Mechanisms Of Cell Migration On 3D Aligned Matrices

 描述（由申请人提供）：细胞迁移是一个基本过程，已对细胞在包被的2D表面上迁移进行了充分研究，但在3D基质或体内背景下了解甚少。理解细胞迁移是重要的，因为它是导致转移的入侵所必需的，也是涉及胚胎发育、组织和伤口修复的基本过程。几十年来，细胞生物学家已经在2D表面上表征了细胞迁移;最近的努力正在转向理解体内3D细胞迁移的更困难的任务。为了阐明这个问题，我们联合收割机将Keely实验室的能力与Hahn实验室结合起来，Keely实验室长期专注于3D癌细胞迁移的生物学和成像，Hahn实验室开发了一系列技术来可视化和操纵活细胞中的信号活动。Keely实验室已经发现并表征了乳腺肿瘤周围胶原结构的一系列变化，随着它们的进展-有越来越多的成束，排列的胶原纤维沉积，这些纤维的重组垂直于肿瘤/间质边界。值得注意的是，胶原蛋白排列促进细胞迁移、转移，并导致患者的不良结局。最近，我们发现细胞在对齐的基质上更持久，并且这种持久性与有限的侧向突起有关。我们假设Rho介导的收缩性是沿着基质排列的轴组织的，并且双轴力稳定限制侧向突起的侧向粘连。相比之下，前缘处的较低应变允许新生动态粘附，这促进向前突起。此外，我们假设，与Src和Rho家族GTP酶相关的信号通路在空间上受到这些侧向和正向粘附的调节和反馈，从而使细胞能够读出其微环境的对齐和地形线索。我们的具体目标是测试这些想法：目的1）确定用于调节双轴应变，粘附和突起的Rho家族GTdR信号传导的空间和时间动力学。目的2）确定β1整合素在将排列转化为有组织的细胞极性和突起中的作用。目的3）探讨Rho GT3和粘附信号通路在肿瘤细胞迁移和转移中的作用。

 DESCRIPTION (provided by applicant): Cell migration is a fundamental process that has been well studied for cells migrating on coated, 2D surfaces, but is poorly understood in the context of a 3D matrix or in vivo. Understanding cell migration is significant, as it is essential o invasion leading to metastasis, as well as a fundamental process involved in embryonic development, tissue organization, and wound repair. For several decades, cell biologists have characterized cell migration on 2D surfaces; recent efforts are turning to the significantly more difficult task of understanding 3D cell migration in vivo. To shed light on this question, we combine here the capabilities of the Keely lab, long focused on the biology and imaging of 3D cancer cell migration, with the Hahn lab, which has developed a range of techniques to visualize and manipulate signaling activity in live cells. The Keely laboratory has discovered and characterized a set of changes in the collagen structure surrounding mammary tumors as they progress -- there is an increasing deposition of bundled, aligned collagen fibers and a reorganization of these fibers to be perpendicular to the tumor/stromal boundary. Notably, collagen alignment facilitates cell migration, metastasis, and leads to poor outcome in patients. Recently, we found that cells are more persistent on an aligned matrix, and that this persistence is associated with limited lateral protrusions. We hypothesize that Rho-mediated contractility is organized along the axis of matrix alignment, and that biaxial forces stabilize lateral adhesions that limit lateral protrusions. In contrast, the lower strain at the leading edge allows nascent dynamic adhesions, which promote forward protrusions. Moreover, we hypothesize that signaling pathways linked to Src and Rho family GTPases are spatially regulated by and feed back to these lateral and forward adhesions to allow cells to read out the alignment and topographic cues of their microenvironment. Our specific aims to test these ideas: Aim 1) Determine the spatial and temporal dynamics of Rho family GTPase signaling used to regulate biaxial strain, adhesions, and protrusions. Aim 2) Determine β1 integrin's role in translating alignment into organized cell polarity and protrusions. Aim 3) Determine the role of Rho GTPase and adhesion signaling pathways during migration and metastasis in vivo.