Investigating mechanisms regulating cell adhesion during tissue remodeling

项目摘要：精确控制细胞间的黏附对维持组织的完整性至关重要 在发育和成体组织中。调节肿瘤黏附的信号异常激活可导致 上皮-间充质转化(EMT)与肿瘤转移，但其机制尚不完全清楚 这些调节信号通过这些信号导致粘附性的丧失。这一过程的一个拟议模型是 细胞黏附基因的转录抑制：EMT调控因子Snail抑制E- 钙粘蛋白黏附分子是黏附连接(AJ)的核心成分，被认为控制 以这种方式粘合。然而，最近的观察表明，蜗牛调节稳定性和 AJs的定位不依赖于E-钙粘附素水平的转录调控。具体机制是通过 目前尚不清楚Snail是如何监管AJ组织的，这突显了我们对 调节黏附和控制细胞决定接受EMT的信号。这样做的长期目标是 项目是确定在发育过程中如何控制细胞间的黏附，以使组织形态发生 和胚层的分离。这项提案的总体目标是确定蜗牛 在果蝇胚胎发育的组织重塑事件中调节AJ组织，以及 确定这一规定如何有助于细胞决定接受EMT。初步数据显示， 异位表达Snail导致E-钙粘附素蛋白定位从细胞连接迅速转移到 细胞内结构。其他观察表明，果蝇腹侧沟中的细胞经历了 由蜗牛依赖的关节失稳引起的关节重塑。值得注意的是，这一规定发生在 消耗母体提供的E-钙粘附素蛋白，且不受异位E-钙粘附素的抑制 表情。这些数据表明，Snail通过额外的POST-2控制AJ组织 转录机制。这项拟议工作的基本原理是不仅要深入了解 这些机制，以及在EMT事件中管理细胞黏附的一般原则。我们的中央 假说认为Snail通过改变连接钙粘附素的稳定性和定位来调节黏附 蛋白质复合体通过一种独立于E-钙粘附素转录调控的机制。这一假设 将通过追求两个具体目标进行测试：我们将(1)确定Snail影响AJ的机制 组织，以及(2)确定蜗牛可以促进果蝇上皮细胞EMT的物理条件 纸巾。我们的方法是创新的，因为它是第一批研究蜗牛的机械基础的方法之一- 钙粘素本地化的依赖转移，进一步是因为它使用了一种整合的策略，将 生物化学和细胞生物学方法。这项拟议的研究意义重大，因为预计它将 促进我们对细胞-细胞黏附的转录后调控的理解，并可能打开新的 了解癌症生物学和发育缺陷的研究途径。

Project Summary: Precise control of cell-cell adhesion is critical for maintaining tissue integrity during development and in adult tissues. Abnormal activation of signals that regulate adhesion in tumors can result in epithelial-mesenchymal transition (EMT) and cancer metastasis, but we do not fully understand the mechanism through which these regulatory signals cause loss of adhesion. A proposed model for this process is transcriptional repression of cell adhesion genes: the EMT regulator Snail represses expression of the E- cadherin adhesion molecule, a core component of Adherens Junctions (AJs), and is thought to control adhesion in this manner. However, recent observations have shown that Snail regulates the stability and localization of AJs independent of transcriptional regulation of E-Cadherin levels. The specific mechanism by which Snail regulates AJ organization remains unknown, highlighting an important gap in our understanding of the signals that regulate adhesion and govern the cellular decision to undergo EMT. The long-term goal of this project is to determine how cell-cell adhesion is controlled during development to enable tissue morphogenesis and segregation of germ layers. The overall objective of this proposal is to identify mechanisms by which Snail regulates AJ organization during tissue remodeling events during embryonic development in Drosophila, and determine how this regulation contributes to a cell’s decision to undergo EMT. Preliminary data indicate that ectopic Snail expression causes a rapid shift in E-Cadherin protein localization from cell junctions to intracellular structures. Other observations have shown that cells in the Drosophila ventral furrow undergo junctional remodeling through Snail-dependent destabilization of AJs. Remarkably, this regulation occurs prior to depletion of maternally provisioned E-Cadherin protein, and is not inhibited by ectopic E-cadherin expression. Together these data indicate that Snail controls AJ organization through an additional post- transcriptional mechanism. The rationale for this proposed work is to gain insight not only into the nature of these mechanisms, but also the general principles governing cell adhesion during EMT events. Our central hypothesis is that Snail modulates adhesion by altering the stability and localization of junctional cadherin protein complexes through a mechanism independent of E-cadherin transcriptional regulation. This hypothesis will be tested by pursuing two specific aims: we will (1) identify the mechanism through which Snail affects AJ organization, and (2) define the physical conditions in which Snail can promote EMT in Drosophila epithelial tissues. Our approach is innovative because it is one of the first to examine the mechanistic basis of Snail- dependent shifts in cadherin localization, and further because it uses an integrative strategy that combines biochemical and cell biological approaches. The proposed research is significant because it is expected to advance our understanding of the post-transcriptional regulation of cell-cell adhesion, and may open new avenues of research for understanding cancer biology and developmental defects.