New Approaches For Identifying Critical Integrin-Mediated Signaling Events In Directed Cell Migration

Abstract

Dynamic regulation of cell adhesion on extracellular matrix (ECM) proteins plays vital roles in the establishment and maintenance of tissue structure. More specifically, the spatial coordination of cell-ECM adhesion foci, or focal contacts, influences direction-sensing in motile cells for a wide range of biological processes in metazoan life. Currently available evidence suggests that positive feedback loops between a few canonical signaling pathways focuses rapid turnover of focal contacts at the leading edge, where broad protrusions guide cells in one direction or another. For instance, Rho GTPases regulate the localization of specific actin cytoskeletal rearrangements, phosphatidylinositol 3-kinase (PI3K) -generated second messengers facilitate polarization of related signaling molecules, and focal adhesion kinase (FAK) may potentially act as a mediator of these distinct signaling pathways. This dissertation aims to elucidate mechanisms that coordinate these signaling pathways, with acknowledgement of the limitations of current methods in studying cell migration (Chapter 1). To address questions relating to the role of certain signaling molecules in the maintenance of direction in migration, novel methods for depositing gradients of ECM molecules have been established, including one novel technique incorporating microfluidics, which is amenable to live cell tracking via standard microscopy (Chapter 2). This microfluidic device enables the relative impact of the overexpression of various key proteins to be examined. The subsequent study suggests the role of FAK in facilitating maintained directional migration, also described as persistent migration (Chapter 3). These results suggest that FAK is a critical facilitator of direction-sensing and spatial regulation of focal contacts.