RUI: Mapping obscurin’s role as a force sensor

所有细胞都必须感知力并对力作出反应，否则物理力量会将它们撕裂。然而，人们对细胞如何将纯物理刺激（如力）转化为生化信号仍知之甚少。一种潜在的机制涉及到巨大的黑蛋白。这种蛋白质的一端与细胞膜结合，另一端与细胞骨架结合。蛋白质的中间部分就像一根绳子，它可以改变形状和拉长，而不需要任何主要的能量贡献。但如果蒙布林被拉伸得太远，它就会开始像弹簧一样，逐渐抵抗额外的拉伸。蒙昧素还含有酶活性区域，当它被拉伸时，酶活性区域被认为是“开启”的。这个项目测试了一种模型，即暗素是一种信号转导机械分子，可以感知并响应活细胞所经历的拉伸物理力。这项研究将深入了解细胞如何“感受”力，它们如何在分子水平上对这种力做出反应，以及这种现象如何影响组织生长。这个项目的更广泛的影响整合了一个强大的本科科学教育计划和社区外展。与詹姆斯麦迪逊大学工程学系的顶点班合作，将开发和建造交互式机器，教授分子生物学的基本概念。这些机器将在当地的县博览会上展出，将科学和工程概念带给经常被排除在其他科学推广活动之外的人群。该项目通过聚焦于一种假定的拉伸传感器蛋白，解决了膜和细胞骨架之间机械转导的新机制。这项研究将确定在细胞内是否处于紧张状态。它还将决定其定位和信号传导与细胞所经历的力直接相关的程度。第三，这项工作将确定模糊蛋白定位是否会影响细胞片的运动，以响应它们所经历的力。为了回答这些问题，该项目将结合FRET显微镜、荧光张力传感器、组织消融技术、细胞生物学方法和计算机建模。从这些实验中获得的数据将用于创建细胞运动的预测模型。总之，这些实验将揭示蒙昧素在细胞机械感觉中的作用。由于在多种细胞类型中都发现了暗布蛋白，因此这项基础科学工作有望对更广泛的特定主题产生影响，包括胚胎发育、细胞侵袭性、内皮细胞生物学、肌肉生物学和心肌细胞。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

All cells must sense and respond to force, otherwise physical forces would tear them apart. However, it is still poorly understood how cells turn a purely physical stimulus, like force, into a biochemical signal. One potential mechanism involves the giant protein obscurin. One end of this protein binds to the cell membrane and the other end binds the cytoskeleton. The middle of the protein acts like a rope in that it is able to change shape and elongate without requiring any major energy contribution. But if obscurin is stretched too far, it begins to act like a spring, gradually resisting additional stretch. Obscurin also contains enzymatically active regions that are thought to ‘turn on’ when it is stretched. This project tests the model that obscurin is a signal transducing mechano-molecule that senses and responds to the physical force of stretch experienced by living cells. This research will provide insights into how cells ‘feel’ force, how they react to this force on a molecular level, and how this phenomenon informs tissue growth. The Broader Impacts of this project integrate a strong undergraduate science education program and community outreach. In collaboration with the capstone class of James Madison University’s engineering department, interactive machines will be developed and built that teach fundamental concepts of molecular biology. These machines will be displayed at local county fairs, bringing science and engineering concepts to a population that is often excluded from other science outreach efforts. This project addresses a novel mechanism of mechano-transduction between membranes and the cytoskeleton of the cell by focusing on the protein obscurin, a putative stretch sensor. This research will determine whether obscurin is under tension within the cell. It will also determine the extent to which its localization and signaling are directly related to the forces experienced by the cell. Third, this work will determine whether obscurin localization influences the motility of sheets of cells in response to the force they experience. To answer these questions, this project will use a combination of FRET microscopy, fluorescent tension sensors, tissue ablation techniques, cell biological approaches, and computer modelling. The data obtained from these experiments will be used to create a predictive model of cell motility. Together, these experiments will reveal obscurin’s role in cellular mechanosensation. Because obscurin is found in multiple cell types, this basic science work is expected to have impact on a broad range of more specific topics, including embryonic development, cell invasiveness, endothelial cell biology, muscle biology, and cardiomyocytes.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.