Cadherin Mechanotransduction

这笔赠款建立在我们的新发现基础上，即E-钙粘附素在上皮细胞-细胞连接处传递机械信号 通过表皮生长因子受体(EGFR)激活激活级联信号。E-钙粘素是一种 上皮细胞-细胞连接处的基本黏附蛋白和E-钙粘附素复合体也转导力量，以 调节细胞形态和上皮屏障的完整性。这些新的发现表明，E-钙粘附素- 信号转导也激活了调控细胞增殖、形态发生和疾病的信号。总的目标是 该计划的目的是确定E-钙粘素机械激活EGFR的初始步骤，并建立 这一机制的更广泛的生理含义。我们的初步数据也表明这一点 力激活信号通路调节应激细胞-细胞连接的细胞骨架强化 E-钙粘附素复合体中的α−连接素。这一意想不到的发现支持了EGFR和E- 钙粘附素是上皮细胞连接的核心力转导机制的重要组成部分。在这 程序，特殊目标1测试机械刺激的E-钙粘素激活EGFR的假设 磷酸化，通过触发假定的E-钙粘素/EGFR复合体的破坏。《特定目标2》将使用 创新的基于荧光的方法学，由合作者赫里斯托娃(约翰斯·霍普金斯)开发 研究E-钙粘蛋白和EGFR在质膜上的直接相互作用。建议的研究包括 基于大量的初步数据，这些数据揭示了蛋白质之间的直接联系。生物物理学研究将 使用E-钙粘蛋白和EGFR突变体的子集，建立这种联系的分子要求。 《特殊目标3》将在一个三维的器官类型中测试这些发现的生理学意义。 人类乳腺上皮组织模型，与韦弗大学(UCSF)合作。研究将确定 E-钙粘蛋白/EGFR复合体是否真的是与整合素协调的中枢力敏感单位 调整形态发生和恶性程度，以响应组织力学。乳腺上皮细胞的三维培养 基因工程表达E-钙粘蛋白突变体(Aim 2)将决定E-钙粘蛋白/EGFR复合体的影响 破坏增殖、形态发生和侵袭，作为基质刚性的函数。整合素很好 已知与EGFR协调以调节乳房组织发育和肿瘤进展。这些研究 可能会使E-钙粘附素成为这个对力量敏感的网络中的一个重要组成部分。

This grant builds on our novel discovery that E-cadherin at epithelial cell-cell junctions transduces mechanical signals, by activating a kinase cascade via the epidermal growth factor receptor (EGFR). E-cadherin is an essential adhesion protein at epithelial cell-cell junctions, and E-cadherin complexes also transduce force, to regulate cell shape and epithelial barrier integrity. These new findings suggest that E-cadherin force- transduction also activates signals that regulate cell proliferation, morphogenesis, and disease. The broad goal of this program is to identify initial steps in the mechanical activation of EGFR by E-cadherin, and to establish the broader physiological implications of this mechanism. Our preliminary data also demonstrate that this force-activated signaling pathway regulates the cytoskeletal reinforcement of stressed cell-cell junctions by α−catenin in E-cadherin complexes. This unexpected finding supports the hypothesis that EGFR and E- cadherin are essential components in the core force-transduction machinery at epithelial cell junctions. In this program, Specific Aim 1 tests the hypothesis that mechanically stimulated E-cadherin activates EGFR phosphorylation, by triggering the disruption of putative E-cadherin/EGFR complexes. Specific Aim 2 will use innovative fluorescence-based methodology, developed by collaborator Hristova (Johns Hopkins) to investigate direct interactions between E-cadherin and EGFR at the plasma membrane. Proposed studies are based on substantial preliminary data, which reveal direct protein-protein association. Biophysical studies will establish the molecular requirements for this association, using a subset of E-cadherin and EGFR mutants. Specific Aim 3 will test the physiological implications of these findings in a three-dimensional, organotypic model of human mammary epithelial tissue, in collaboration with Weaver (UCSF). Studies will determine whether E-cadherin/EGFR complexes are indeed central force-sensing units that coordinate with integrins to tune morphogenesis and malignancy, in response to tissue mechanics. 3D cultures of breast epithelial cells engineered to express E-cadherin mutants (Aim 2) will determine the impact of E-cadherin/EGFR complex disruption on proliferation, morphogenesis, and invasion, as a function of matrix rigidity. Integrins are well known to coordinate with EGFR to regulate breast tissue development and tumor progression. These studies would potentially establish E-cadherin as an essential component in this force-sensitive network.