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于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，对于伤口修复至关重要。在单个细胞的水平上，形状变化的周期允许某些细胞类型在胚胎发生、免疫功能和（更隐蔽地）转移过程中迁移。细胞形态由细胞骨架（肌动蛋白丝和微管网络）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学的原理和机制。重要的是，肌动蛋白和微管的网络并不是孤立地起作用，而是存在前所未有的串扰程度，包括调节相互作用和机械相互作用。微管加末端跟踪蛋白（或 TIP）是一类选择性定位于生长和收缩微管尖端的分子。自 1999 年发现以来，TIP 与几乎所有微管依赖性细胞功能有关，包括微管动态不稳定性的调节、细胞器和染色体运输、有丝分裂纺锤体的组装、细胞极性的建立和细胞迁移。在本提案中，我们重点关注 TIP，特别强调肌动蛋白-微管串扰，因为这代表了两个细胞骨架网络之间相对未经探索的功能界面。我们的核心假设是，微管两端是动态平台，可将信息传递到控制细胞形状的皮质调节网络，并充当肌动蛋白和微管之间的结构整合位点。我们将使用我们用培养的果蝇细胞系开发的新测定法来测试这些想法，因为该模型系统适合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，这些细胞成分在形态发生和细胞迁移等过程中介导细胞形状的变化。该提案的目标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于胚胎发育至关重要 - 如果它们在发育过程中不同步或根本无法发生，则可能导致先天性出生缺陷。同样，细胞运动是伤口愈合和免疫反应等过程的基础。细胞运动不当也是动脉粥样硬化、炎症和转移的根本原因。

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.