Mechanisms Of Cell Migration On 3D Aligned Matrices

 描述(申请人提供)：细胞迁移是细胞在涂覆的2D表面上迁移的一个基本过程，已经得到了很好的研究，但在3D基质或体内的背景下了解较少。了解细胞迁移具有重要意义，因为它是侵袭导致转移的关键，也是胚胎发育、组织组织和伤口修复的基本过程。几十年来，细胞生物学家一直在描述细胞在2D表面上的迁移；最近的努力转向了更困难的任务，即了解3D细胞在体内的迁移。为了阐明这个问题，我们在这里将凯利实验室和哈恩实验室的能力结合起来，前者长期专注于3D癌细胞迁移的生物学和成像，后者开发了一系列技术来可视化和操纵活细胞中的信号活动。基利实验室发现并表征了乳腺肿瘤发展过程中周围的胶原蛋白结构的一系列变化--成束排列的胶原蛋白纤维越来越多地沉积，这些纤维的重组垂直于肿瘤/基质边界。值得注意的是，胶原蛋白的排列促进了细胞的迁移和转移，并导致患者预后不良。最近，我们发现细胞在排列的基质上更持久，这种持久性与有限的侧向突起有关。我们假设Rho介导的收缩能力是沿着基质排列的轴线组织的，并且双轴力稳定了限制侧向突出的侧向粘连。相反，前缘的较低张力允许新生的动态粘连，这促进了向前突出。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向前粘连的调节并反馈到这些粘连，从而允许细胞读出其微环境的排列和地形线索。我们的具体目的是验证这些想法：目的1)确定Rho家族GTP酶信号用于调节双轴应变、粘连和突起的空间和时间动力学。目的2)确定β-1整合素在将序列转换为有组织的细胞极性和突起中的作用。目的3)探讨Rho GTP酶和黏附信号通路在体内迁移转移中的作用。

 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.