No abstract

Cell migration and spreading involve the coordination of membrane trafficking, actomyosin contraction, and modifications to plasma membrane tension and area. The biochemical or biophysical basis for this coordination is however unknown. In this study, we show that during cell spreading, lamellipodia protrusion flattens plasma membrane folds and blebs and, once the plasma membrane area is depleted, there is a temporary increase in membrane tension by over twofold that is followed by activation of exocytosis and myosin contraction. Further, an artificial increase in plasma membrane tension stopped lamellipodia protrusion and activated an exocytotic burst. Subsequent decrease in tension restored spreading with activation of contraction. Conversely, blebbistatin inhibition of actomyosin contraction resulted in an even greater increase in plasma membrane tension and exocytosis activation. This spatiotemporal synchronization indicates that membrane tension is the signal that coordinates membrane trafficking, actomyosin contraction, and plasma membrane area change. We suggest that cells use plasma membrane tension as a global physical parameter to control cell motility.

A key molecular link between cells and the extracellular matrix is the binding between fibronectin and integrins ␣ 5 ␤ 1 and ␣ v ␤ 3 . However, the roles of these different integrins in establishing adhesion remain unclear. We tested the adhesion strength of fibronectin-integrin-cytoskeleton linkages by applying physiological nanonewton forces to fibronectin-coated magnetic beads bound to cells. We report that the clustering of fibronectin domains within 40 nm led to integrin ␣5␤1 recruitment, and increased the ability to sustain force by over six-fold. This force was supported by ␣5␤1 integrin clusters. Importantly, we did not detect a role of either integrin ␣v␤3 or talin 1 or 2 in maintaining adhesion strength. Instead, these molecules enabled the connection to the cytoskeleton and reinforcement in response to an applied force. Thus, high matrix forces are primarily supported by clustered ␣5␤1 integrins, while less stable links to ␣v␤3 integrins initiate mechanotransduction, resulting in reinforcement of integrin-cytoskeleton linkages through talindependent bonds.