IRSP53和mDia2在细胞骨架和质膜之间的界面上的作用。需要研究的丝状伪足相关结构包括控制细胞运动的前沿丝状伪足、在脑内建立突触并分化为树突棘的树状丝状伪足，以及参与内皮通透性屏障形成的连接性丝状伪足。该项目的基本假设是，在丝状伪足突起过程中，两种分子机制--肌动蛋白细胞骨架组装和质膜动力学--紧密合作，这种平衡可以在不同方向上调节，以产生各种服务于不同功能的丝状伪足结构。我们研究策略的一个关键要素是通过铂复制电子显微镜获得高分辨率的结构信息，并将其与先进光学显微镜记录的同一细胞的动态行为相关联。当与现代功能方法相结合时，这种策略具有独特的能力，通过在亚细胞和超分子水平上提供桥接信息来填补单个分子的性质和细胞行为之间的现有差距。 公共卫生相关性：这些结果将有助于理解丝状伪足形成的分子机制，以及丝状伪足在细胞迁移、细胞间通讯和组织形态发生等发育和疾病基本过程中的作用。这些信息将有助于设计药物、治疗和诊断工具。

DESCRIPTION (provided by applicant): The broad objective of this proposal is to understand how cells communicate with each other and with the environment in a meaningful way. To find a target surface for adhesion or a final destination for migration, a cell should be able to explore the environment. Filopodia are long slender cellular protrusions which are believed to be the cell's sensory and guiding organelles. They can recognize environmental cues, detect appropriate surfaces and determine the direction of cell locomotion. Filopodia are especially important for navigation of normal tissue cells and for metastasis of cancer cells, but also for initialization of specialized structures, such as synapses, junctions, and communication pathways. This project will focus on studying the structural basis of the dynamic behavior of filopodia during their formation, protrusion, and differentiation into specialized structures. The specific aims are designed to investigate the filopodial machinery in depth with a major focus on interaction of the filopodial cytoskeleton with the plasma membrane, and in breadth, by analyzing the structural and dynamic variations among filopodia formed by different cell types in different conditions and for different purposes. Roles of central players of filopodial machinery, IRSp53 and mDia2, functioning at the interface between the cytoskeleton and plasma membrane will be investigated by combination of structural, kinetic, functional and molecular genetics approaches. The range of filopodia-related structures to be investigated includes leading edge filopodia that control cell motility, dendritic filopodia that establish synapses in brain and differentiate into dendritic spines, and junctional filopodia that are involved in formation of permeability barrier in endothelium. The underlying hypothesis for this project suggests that two molecular machineries, actin cytoskeleton assembly and plasma membrane dynamics, tightly cooperate with each other during filopodia protrusion and this balance can be tuned in different directions to produce a variety of filopodia-based structures serving different functions. A key element of our research strategy is to obtain high resolution structural information by platinum replica electron microscopy and correlate it with the dynamic behavior of the same cell recorded by advanced light microscopy. When additionally combined with modern functional approaches, this strategy has a unique ability to fill the existing gap between properties of individual molecules and behavior of a cell by providing bridging information at subcellular and supramolecular levels. PUBLIC HEALTH RELEVANCE: The results will contribute to understanding of the molecular mechanisms of filopodia formation and the roles of filopodia in such fundamental processes in development and disease as cell migration, cell-cell communication, and tissue morphogenesis. This information will help to design drugs, treatments and diagnostic tools.