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

合作研究：细胞外基质引导的细胞迁移中“感觉”梯度感知的机制

• 批准号: 1706087
• 负责人: Jason Haugh
• 金额: $ 28.53万
• 依托单位: North Carolina State University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-01 至 2021-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1706087&HistoricalAwards=false
• 关键词: Collaborative; Research; Mechanisms; Gradient; Sensing

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.

细胞迁移是人体许多过程的重要步骤，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，细胞必须通过触觉和感觉积极地感知基质的特性。这个合作项目的首要目标是统一或区分细胞对与矩阵接触的数量（通过触摸产生的粘性）和质量（通过感觉产生的硬度）的感知。多学科研究将与研究生培训、工程课程教学以及促进科学交流的外展活动相结合。细胞迁移是组织发育和体内平衡的一个基本过程。趋化性的分子决定因素，即受可溶性引诱剂梯度影响的细胞迁移，已经被很好地理解，并且出现了一种范式，将某些细胞内信号通路表征为分子“指南针”。相比之下，由于细胞必须主动地遇到和响应物理线索（即通过“感觉”）而不是被动地感知扩散配体，因此人们对附着性和硬性（分别由固定配体密度梯度和机械刚度梯度引导的迁移）知之甚少，需要一个新的范式。在目标1下，目的是定义控制迁移细胞中耦合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板足突的持续时间，并由f -肌动蛋白束在空间上协调。高分辨率，活细胞显微镜和分子扰动的假设信号和细胞骨架过程将被应用于系统地将调节肌动蛋白细胞骨架的假设机制的扰动与板足突动力学的变化联系起来。这些研究有望揭示细胞运动是如何受到生理信号的影响的。在目标2中，作者提出了在粘附、信号传导和细胞骨架动力学水平上阐明触致性偏向的本质。假设f -肌动蛋白束/丝状足直接，板足繁殖，触致探索。一个相关的假设是，触致偏倚（比较上下梯度）表现为信号传导和/或细胞骨架动力学的差异，并与粘附结合在一起。在微流体装置产生的触致梯度迁移过程中，提出了分析粘附、信号传导和细胞骨架结构的动力学。这些研究将描述触致性过程中的前沿运动动力学，这是一种鲜为人知的定向迁移模式，与趋化性不同。在目标3下，提出的工作将阐明趋向性的分子和生物物理决定因素，并定义趋向性和趋向性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相结合，牵引力显微镜是一种测量弹性凝胶局部应力的方法。这些研究将测试富筋膜蛋白丝状足作为感觉细胞器指导多径性迁移的作用。研究结果将进一步阐明细胞突起、黏附动力学和黏附力之间的关系。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，他们将从事研究和促进科学传播的推广活动。