Defining Mechanical Landscapes at Cell-Cell Junctions

项目总结 机械力在许多内在的和集体的细胞过程中扮演着基本的角色，包括组织 再生、形态发生和肿瘤转移。虽然广泛的研究集中在部队 在细胞和细胞外基质之间，单个细胞之间的机械相互作用似乎是 很重要，但特点不佳。众所周知，这些细胞间力在伤口愈合过程中是至关重要的， 癌细胞侵袭，以及其他发育和动态平衡过程。然而，分子原理 控制这些精细平衡的机械转导事件的机制仍然知之甚少。为了描绘出 这些集体细胞过程的机制，测量细胞间力和相互关系是至关重要的 确定的机械景观与特定的分子机制，调节细胞信号。 我们的实验室已经提出了精确和易于使用的基于DNA的传感器来可视化和量化细胞间 力量。我们和其他人最近开发了一种有效的基于脂质的方法来锚定设计师DNA 哺乳动物细胞膜外表面上的序列。通过采用这种方法， 膜锚定的DNA探针可以灵敏地成像细胞-细胞之间的大范围分子作用力 交汇点。目前的机械生物学研究是基于通常仅在少数情况下进行的技术 专业实验室。所建议的传感器与容易获得的荧光兼容 显微镜，高度坚固和多功能，易于准备和使用。为了进一步发展和适应这些 传感器用于研究细胞间的机械敏感事件，未来的研究计划是：(1)工程和 优化基于DNA的拉力和压力传感器，以测量大范围的细胞间力 单分子水平。(2)以表征良好的钙粘素为基础的机械转导为例， 量化和监测集体细胞迁移和神经板成形过程中细胞-细胞连接处的作用力。(3) 应用这些传感器来研究Notch激活在免疫细胞激活中的机械作用。凹槽 信号在不同的发育和疾病过程中高度保守。细胞间配体诱导 在激活缺口时需要机械力。取决于环境和机械环境， 缺口的激活在组织生长和免疫反应的调节中可能起到相反的作用。我们的 这些结果将为阐明Notch信号激活的机制提供独特的见解。 我们的长期目标是使细胞间力测量在生命科学实验室得到广泛应用。 这些新型传感器将被广泛用于理解开发的基本机械原理， 生理学和疾病，这也将成为制定新战略的关键基础 组织工程学、再生医学和细胞疗法。

PROJECT SUMMARY Mechanical forces play fundamental roles in many intrinsic and collective cellular processes, including tissue regeneration, morphogenesis, and tumor metastasis. While extensive studies have focused on the forces between cells and extracellular matrices, mechanical interactions among individual cells appear to be important yet poorly characterized. These intercellular forces are known to be critical during wound healing, cancer cell invasion, and other developmental and homeostatic processes. However, the molecular principles that govern these finely balanced mechanotransduction events are still poorly understood. To depict the mechanisms of these collective cellular processes, it is essential to measure intercellular forces and correlate the determined mechanical landscapes with the specific molecular machineries that regulate cellular signaling. Our lab have proposed precise and easy-to-use DNA-based sensors to visualize and quantify intercellular forces. We and others recently developed an efficient lipid-based approach to anchor designer DNA sequences onto the external surfaces of mammalian cell membranes. By employing this approach, membrane-anchored DNA probes allow sensitive imaging of a broad range of molecular forces at cell-cell junctions. Current mechanobiology studies are based on techniques typically performed in only a few specialized laboratories. The proposed sensors are compatible with readily accessible fluorescence microscopes, highly robust and versatile, and easy to prepare and use. To further develop and adapt these sensors to study intercellular mechanosensitive events, the future research plan is to: (1) engineer and optimize DNA-based tensile and compressive force sensors to measure a broad range of intercellular forces at the single-molecule level. (2) Use well-characterized cadherin-based mechanotransduction as an example, quantify and monitor forces at cell-cell junctions during collective cell migrations and neural plate shaping. (3) Apply these sensors to investigate the mechanical roles of Notch activation in immune cell activation. Notch signaling is highly conserved in different developmental and disease processes. Intercellular ligand-induced mechanical forces are required in Notch activation. Dependent on the environmental and mechanical context, Notch activation can have contrary effect in the regulation of tissue growth and immune responses. Our results will provide unique insights to elucidate the mechanical mechanisms of Notch signal activation. Our long-term goal is to make intercellular force measurements widely implemented in life science laboratories. These novel sensors will be broadly used to understand the basic mechanical principles of development, physiology, and disease, which will also serve as the critical foundation for developing novel strategies in tissue engineering, regenerative medicine, and cell therapy.