Physical mechanisms of 3D cell motility

项目摘要 细胞通过三维(3D)细胞外基质(ECM)的运动是正常 生理和疾病，包括伤口愈合和肿瘤转移。了解细胞如何移动 通过结构多样化的3D矩阵将是设计旨在控制细胞迁移的治疗方法的关键 在身体里。在3D迁移过程中，转移的肿瘤细胞和伤口愈合的成纤维细胞都面临着 同样的问题是：如何有效地移动笨重僵硬的原子核。而肌动球蛋白的收缩能力是 对于细胞在3D基质中移动它们的核是必不可少的，目前还不清楚它是如何被ECM调节的 结构或物理上耦合到原子核。另一层复杂性来自于3D 基质结构可以控制肌动球蛋白的收缩能力，以决定细胞用来移动的突起的类型(即 迁移可塑性)。通过了解3D ECM的结构如何影响 细胞核和肌动球蛋白的收缩能力，我们的目标是创建一个概念框架来解释人类如何以及为什么 细胞在不同的3D迁移机制之间切换。我们最近发现，人类细胞在一个 线状弹性3D基质依赖于基于整合素的细胞-基质黏附和肌球蛋白收缩的力量 把细胞核向前拉，就像活塞一样，从使用低压片状脂膜转为使用高压片状脂膜 叶状突。这个项目将检验原子核上的机械应力重新编程的假设 细胞内的结构和极性为细胞核提供能量，从而通过3D矩阵为细胞提供能量。至 为了实现这些目标，我们将结合生物物理和细胞生物学的方法来测量机械应力 并确定不同细胞骨架元素之间的分子连接 高压3D运动。目标1将操纵基质和核的物理结构，并 测量细胞组装和激活核活塞机制的能力。这一方法将 确定为什么3D矩阵可以重新编程细胞力生产以将细胞切换到压力驱动的3D 迁移。目标2将确定由3D基质细胞特异性激活的肌动球蛋白机制 产生压力和控制迁徙可塑性的相互作用。这些实验将清楚地区分 肌动球蛋白细丝负责将核活塞从对基质有反应的活塞上拉出来 通过增加牵引力来提高刚度。目标3将确定波形蛋白中间体的机制 细丝向细胞核传递力量，以维持细胞内的极化和细胞的定向运动 3D矩阵的狭窄范围。这些新方法将确定原子核是否是机械传感器。 这通过调节肌动球蛋白的收缩能力和3D细胞迁移模式来响应3D基质结构。 这增强了对定向3D细胞运动和迁移的基本原理的理解 可塑性将导致新的治疗策略，以控制体内正常和异常的细胞运动。

Project Summary Cell movement through three-dimensional (3D) extracellular matrix (ECM) is an essential component of normal physiology and disease, including wound healing and tumor metastasis. Understanding how cells move through structurally diverse 3D matrices will be essential to design therapies aimed at controlling cell migration in the body. During 3D migration, both metastatic tumor cells and wound healing fibroblasts are faced with the same problem: how to efficiently move the bulky, stiff nucleus. While the power of actomyosin contractility is essential for cells to move their nuclei in 3D matrices, it is not understood how it is regulated by the ECM structure or physically coupled to the nucleus. An additional layer of complexity comes from the fact that the 3D matrix structure can govern actomyosin contractility to dictate the type of protrusions cells use to move (i.e. migratory plasticity). By understanding how the structure of the 3D ECM affects the physical properties of the nucleus and actomyosin contractility, we aim to create a conceptual framework to explain how and why human cells switch between distinct 3D migration mechanisms. We recently discovered that human cells moving in a linearly elastic 3D matrix rely on integrin-based cell-matrix adhesions and the power of actomyosin contractility to pull the nucleus forward, like a piston, and switch from using low-pressure lamellipodia to high-pressure lobopodial protrusions. This project will test the hypothesis that mechanical stress on the nucleus reprograms intracellular architecture and polarity to power the nucleus, and thereby the cell through 3D matrices. To achieve these goals, we will combine biophysical and cell biology approaches to measure mechanical stress on the nucleus and determine the molecular connections between discreet cytoskeletal elements required for high-pressure 3D motility. Aim 1 will manipulate the physical structures of the matrix and the nucleus and measure the ability of the cells to assemble and activate the nuclear piston mechanism. This approach will establish why the 3D matrix can reprogram cellular force production to switch cells to pressure-driven 3D migration. Aim 2 will identify the actomyosin machinery that is specifically activated by 3D matrix-cell interactions to generate pressure and govern migratory plasticity. These experiments will clearly distinguish the actomyosin filaments responsible for pulling the nuclear piston forward from those that respond to matrix stiffness by increasing traction force. Aim 3 will determine the mechanisms by which vimentin intermediate filaments transmit force to the nucleus to sustain intracellular polarization and directional cell movement in the narrow confines of the 3D matrix. These novel approaches will determine if the nucleus is a mechanosensor that responds to 3D matrix structure by governing actomyosin contractility and the mode of 3D cell migration. This enhanced understanding of the fundamental principles of directional 3D cell motility and migratory plasticity will lead to new therapeutic strategies to control normal and abnormal cell movement in the body.