Correlation of cellular traction forces and dissociation kinetics of adhesive protein zyxin by means of multidimensional microscopy

LORENA SIGAUT; LÍA I PIETRASANTA; MICAELA BIANCHI

Conferencia; 27th IUPAP International Conference on Statistical Physics, StatPhys27; 2019

Cells exert traction forces on the extracellular matrix to which they are adhered through the formation of focal adhesion. Spatial-temporal regulation of traction forces is crucial in cell adhesion, migration, cellular division and remodeling of the extracellular matrix. For quantitative measurements of the direction and magnitude of cellular traction force are required a combination of cell experiments, image processing and force reconstruction based on elasticity theory. Traction Force Microscopy (TFM) is a unique tool to assess the changes in the average traction force exerted by individual cells. TFM utilizes elastic substrates and optical microscopy to track substrate surface displacements through the spatial correlation of fluorescent particles embedded in the substrate. In order to study the effects of the elasticity of the substrate in the generation of forces and in the binding kinetics of one of the protein components of the focal adhesion, we quantify the traction forces exerted by mouse epithelial mammalian living cells cultivated on polyacrylamide substrates of different elasticity coated with fibronectin. Also, in these conditions, we characterize the dissociation kinetics of zyxin, one of the focal adhesion proteins postulated as mechanosensor. From TFM's experiments we were able to reconstruct traction forces maps of cells cultivated on different substrates, observing that the magnitude of the force generated by the cells increases when they are grown on more rigid substrates. On the other hand, the study of the unbinding kinetics of zyxin by means of the fluorescence recovery after photobleaching (FRAP) revealed a decrease in the dissociation constant of zyxin from the focal adhesion, as the rigidity of the substrate increases. Furthermore, with the aim to explore the correlation between the unbinding kinetic of zyxin and the magnitude of the traction forces exerted by a focal adhesion, we performed combined experiments of TFM and FRAP in a single cell. The analysis of these experiments would indicate a correlation ?at the focal adhesion level- between the unbinding kinetic of zyxin and the magnitude of the traction forces exerted by focal adhesions: suggesting that focal adhesions that exert greater traction forces, present smaller dissociation constant for zyxin.