Control of directional extension of lamellipodia by extracellular matrix and mechanical forces

Directional cell migration is central to numerous physiological processes including development, wound healing, and the immune response. Although a great deal is known about the molecules that invoke a motile response, the mechanism by which this response becomes spatially localized within the cell to determine the site where motile structures, such as lamellipodia and filopodia, are generated is less well characterized. We hypothesized that the direction of cell motility is governed by mechanical interactions between cells and extracellular matrix (ECM) that are significantly influenced by local variations in ECM topography on the subcellular scale. To test this hypothesis, a model system was developed to mimic the physical constraints experienced by individual cells within tissue. Constraining cell shape on polygonal adhesive islands consistently restricted the generation of motile processes to the cell vertices. Focal adhesions assembled specifically in these corner regions, and colocalized with sites of matrix fibrillogenesis and elevated traction forces on the ECM. We then asked how the mechanical cues from the microfabricated ECM substrates integrate with biochemical signals, specifically through the Rho-family small GTPases, to control the direction of lamellipodia extension. Constitutive activation of Rac GTPase eliminated the corner localization of lamellipodia and focal adhesions. Inhibition of Rho or its effectors eliminated all lamellipodia extension of Rho GTPase but in cells expressing active Rac it rescued the corner pattern of adhesions and motile extensions, pointing to the importance of a balance between Rac and Rho as regulators of focal adhesion formation and lamellipodia positioning.