The ability of living cells to sense and respond to the mechanical properties of their surroundings underlies a wide range of cell and tissue engineering applications, particularly in the area of stem cell biology. However, the underlying mechanisms by which cells sense intrinsically mechanical stimuli remain poorly understood, due largely to a lack of tools that measure forces within living cells and organisms. Our laboratory uses genetically encoded molecular tension sensors to measure the mechanical forces experienced by individual proteins in living cells. In this presentation I outline ongoing applications of this technology.
We use a FÃÂ¶rster Resonance Energy Transfer (FRET)-based molecular tension sensor to test the origin and magnitude of tensile forces transmitted through the cytoplasmic domain of E-cadherin, a protein principally responsible for intercellular adhesion in a wide variety of cell types. We find that the cytoskeleton exerts pN-tensile force on E-cadherin and that tension is increased at cell-cell contacts when adhering cells are stretched. While the detailed mechanism remains to be characterized, our observations support the presence of mechanically activated signalling pathways at cell-cell contacts, with likely significance in stem cell and tissue engineering.
A conceptually similar strategy has allowed us to image the mechanical forces experienced by the cytoskeletal protein spectrin in touch-sensitive neurons in the model organism C. elegans. Previous work shows that spectrins are expressed in the nervous system in mammals and disrupted in human brain disorders. We find that neuronal spectrin is under mechanical prestress, and that an intact, prestressed spectrin network is required for touch sensitivity. Based on these and other data, we suggest that spectrin-dependent prestress plays a central and conserved role in the architecture and physiological function of the central and peripheral nervous systems.
Finally, I describe recent advances that allow us to directly visualize cell-generated forces with single-molecule sensitivity. We apply this technique to determine the distribution of forces generated by individual integrins, a class of cell adhesion molecules with prominent roles throughout cell and developmental biology. We observe strikingly complex distributions of tensions within individual integrin-based adhesions. FRET values measured for single probe molecules suggest that relatively modest tensions at the molecular level are sufficient to drive both robust cellular adhesion and matrix sensing.