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 ?1 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.
Our laboratory has defined a set of changes in the collagen structure surrounding mammary tumors as they progress. These changes manifest in predictable ways, such that 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, perpendicular collagen alignment predicts poor outcome in breast cancer patients. Likely the reason that aligned collagen predicts outcome is that collagen alignment facilitates cell migration, which allows cells to spread throughout the body. In this proposal, we will investigate the molecular mechanisms by which breast cancer cells are able to recognize and make use of aligned collagen for efficient invasion leading to metastasis. We will make use of mouse models and human breast cancer cells to understand the mechanisms involved. The long term impact of this proposal is the potential to identify means by which we can block these mechanisms, and the identification of therapeutic targets for future studies.
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