Mechanisms of cytoskeletal crosstalk during cellular motility

描述(由申请人提供)：细胞改变其形状的能力对大多数生物体的个体发育至关重要。例如，在组织内，细胞形态的变化在形态发生过程中驱动组织重塑，并对伤口修复至关重要。在单个细胞的水平上，形状变化的周期允许某些类型的细胞在胚胎发育、免疫功能和(更隐蔽的)转移期间迁移。细胞形态由细胞骨架--肌动蛋白细丝和微管组成的网络--决定。该项目的长期目标是通过研究调节和整合细胞骨架动力学的途径，在分子水平上了解细胞形态的基本原理和机制。重要的是，肌动蛋白和微管网络并不是孤立地发挥作用，而是存在着前所未有的串扰，既有调节作用，也有机械作用。微管加末端跟踪蛋白(或TIPS)是一类选择性地定位于生长和收缩的微管末端的分子。自1999年发现以来，TIPS几乎参与了微管依赖的所有细胞功能，包括微管动态不稳定性的调节、细胞器和染色体的运输、有丝分裂纺锤体的组装、细胞极性的建立和细胞迁移。在这项提案中，我们将重点放在TIPS上，特别强调肌动蛋白-微管的串扰，因为这代表了两个细胞骨架网络之间相对未被探索的功能接口。我们的核心假设是，微管+末端是动态的平台，向控制细胞形状的皮质调控网络传递信息，也是肌动蛋白和微管之间结构整合的位置。我们将使用我们开发的新的分析方法来测试这些想法，因为这个模型系统适用于高分辨率光学显微镜、生化分析和使用RNAi的基因抑制。这些研究的结果将有助于对细胞成分网络的基本了解，这些细胞成分网络在形态发生和细胞迁移等过程中调节细胞形状的变化。这项建议的目的是了解细胞形态发生和运动的机制基础。细胞形状变化的正确执行对胚胎发育至关重要--如果它们在发育过程中不同步，或者根本没有发生，这可能会导致先天性出生缺陷。像WISE一样，细胞运动是伤口愈合和免疫反应等过程的基础。不适当的细胞运动也是动脉粥样硬化、炎症和转移的潜在原因。

DESCRIPTION (provided by applicant): The ability of cells to alter their shape is critical to the ontogeny of most organisms. Within tissues, for example, changes in cellular morphology drive tissue remodeling during morphogenesis and are essential for wound repair. At the level of the individual cell, cycles of shape change allow some cell types to migrate during embryogenesis, immune function, and (more insidiously) during metastasis. Cellular morphology is dictated by the cytoskeleton - the network of actin filaments and microtubules. The long-term goal of this project is to understand the principles and mechanisms underlying cellular morphology at the molecular level by studying the pathways that regulate and integrate cytoskeletal dynamics. Importantly, the networks of actin and microtubules do not act in isolation, rather there is an unprecedented degree of cross-talk, both regulatory interactions and mechanical interactions. Microtubule plus end-tracking proteins (or +TIPs) are a class of molecules that selectively localize to the tips of growing and shrinking microtubules. Since their discovery in 1999, +TIPs have been implicated in almost every microtubule-dependent cellular function including regulation of microtubule dynamic instability, organelle and chromosomal transport, assembly of the mitotic spindle, establishment of cellular polarity, and cell migration. In this proposal, we focus on +TIPs with a particular emphasis on actin-microtubule cross-talk as this represents a relatively unexplored functional interface between the two cytoskeletal networks. Our core hypothesis is that microtubule plus ends are dynamic platforms that deliver information to cortical regulatory networks governing cell shape and also act as sites of structural integration between actin and microtubules. We will test these ideas using novel assays we have developed with cultured Drosophila cell lines as this model system is amenable to high-resolution light microscopy, biochemical analyses, and gene inhibition using RNAi. The results of these studies will contribute to a basic understanding about the network of cellular components that mediate changes in cellular shape during processes such as morphogenesis and cell migration. The goal of this proposal is to understand the mechanistic basis of cellular morphogenesis and motility. The proper execution of cellular shape changes is essential for embryonic development - if they are not synchronized during development, or fail to occur at all, this can result in congenital birth defects. Like wise, cellular motility underlies processes such as wound healing and immune response. Improper cell motility is also an underlying cause of atherosclerosis, inflammation, and metastasis.