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，而集体细胞运动又是适当组织组装和形态发生所需的。在脊椎动物原肠胚形成期间，一系列戏剧性的细胞重排最终导致胚层分离和脊椎动物体型的建立。随着这些事件的展开，所涉及的细胞和组织产生并响应由钙粘蛋白和整合素粘附承担的机械力。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，机械力通过组织和指导负责大量组织运动的细胞行为（包括细胞在细胞外基质（ECM）基质上的集体迁移），在形态发生中起着至关重要的指导作用。使用两栖动物非洲爪蟾的中内胚层作为一个强大的集体细胞运动模型，我们发现了一种新的机械响应钙粘蛋白复合物与角蛋白中间丝（KIF）细胞骨架的联系。这些钙粘蛋白依赖的细胞-细胞粘附抵抗并平衡在每个细胞前部产生的整合素依赖的牵引力。由此产生的力的各向异性是至关重要的组织集体细胞运动中的中内胚层。在这个项目期间，我们将确定参与指导细胞增殖活动的步骤，以响应钙粘蛋白上的牵引力，并建立中间丝对这一过程的贡献。有三个具体目标。第一个将建立关系 钙粘蛋白牵引力和Rho家族GTCRac 1的激活之间的关系。我们将解决的假设，有一个拮抗关系Rac活性和KIF细胞骨架，调节这些细胞中的增殖活性。第二个目标将建立两种类型的力的相对贡献的圆形中内胚层组织的逐步关闭：分布的牵引应力产生的单个细胞的纤连蛋白与收缩力产生的圆周排列的细胞在组织的前缘。最终的目标将采用基于光学的方法来选择性地干扰钙粘蛋白与肌动蛋白或角蛋白丝细胞骨架的连接。这些研究将阐述钙粘蛋白-KIF复合物对机械感知、极化细胞外基质活性的调节、与整合素-ECM粘附的串扰以及集体细胞间的协调的功能重要性。 迁移这项研究与组织再生和修复、正常组织形态发生及其在疾病中的失调（包括癌症和转移）的基本问题具有广泛的相关性。

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.