Probing the logic of Hippo-based mechanotransduction with optogenetics

物理力在细胞决策过程中发挥着重要作用，如增殖、分化和迁移。细胞力学的扰动导致发育障碍和包括癌症在内的疾病。因此，了解物理力和细胞机械转换器之间的相互调节对于细胞生理学和病理生理学非常重要。解决这个问题是复杂的紧耦合的个别子过程内的肌动蛋白细胞骨架，并与其他mechanosensory结构。它需要工具来测量和操纵特定的，空间和时间上定义的，细胞的物理和生化特性。为了实现这一点，我建议使用光遗传学探测肌动蛋白依赖的机械通过河马途径，一个重要的细胞mechanosensor的逻辑。总的来说，我追求三个目标。首先，通过控制机械输入的大小和动态来揭示Hippo通路的信号接收和传输逻辑。第二，询问力学和雅普激活在控制细胞命运中的相对贡献。第三，确定肌动蛋白细胞骨架的特征，这些特征是由Hippo通路感知的。使用组合的共聚焦和原子力显微镜将允许精确的光遗传学操作和细胞力学和Hippo信号的同时分析。为了探索这些发现在一个需要Hippo信号传导的系统中的意义，该研究将在小鼠胚胎干细胞中进行。从这些结果中，我期望对细胞力学和行为的相互调节有基本的了解。

Physical forces play a fundamental role for cellular decision-making processes such as proliferation, differentiation, and migration. Perturbation of cellular mechanics causes both developmental disorders and diseases including cancer. Understanding the reciprocal regulation between physical forces and cellular mechanotransducers is therefore important for cell physiology and pathophysiology. Addressing this problem is complicated by the tight coupling of individual sub-processes both within the actin cytoskeleton and in connection with other mechanosensory structures. It requires tools for measuring and manipulating specific, spatially and temporally defined, physical and biochemical properties of the cell. To achieve this, I propose to use optogenetics to probe the logic of actin-dependent mechanotransduction through the Hippo pathway, an important cellular mechanosensor. Overall, I pursue three goals. First, to reveal the logic of signal reception and transmission of the Hippo pathway by controlling the magnitude and dynamics of the mechanical input. Second, to interrogate the relative contributions of mechanics and YAP activation in control of cell fate. And third, to identify the features of the actin cytoskeleton that are sensed by the Hippo pathway. Using a combined confocal and atomic force microscope will allow precise optogenetic manipulations and simultaneous analysis of cell mechanics and Hippo signaling. To probe the significance of the findings in a system that requires Hippo signaling for its physiology, the study will be performed in mouse embryonic stem cells. From the results, I expect fundamental insights into the reciprocal regulation of cell mechanics and behavior.