Rho GTPase Dynamics in the Regulation of Cellular Signaling and Migration

Cell migration is critical to the development and maintenance of higher organisms, and is required for the patterning of the nervous system, for the development of organs, and for responses to wounds or sites of inflammation. Because cell migration is so widely utilized, it must be very tightly controlled, as is apparent when the process goes awry, such as in cancer cell metastasis or chronic inflammation. Growth factors and other cues mediate the activation of a variety of pathways that induce cell migration. Despite these many pathways, a family of proteins called Rho GTPases are universally engaged to cause changes in the cellular cytoskeleton, leading to cell migration. Because Rho GTPases are so critical to cell migration, yet can be used to mediate many different types of cellular responses, they must be precisely controlled. In most cases, it is the timing and placement of Rho GTPase activity that defines cellular behaviors in response to specific signals. However, tools to investigate the spatiotemporal dynamics of Rho GTPase activity in live cells have only recently been developed. I this work, I characterize the spatial and temporal dynamics of the Rho GTPases in cell migration through the development of sensors for Rho GTPase activity for live cell imaging. I establish the spatial and temporal dynamics of RhoA, Rac1, and Cdc42 at the leading edge of migrating cells, and the role of RhoG in its ability to precisely position and activate Rac1 at the leading edge of cells. This work provides the first thorough characterization of the roles of these GTPases at the leading edge relative to one another and the mechanisms by which they are regulated. This work also demonstrates preliminary studies on the roles of these GTPases during leukocyte transendothelial migration. It is critical to gain an understanding of the mechanisms by which cells control cell migration via the Rho GTPases. Aberrant signaling through the GTPases leads to a variety of disease processes. Thus, a better understanding of normal Rho GTPase signaling will provide a framework for understanding how cell migration goes awry and how it can be potentially treated.Doctor of Philosoph