Collaborative Research: Mechanisms of Gradient Sensing by 'Feel' in Cell Migration Directed by Extracellular Matrix

细胞迁移是身体许多过程中的重要步骤，例如伤口愈合和对感染的反应。细胞与基质接触，使它们能够爬行和探索。要做到这一点，细胞必须通过触摸和感觉主动感知基质属性。这个合作项目的首要目标是统一或区分与基质接触的数量（通过触摸的粘性）与质量（通过感觉的硬度）的细胞感测。多学科研究将与研究生培训，工程课程的教学和推广相结合，这将促进科学交流。细胞迁移是组织发育和稳态的基本过程。趋化性的分子决定因素，细胞迁移偏向于一个梯度的可溶性引诱剂，是合理的理解，并出现了一个范例，表征某些细胞内信号转导途径作为一个分子“罗盘”。相比之下，分别由固定化配体密度梯度和机械刚度梯度引导的趋触性和硬旋转性（haptotaxis and durotaxis）迁移知之甚少，并且需要新的范例，考虑到细胞必须主动遇到并响应物理线索（即，通过“感觉”）而不是通过扩散配体的被动感测。在目标1下，目的是定义控制迁移细胞中偶联细胞骨架动力学的反馈机制。据推测，粘附介导的信号通路控制的持续时间板状伪足突起和空间协调的F-肌动蛋白束。高分辨率，活细胞显微镜和分子扰动的假定信号和细胞骨架过程将被应用到系统地涉及扰动的假定机制调节肌动蛋白细胞骨架的变化，在lamellipodial突起的动力学。这些研究有望显示细胞运动如何受到物理线索的影响。在目标2下，建议在粘附、信号传导和细胞骨架动力学水平上阐明趋触性偏倚的性质。据推测，F-肌动蛋白束/丝状伪足直接，和板状伪足传播，haptotactic探索。一个相关的假设是，趋触偏倚（比较向上与向下梯度）表现为信号传导和/或细胞骨架动力学的差异，通过粘附整合。它提出了分析动态的粘附，信号，和细胞骨架结构迁移过程中的haptotactic梯度产生的微流体装置。这些研究将表征趋触性过程中的前沿运动动力学，这是一种与趋化性不同的定向迁移模式。根据目标3，拟议的工作将阐明硬脊膜扩张的分子和生物物理决定因素，并定义触觉和硬脊膜扩张之间共同或不同的特征。将生成具有交联梯度的水凝胶，并将其与牵引力显微镜（一种用于测量弹性凝胶中局部应力的方法）相结合。这些研究将测试富含fascin的丝状伪足作为引导durotactic迁移的感觉细胞器的作用。结果将进一步阐明细胞突起，局灶性粘附动力学和durotaxing细胞施加的牵引力之间的关系。该项目的跨学科性质为两名研究生的培训提供了一个独特的环境，他们将从事促进科学传播的研究和外联活动。

Cell migration is an important step in many processes in the body, such as the healing of wounds and the response to infection. The cells are in contact with a matrix that allows them to crawl and explore. To do so, cells must actively sense matrix properties by touch and feel. An overarching goal of this collaborative project is to unify or distinguish cell sensing of the quantity (stickiness by touch) versus quality (stiffness by feel) of contacts with the matrix. The multidisciplinary research will be integrated with graduate student training, teaching of engineering courses, and outreach that will foster science communication.Cell migration is a fundamental process in tissue development and homeostasis. The molecular determinants of chemotaxis, cell migration biased by a gradient of a soluble attractant, are reasonably well understood, and a paradigm has emerged characterizing certain intracellular signaling pathways as a molecular "compass". In contrast, haptotaxis and durotaxis, migration directed by gradients of immobilized ligand density and of mechanical stiffness, respectively, are poorly understood and require a new paradigm, considering that cells must actively encounter and respond to physical cues (i.e., by "feel") rather than by passive sensing of diffusible ligands. Under Objective 1, the aim is to define feedback mechanisms that control coupled cytoskeletal dynamics in migrating cells. It is hypothesized that adhesion-mediated signaling pathways control the duration of lamellipodial protrusion and are spatially coordinated by F-actin bundles. High-resolution, live-cell microscopy and molecular perturbations of the putative signaling and cytoskeletal processes will be applied to systematically relate perturbations of putative mechanisms regulating the actin cytoskeleton to changes in lamellipodial protrusion dynamics. These studies are expected to show how cell motility is biased by physical cues. Under Objective 2, it is proposed to elucidate the nature of haptotactic bias at the level of adhesion, signaling, and cytoskeletal dynamics. It is hypothesized that F-actin bundles/filopodia direct, and lamellipodia propagate, haptotactic exploration. A related hypothesis is that the haptotactic bias (comparing up- vs. down-gradient) manifests as differences in signaling and/or cytoskeletal dynamics, integrated by adhesions. It is proposed to analyze the dynamics of adhesion, signaling, and cytoskeletal structures during migration on haptotactic gradients generated using microfluidic devices. These studies will characterize leading-edge motility dynamics during haptotaxis, a poorly understood mode of directed migration that is distinct from chemotaxis. Under Objective 3, the proposed work will elucidate the molecular and biophysical determinants of durotaxis and define features that are common or distinct between haptotaxis and durotaxis. Hydrogels with gradients of crosslinking will be generated and integrated with traction force microscopy, a method for measuring local stress in an elastic gel. These studies will test the role of fascin-rich filopodia as sensory organelles guiding durotactic migration. The results will further elucidate the relationship between cell protrusion, focal adhesion dynamics, and traction force exerted by durotaxing cells. The interdisciplinary nature of the project offers a unique environment for the training of two graduate students, who will be engaged in research and an outreach activity fostering science communication.