The general goal of this renewal proposal is to understand the biomechanical mechanism by which extracellular matrix (ECM) regulates angiogenesis during tumor development, with a specific focus on how physical interactions between capillary endothelial (CE) cells and their ECM adhesions control directional cell motility. During the last grant period, we showed that mechanical changes at the cell-ECM interface govern the direction in which cells move because local variations of physical force distributions dictate where cells will form focal adhesions (FAs) and extend new motile processes when stimulated with soluble motility factors. Analysis of this motility steering mechanism and the mechanism of FA repositioning revealed a central role for transfer of mechanical forces across transmembrane integrin receptors which elicit signaling responses that, in turn, activate additional p1 integrin receptors. Other signaling molecules, including the small GTPases, Rho and Rac, also contribute to the mechanism by which ECM influences FA location, and cells that lack the FA protein paxillin fail to exhibit spatial coupling between FA formation and lamellipodia extension. In separate studies, we discovered that an upstream regulator of Rho, p190RhoGAP, may link cytoskeletal signaling to cell motility and angiogenesis by another mechanism: this Rho inhibitor regulates the activity of the transcription factor TFII-I and thereby controls expression of the vascular endothelial growth factor (VEGF) receptor VEGFR2. Thus, the specific aims include: 1) To explore how stress- dependent activation of (31 integrin and Rho alter focal adhesion position, 2) To determine how focal adhesions govern lamellipodia positioning and directional cell migration, and 3) To analyze how cytoskeletal signaling through p190RhoGAP influences VEGFR2 gene expression. These studies will include in vitro mechanistic experiments as well as in vivo studies in a tumor angiogenesis model to determine the potential clinical relevance of our findings. Understanding the molecular basis of this mechanical signaling response that controls direction migration of capillary blood vessel cells could lead to identification of new molecular targets for therapeutic intervention in virtually all solid cancers that require continuous angiogenesis for their own growth and expansion.
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