Cell-Cell and Cell-Matrix Interactions in Morphogenesis

描述（由申请人提供）：拟议研究的主要目标是阐明负责协调集体细胞运动的机制，而集体细胞运动反过来又需要适当的组织组装和形态发生。在脊椎动物原肠胚形成过程中，一系列戏剧性的细胞重排最终导致胚层的分离和脊椎动物身体计划的建立。随着这些事件的展开，相关的细胞和组织产生并响应钙粘蛋白和整合素粘附所承受的机械力。很少有研究试图将这种机械信号的涨落与体内单个细胞或细胞群的特定运动行为的出现联系起来。我们的总体假设是，机械力在形态发生中发挥着重要的指导作用，通过组织和指导细胞行为，包括细胞在细胞外基质（ECM）基质上的集体迁移。利用两栖动物非洲爪蟾（Xenopus laevis）的中胚层作为集体细胞运动的稳健模型，我们发现了一种与角蛋白中间丝（KIF）细胞骨架相连的新型机械响应钙粘蛋白复合物。这些钙粘蛋白依赖的细胞-细胞粘附抵抗并平衡每个细胞前部产生的整合素依赖的牵引力。由此产生的力各向异性对中胚层细胞集体运动的组织至关重要。在这个项目期间，我们将确定指导细胞突出活动的步骤，以响应钙粘蛋白上的拉力，并确定中间细丝对这一过程的贡献。有三个具体目标。第一个将建立关系

DESCRIPTION (provided by applicant): The primary goal of the proposed research is to elucidate mechanisms responsible for the coordination of collective cell movements that, in turn, are required for proper tissue assembly and morphogenesis. During vertebrate gastrulation, a dramatic series of cellular rearrangements culminates in the segregation of germ layers and the establishment of the vertebrate body plan. As these events unfold, the cells and tissues involved generate and respond to mechanical forces that are borne by both cadherin and integrin adhesions. Few studies have attempted to link this ebb and flow of mechanical signals to the emergence of specific motile behaviors of individual cells or groups of cells in vivo. Our overall hypothesis is that mechanical forces play an essential instructive role in morphogenesis by organizing and directing the cellular behaviors responsible for bulk tissue movements including the collective migration of cells on extracellular matrix (ECM) substrates. Using the mesendoderm of the amphibian Xenopus laevis as a robust model for collective cell motility, we have discovered a novel mechanically-responsive cadherin complex with links to the keratin intermediate filament (KIF) cytoskeleton. These cadherin-dependent cell-cell adhesions resist and balance integrin-dependent traction forces generated at the front of each cell. The resulting force anisotropy is crucial to the organization of collective cell movements in the mesendoderm. In this project period we will determine the steps involved in directing cell protrusive activity i response to tugging forces on cadherins and establish the contributions of intermediate filaments to this process. There are three specific aims. The first will establish the relationship between cadherin tugging forces and activation of the Rho family GTPase Rac1. We will address the hypothesis that there is an antagonistic relationship between Rac activity and the KIF cytoskeleton that regulates protrusive activity in these cells. The second aim will establish the relative contributions of two types of forces to the progressive closure of the circular mesendoderm tissue: distributed traction stresses generated by individual cells on fibronectin vs. contractile forces generated by the circumferentially arranged cells at the leading-edge of the tissue. The final aim will employ optically based approaches to selectively perturb cadherin linkages to either the actin or keratin filament cytoskeletons. These studies will address the functional importance of the cadherin-KIF complex to mechanosensing, regulation of polarized protrusive activity, crosstalk with integrin-ECM adhesions, and the coordination of collective cell migration. This research has broad relevance to fundamental problems of tissue regeneration and repair, normal tissue morphogenesis and its dysregulation in diseases, which include cancer and metastasis.