Analysis of PECAM-1-dependent mechanotransduction in endothelial cells
Final Report Abstract
The ability to sense and respond to mechanical forces is an intrinsic property of cells and fundamental to developmental and physiological processes. Mechanotransduction is also central to a wide range of diseases such as muscular dystrophies, cancer or atherosclerosis. However, the molecular mechanisms of mechanotransduction are largely unknown, because we lack suitable methods to analyze force sensation and force transduction under physiological conditions. During the course of the project, I developed a new microscopic technique to measure and visualize mechanical tension across single proteins in cells. It is based on a fluorescence resonance energy transfer (FRET) biosensor sensitive to mechanical forces in the pi co-Newton range. To evaluate the technique it was applied to vinculin, a protein involved in integrin-dependent mechanotransduction. Interestingly, the sensor revealed a polarized distribution of force across this molecule during cell migration. Vinculin is exposed to high mechanical forces in protruding areas, but under low mechanical forces in retracting parts of the cell. Furthermore, I can show that localization of vinculin and vinculin force transmission are regulated separately. This is surprising, because these processes were expected to be intimately linked. Together, these experiments demonstrate that the tension sensor method allows a more precise analysis of mechanotransduction and can be efficiently used to determine the spatiotemporal distribution of force across single molecules in living cells. Because of its broad applicability this technique will be widely used to address mechano-biological questions in great detail. Indeed, new tension sensor constructs for proteins such as Pecam-1, VE-Cadherin and talin-1 have been developed and will be the basis for my future research on mechanotransduction.
