Cell adhesion, signal transduction and cytoskeletal regulation in Drosophila

描述（由申请人提供）：细胞粘附和细胞骨架调节的协调对胚胎发生期间的组织和器官组装至关重要，也对成人的伤口修复和组织稳态至关重要。细胞粘附的破坏是肿瘤转移的关键步骤，并在皮肤和心脏疾病中发挥作用。我们开发了一个模型系统来研究细胞粘附和肌动蛋白细胞骨架的调节耦联，使用果蝇。该系统中可用的工具使我们能够将非常强大的遗传方法与在完整动物的背景下研究细胞生物学事件的能力结合起来，通常是实时的。许多实验室的工作提供了一个静态的“教科书”模型，说明核心钙粘蛋白：粘附连接处的连环蛋白复合物（AJs）如何介导粘附并将粘附连接连接到肌动蛋白。然而，胚胎和成人的细胞远不是静止的。此外，教科书模式的某些关键特征也受到了质疑。我们目前的挑战是修改这个模型，揭示粘附和细胞骨架调节是如何协调和调节的，从而使发育中的胚胎中发现的显著多样化的细胞行为成为可能。在这里，我们讨论这一领域的两个广泛的未解决的问题，每个问题都为我们的一个具体目标提供了基础。目的1：明确AJs与肌动蛋白细胞骨架连接的机制，并探讨这些机制如何影响顶端收缩、AJs重塑和其他过程。规范模型表明AJs和肌动蛋白之间存在直接的机械联系，由钙粘蛋白之间的相互作用介导。连环蛋白,? ?连环蛋白和肌动蛋白。然而，最近的研究对这种说法提出了质疑。确定AJs是否以及如何与肌动蛋白细胞骨架相连是我们这个领域的一个关键问题。我们假设一个直接的机械连接是关键的坚固细胞形状的变化，如那些顶端收缩。基于我们的初步数据，我们进一步假设独木舟和Rap1有助于调解这种联系。我们将验证这一假设，研究Cno和Rap1的作用机制，并探索它们是否在根尖收缩和AJ重塑事件中发挥更普遍的作用。目的2定义粘附和肌动蛋白动力学协调调节的机制。为了完成复杂的细胞形态发生行为，细胞必须密切协调黏附和肌动蛋白动力学。定义这种情况发生的机制是该领域的一项关键任务。我们确定了酪氨酸激酶Abl和肌动蛋白调节因子Ena在形态发生中的关键作用。我们使用它们作为理解协调细胞粘附和细胞骨架调节机制的模型。我们假设像Ena这样的肌动蛋白调节因子在AJs中处于非活性状态。我们假设Abl通过调节Ena的定位和/或功能来调节AJs及附近的肌动蛋白组装，作为支架，直接影响肌动蛋白。我们假设Ena通过不同的结构域和伙伴帮助产生不同的肌动蛋白结构，并且Ena与丝状足中的其他肌动蛋白调节因子相结合。我们将检验这些假设。

DESCRIPTION (provided by applicant): The coordination of cell adhesion and cytoskeletal regulation is critical for tissue and organ assembly during embryogenesis, and also during wound repair and tissue homeostasis in adults. Disruption of cell adhesion is a critical step in tumor metastasis, and plays a role in skin and heart diseases. We have developed a model system to study coupling of cell adhesion and regulation of the actin cytoskeleton, using the fruit fly Drosophila. Tools available in this system allow us to combine very powerful genetic approaches with the ability to study cell biological events in the context of intact animals, often in real time. Work in many labs provided a static "textbook" model for how the core cadherin:catenin complex at adherens junctions (AJs) mediates adhesion and links adhesive junctions to actin. However, cells in embryos and adults are far from static. Further, certain key features of the textbook model have been called into question. Our current challenge is to revise this model, revealing how adhesion and cytoskeletal regulation are coordinated and regulated to enable the remarkably diverse cell behaviors found in developing embryos. Here we address two broad unanswered questions in this area, each providing the basis for one of our Specific Aims. Aim 1 Define mechanisms by which AJs are connected to the actin cytoskeleton and explore how these affect apical constriction, AJ remodeling and other processes. The canonical model suggests that there is a direct mechanical connection between AJs and actin, mediated by linked interactions between cadherins, ?-catenin, ??catenin, and actin. However, recent work called this into question. Defining whether and how AJs are linked to the actin cytoskeleton is a key question for our field. We hypothesize that a direct mechanical connection is critical in robust cell shape changes like those of apical constriction. Based on our preliminary data, we further hypothesize that Canoe and Rap1 help mediate this link. We will test this hypothesis, examining mechanisms by which Cno and Rap1 act, and exploring whether they play a more general role in apical constriction and AJ remodeling events. Aim 2 Define mechanisms by which adhesion and actin dynamics are coordinately regulated. To accomplish the complex cell behaviors of morphogenesis, cells must closely coordinate adhesion and actin dynamics. Defining mechanisms by which this occurs is a key task for the field. We identified critical roles for the tyrosine kinase Abl and the actin regulator Ena in morphogenesis. We use them as a model for understanding mechanisms coordinating cell adhesion and cytoskeletal regulation. We hypothesize actin regulators like Ena are stored in an inactive state at AJs. We hypothesize that Abl regulates actin assembly at AJs and nearby by regulating Ena localization and/or function, acting as a scaffold, and influencing actin directly. We hypothesize that Ena helps generate distinct actin structures using different domains and partners, and that Ena is integrated with other actin regulators in filopodia. We will test these hypotheses.