Subcellular Mechanical Force Transduction

描述（由申请人提供）：该项目的重点是在细胞与其细胞外基质之间的粘附部位所经历的亚细胞力在调节细胞组织和功能中的作用。细胞在空间上分辨和响应这种局部力的适应性过程对于细胞运动所需的粘附重塑至关重要，其中细胞在分解其他粘附的同时加强特定的粘附。在这一领域的先前工作已经表明，粘着斑组装响应于施加到它们的力，但由于粘附到细胞外基质也诱导生化信号，触发细胞收缩性，它仍然不清楚是否施加到特定的粘附力导致任何机械串扰与分布在整个细胞的剩余的粘附。该项目将联合收割机的专业知识，2个研究人员采用一种新的技术，同时允许局部机械刺激的粘附表面的细胞和空间分辨测量的局部力场产生的整个细胞在响应于这种刺激。有人提出，局部刺激和整体机械反应之间的关系对于细胞运动所需的细胞骨架内的机械协调至关重要。1的研究人员最近开发了一种技术，其中的偏转阵列的微型职位报告的细胞骨架张力和局部力场所产生的细胞附着到阵列。在这个项目中，嵌入在单个柱子中的纳米工程磁性材料将用于使这些柱子变形，从而对附着的细胞施加可调的亚细胞机械应力，而这些应力的影响同时由周围的柱子测量。该项目的具体目标1将是表征磁柱器械在纳牛顿水平向细胞施加明确定义的力的能力，校准这些力，并证明应力向细胞的传递。具体目标2是通过研究收缩性和细胞力分布的变化来测量细胞对局部力的整体反应，具体目标3是测量局灶性粘连对局部施加的力的非局部反应，以确定局灶性粘连是否由于细胞力的分布变化而在整体上重新分布，并确定焦点粘附分布如何随局部施加的力的强度而变化。这种新技术的发展将导致对细胞如何将机械应力转换为适应性，协调的细胞骨架反应的新理解，并将开辟一条通往炎症，癌症转移和组织发育关键的细胞运动机制的新见解的途径。

DESCRIPTION (provided by applicant): This project focuses on the role that sub-cellular forces experienced at sites of adhesion between cells and their extracellular matrix play in regulating cell organization and function. The adaptive process by which cells spatially resolve and respond to such localized forces is critical to the adhesion remodeling required for cell motility, where cells reinforce specific adhesions while disassembling others. Previous work in this area has shown that focal adhesions assemble in response to forces applied to them, but because adhesion to extracellular matrix also induces biochemical signals that trigger cell contractility, it remains unclear whether forces applied to specific adhesions result in any mechanical crosstalk with the remaining adhesions distributed throughout the cell. This project will combine the expertise of 2 investigators to employ a new technique that simultaneously allows local mechanical stimulation of the adherent surface of a cell and spatially-resolved measurement of the local force fields generated throughout the cell in response to this stimulation. It is proposed that the relationship between local stimulation and global mechanical response is critical to the mechanical coordination within the cytoskeleton required for cell motility. 1 of the investigators has recently developed a technique wherein the deflections of an array of microfabricated posts report the cytoskeletal tension and local force fields generated by a cell attached to the array. In this project, nanoengineered magnetic material embedded in individual posts will be used to deform those posts, thereby exerting tunable subcellular mechanical stresses to attached cells, while the effects of the these stresses are simultaneously measured by the surrounding posts. Specific Aim 1 of the project will be to characterize the ability of the magnetic post device to apply well-defined forces at the nanonewton level to cells, to calibrate these forces, and to demonstrate the transmission of stresses to the cells. Specific Aim 2 will be to measure the global response of a cell to local forces through studies of changes in contractility and in the distribution of cellular forces, Specific Aim 3 will be to measure the non-local response of focal adhesions to locally applied forces, to determine whether focal adhesions re-distribute globally as a result of the distributed changes in cellular forces, and to determine how focal adhesion distributions vary as a function of the strength of a locally applied force. Development of this novel technique will lead to new understanding of how mechanical stresses are transduced by cells into an adaptive, coordinated cytoskeletal response, and will open a pathway toward new insights into the mechanisms of cell motility critical for inflammation, cancer metastasis, and tissue development.