Direct and Quantitative Probing of Desmosome Mechanotransduction

项目摘要 细胞间粘附连接，包括桥粒和粘附连接，连接上皮细胞内， 组织提供机械完整性，调节细胞分选和迁移，并控制化学信号， 进一步指示小区决策。特别地，桥粒抵抗机械应力并响应机械应力。 调节复杂细胞行为以促进分化、促进迁移和伤口愈合的信号，以及 介导在发育和许多组织（包括皮肤和心脏）的疾病中至关重要的其他功能。 虽然在确定桥粒内潜在的承重元件方面已经取得了重大进展， 关于它们在上皮力学和机械传导中的作用存在巨大的知识缺口。具体而言是 对桥粒-中间丝连接的力调节机制知之甚少。 关于桥粒的分子成分何时甚至是否具有 机械载荷，这是机械转换的第一步。更重要的是，仍然没有解决的是， 特定的桥粒组分作为机械传感器， 在适应组织中的机械应力中起关键作用的信号通路。我们之前的研究表明， 桥粒-中间丝连接在调节细胞力学中的能力，长期以来一直被认为是 被认为只属于粘附连接。这表明桥粒组分可能 参与武力管制。在这个MIRA项目中，基于这些发现并利用新开发的 单细胞-细胞粘附询问平台，我们将研究重要的，但研究较少的作用， 桥粒在上皮力学和力学转导中起作用。我们将重点进行两项主要研究 推进领域：1）研究桥粒在维持上皮细胞的机械完整性中的作用- 细胞连接和2）确定其在与细胞连接协调的连接处转导机械线索的潜力， 粘附连接处和肌动蛋白网络处的机械敏感分子。通过一系列关于 上皮细胞与桥粒成分窝藏损失和获得的功能突变，我们将量化 每个桥粒蛋白在应激条件下维持上皮机械完整性的贡献。 我们将回答这个问题：肌动蛋白为基础的收缩性如何影响桥粒调节内张力 肌动蛋白基粘合剂网络我们期待着提供第一个直接观察桥粒作为一个 在细胞-细胞连接处的机械转导位点。建议的研究将使我们更了解如何 桥粒协调化学和转录途径，以响应细胞中的机械张力。 上皮细胞，为其他组织中桥粒机械传感的类似研究奠定基础，并最终 提供新的知识，以帮助开发对这些干扰引起的疾病的治疗方法。 粘附连接和它们的机械感应路径。

PROJECT SUMMARY Intercellular adhesive junctions, including desmosomes and adherens junctions, connect epithelial cells within a tissue to provide mechanical integrity, regulate cell sorting and migration, and control chemical signals that further instruct cell decisions. In particular, desmosomes resist mechanical stress and respond to mechanical cues to regulate complex cell behaviors to promote differentiation, facilitate migration and wound healing, and mediate other functions critical in development and in diseases of many tissues, including the skin and heart. While significant progress has been made identifying potential load-bearing elements within desmosomes, a huge knowledge gap exists about their roles in epithelial mechanics and mechanotransduction. Specifically, the mechanisms of force regulation across the desmosome-intermediate filament linkage are poorly understood. There is limited direct evidence for when and even whether the molecular components of desmosomes bear mechanical loads, the first step towards mechanotransduction. More importantly, still unaddressed is whether specific desmosome components act as mechanosensors that determine the strength and duration of chemical signaling pathways critical in adapting to mechanical stresses in tissues. Our previous studies showed the capacity of desmosome-intermediate filament linkage in regulating cell mechanics, a role that has long been regarded to belong solely to adherens junctions. This suggests the potential for desmosomal components to participate in force regulation. In this MIRA project, building on these findings and leveraging a newly developed single cell-cell adhesion interrogation platform, we will examine the important, but less studied, role that desmosomes play in epithelial mechanics and in mechanotransduction. We will focus on two major research thrust areas: 1) investigate the role desmosome plays in maintaining the mechanical integrity of epithelial cell- cell junctions and 2) determine its potential in transducing mechanical cues at the junction in coordination with mechanosensitive molecules at the adherens junctions and the actin network. Through a series of studies on epithelial cells with desmosomal components harboring loss- and gain-of-function mutations, we will quantify the contribution from each desmosomal protein in maintaining epithelial mechanical integrity in stressed conditions. We will answer the question: How does actin-based contractility affect desmosome regulation of tension within actin-based adhesive networks? We anticipate providing the first direct observation of desmosome serving as a mechanotransduction site at the cell-cell junction. The proposed studies will enhance our understanding of how desmosomes coordinate chemical and transcriptional pathways in response to mechanical tension in the epithelia, lay the groundwork for similar studies of desmosome mechanosensing in other tissues, and ultimately provide new knowledge to aid in developing treatments for disorders resulting from interference with these adhesive junctions and their mechanosensing pathways.