Biophysical control of cell migration by mechanical gradients in cell microenvironment

细胞迁移是一种基本的生物学现象，对单细胞和多细胞生物的生命至关重要。简单的生物体，如阿米巴原虫，必须迁移以寻找食物和交配。多细胞生物（如脊椎动物）的复杂性使得个体细胞和细胞群在发育形态发生期间定位自身的必要性，这主要通过定向细胞迁移来实现。在成人中，定向迁移在广泛的生理和病理过程中起着重要作用，包括皮肤和肠上皮的更新、免疫系统功能和伤口愈合，以及癌症转移和心血管疾病。在组织环境中，细胞迁移受到许多调节信号传导网络活性的引导因子的严格调节，这些信号传导网络反过来控制细胞内机制，例如肌动蛋白细胞骨架和粘着斑，从而驱动细胞迁移。在过去的几十年里，人们的注意力集中在了解化学信号，如可溶性生长因子，如何引导细胞迁移。然而，最近的研究强调了细胞微环境中的机械信号的重要性，包括细胞外基质（ECM）的密度和刚度在调节细胞迁移中的作用。通过使用合成ECM，已经表明许多细胞类型显示出向更硬ECM区域迁移的强烈倾向-这一过程称为硬旋转。趋硬性被认为是上皮-间充质转化和发育形态发生的关键，但其异常调节与癌细胞的恶性性质有关。尽管硬旋转具有明显的生物学和临床意义，但细胞感知机械信号并将其转化为细胞反应（如迁移）的确切机制在很大程度上是未知的。在这里，我们建议揭示这些机制，通过使用国家的组合--通过将这些方法与定量图像分析和细胞内信号传导途径的遗传/药理学扰动相结合，我们将揭示将机械引导线索转化为定向细胞迁移的特定分子参与者。该提案的具体目标包括：（1）解开由ECM刚度梯度引导的细胞中信号分子的时空动力学;（2）确定基于整合素的粘着斑的机械传感如何调节由ECM刚度梯度引导的细胞中的信号活动。这项研究将揭示基本的生物学机制，这些机制支持细胞如何感知微环境的机械特性，并将这些信息转化为特定的细胞反应-定向迁移。

Cell migration is a fundamental biological phenomenon that is critical for life of unicellular and multicellular organisms. Simple organisms, such as amebae, have to migrate to find food and to mate. Complexity of multicellular organisms, such as vertebrates, brings the necessity for individual cells and groups of cells to position themselves during developmental morphogenesis that is largely achieved through directed cell migration. In the adult, directed migration plays an important role in a wide range of physiological and pathological processes, including renewal of skin and intestinal epithelium, immune system function, and wound healing, as well as cancer metastasis and cardiovascular diseases. In tissue environment, cell migration is tightly regulated by numerous guidance cues that modulate activity of signaling networks, which in turn control intracellular machineries, e.g. actin cytoskeleton and focal adhesions, driving cell migration. In the last few decades, much attention has focused on understanding how chemical signals, such as soluble growth factors, guide cell migration. However, more recent studies have highlighted the importance of the mechanical signals in cell microenvironment, including density and stiffness of the extracellular matrix (ECM) in regulating cell migration. By using synthetic ECMs it has been shown that many cell types show a strong propensity to migrate toward areas of stiffer ECM - a process known as durotaxis. Durotaxis is thought to be critical to epithelial--to--mesenchymal transition and developmental morphogenesis, but its abnormal regulation is associated with the malignant nature of cancer cells. Despite the apparent biological and clinical significance of durotaxis, the exact mechanisms by which the cells sense mechanical cues and transduce them into cellular responses, such as migration,  are largely unknown. Here, we propose to reveal these mechanisms by using a combination of state--of--the--art biophysical techniques: synthetic hydrogels with tunable mechanical properties to mimic stiffness landscape of the biological tissues and genetically encoded FRET biosensors to visualize signaling activity across migrating cells. By coupling these approaches with quantitative image analysis and genetic/pharmacological perturbations of intracellular signaling pathways we will reveal specific molecular players that transduce mechanical guidance cues into directed cell migration. The specific aims of this proposal include: (1) To unravel the spatiotemporal dynamics of signaling molecules in the cells guided by a gradient of ECM stiffness; (2) To determine how mechanosensing by integrin-based focal adhesions modulates signaling activities in the cells guided by a gradient of ECM stiffness. The proposed study will uncover fundamental biological mechanisms that underpin how cells sense mechanical properties of the microenvironment and convert this information into a specific cellular response - directed migration.