Mechanosensitive signaling and cell adhesion during migration

迁移过程中的机械敏感信号传导和细胞粘附

• 批准号: 7409323
• 负责人: Gregory Weber
• 金额: $ 4.68万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-03-15 至 2010-03-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7409323
• 关键词: Actins; Address; Adhesions; Adhesives; Adult; Affect; Arts; Atomic Force Microscopy; Behavior; Biological Assay; Biological Models; Biomechanics; C cadherin; Cadherins; Cell Adhesion; Cell Polarity; Cell-Cell Adhesion; Cell-Matrix Junction; Cells; Chemotaxis; Chimeric Proteins; Complex; Condition; Confocal Microscopy; Cues; Cultured Cells; Development; Disease; Elements; Environment; Exhibits; Family; Fibronectins; Fluorescence Resonance Energy Transfer; Focal Adhesions; Green Fluorescent Proteins; Guanosine Triphosphate Phosphohydrolases; Immigration; In Vitro; Individual; Integrins; Investigation; Life; Ligands; Link; Mechanics; Mediating; Microscopy; Modeling; Molecular; Morphogenesis; Movement; Pathology; Physiology; Platelet-Derived Growth Factor; Process; Regulation; Resolution; Role; Signal Transduction; Speed; System; Techniques; Testing; Time; Tissues; Traction; Xenopus; cell motility; cell type; cellular imaging; computerized data processing; extracellular; in vivo; insight; magnetic beads; migration; response; rho; spatiotemporal

DESCRIPTION (provided by applicant): Cell migration is a complex process that involves both physical adhesive mechanics and intracellular signaling, which are profoundly interdependent on one another. Mechanisms of cell migration have been studied intensively using single cells cultured in vitro, but our understanding of cell migration in vivo remains limited. Cell migration in vivo often occurs in multicellular arrays that produce bulk tissue movements critical to morphogenesis, normal adult physiology, and various pathologies. While the function of mechanical signaling generated through cell-matrix traction forces has frequently been the subject of investigation, mechanosensitive responses to tension in cell-cell adhesions and the subsequent effects of these adhesive contacts on cell migration have not been elucidated. I will test the hypothesis that mechanical signaling, initiated by cadherin-mediated cell-cell adhesions in intact migratory tissues, has a central role in coordinating cell polarization and directional migration of individual cells to direct intact tissue movements. Mechanisms of cell migration are widely conserved between different species, tissues, and cell types. The migratory mesendoderm that traverses the fibronectin matrix of the blastocoel roof during Xenopus gastrulation will be used as a model system for these studies. This model provides an ideal system in which to study normal in vivo migratory processes because the motile behaviors of the intact tissue are retained when cultured in vitro and thus, the system is accessible to a variety of experimental manipulations. There are four specific aims proposed to address the significance of cell-cell adhesion in the propagation of mechanical signals that regulate mesendoderm cell polarity and migration. In specific aim 1, mesendoderm explants and a magnetic bead "pull" assay will be used to establish the role of biomechanical tension on C-cadherin dependent adhesions as an important regulator of polarization and directional migration in this tissue. Cadherins and integrins provide a physical link between extracellular ligands and internal cytoskeletal elements. GFP fusion proteins will be used to track cytoskeletal dynamics as a consequence of mechanical signaling initiated by cadherin adhesions in specific aim 2. Actin cytoskeletal dynamics involved in migration is signaled by spatiotemporal activation of the small Rho family GTPases. In specific aim 3, FRET analysis will be used to examine the local subcellular activation of Rho family GTPases as a result of C-cadherin and a5 (31 integrin-dependent mechanosensitive cell signaling. Finally, traction force microscopy and reflection contrast microscopy approaches will be used in specific aim 4 to determine the specific force relationship between cell-matrix adhesions and C-cadherin cell-cell contacts in migrating mesendoderm.

描述（由申请人提供）：细胞迁移是一个复杂的过程，涉及物理粘附力学和细胞内信号传导，它们彼此之间深刻地相互依赖。人们已经利用体外培养的单细胞对细胞迁移机制进行了深入研究，但我们对体内细胞迁移的了解仍然有限。体内细胞迁移通常发生在多细胞阵列中，产生对形态发生、正常成人生理学和各种病理学至关重要的大量组织运动。虽然通过细胞基质牵引力产生的机械信号的功能经常成为研究的主题，但对细胞间粘附张力的机械敏感反应以及这些粘附接触对细胞迁移的后续影响尚未阐明。我将检验以下假设：由完整迁移组织中钙粘蛋白介导的细胞间粘附引发的机械信号传导在协调细胞极化和单个细胞的定向迁移以指导完整组织运动方面具有核心作用。细胞迁移机制在不同物种、组织和细胞类型之间广泛保守。在非洲爪蟾原肠胚形成过程中穿过囊胚腔顶部纤连蛋白基质的迁移中内胚层将被用作这些研究的模型系统。该模型提供了一个研究正常体内迁移过程的理想系统，因为在体外培养时完整组织的运动行为得以保留，因此该系统可进行各种实验操作。提出了四个具体目标来解决细胞间粘附在调节中内胚层细胞极性和迁移的机械信号传播中的重要性。在具体目标 1 中，中内胚层外植体和磁珠“拉力”测定将用于确定生物力学张力对 C-钙粘蛋白依赖性粘附的作用，作为该组织中极化和定向迁移的重要调节剂。钙粘蛋白和整合素提供细胞外配体和内部细胞骨架元件之间的物理连接。 GFP 融合蛋白将用于跟踪细胞骨架动力学，这是特定目标 2 中钙粘蛋白粘附引发的机械信号传导的结果。参与迁移的肌动蛋白细胞骨架动力学由小 Rho 家族 GTP 酶的时空激活发出信号。在具体目标 3 中，FRET 分析将用于检查 C-钙粘蛋白和 a5 (31 整合素依赖性机械敏感细胞信号传导) 导致的 Rho 家族 GTPases 的局部亚细胞激活。最后，在具体目标 4 中将使用牵引力显微镜和反射对比显微镜方法来确定迁移中内胚层中细胞-基质粘附和 C-钙粘蛋白细胞-细胞接触之间的特定力关系。