Molecular Dynamics of Mechanosensing and Mechanotransduction at the Focal Adhesions

Focal adhesions are critical to cellular processes such as cell migration and cell-substrate adhesion. Focal adhesion formation can be mechanically regulated: forces either from outside the cell or from contracting actin filaments can induce rapid growth and maturation of the focal adhesions. One hypothesis explored here contends that force-induced focal adhesion formation results from mechanosensing by individual protein components. Molecular dynamics computational simulations are developed to evaluate mechanosensing by talin and vinculin. In Part 1 of this dissertation, two force-induced conformational changes are suggested for talin's activation: (i) the cryptic vinculin-binding sites (VBS) can be activated by stretch of an individual talin rod domain, and (ii) talin can adopt multiple dimer orientations in response to forces applied from outside the cell. The mechanisms of vinculin activation are then explored in Part 2. The trajectory of a vinculin conformational changes that would render it activated is predicted along with the structure of an activated vinculin. Domain 1 (D1) is predicted to separate from the vinculin tail (Vt) during activation. In this context, the PIP2 from the cell membrane is shown to preferentially bind basic residues on the vinculin surface and recruit vinculin to membrane proximal regions, potentially allowing for vinculin phosphorylation. The impact of phosphorylation on the vinculin structure is simulated and it is suggested that phosphorylation could prime vinculin for activation by reducing the strength of inter-domain interactions stabilizing the auto-inhibited vinculin conformation. In Part 3 of this dissertation, the interaction of activated vinculin with its binding partners is simulated. It is demonstrated that a talin VBS can only link vinculin prior to activation but can completely bind vinculin following activation. Vinculin activation by movement at D1 is shown to be necessary and sufficient for linking vinculin to actin filaments. Furthermore, simulation of F-actin caping by vinculin suggests that a second vinculin conformational change, releasing Vt from all vinculin head domains, facilitates effective capping of the actin filament. Three vinculinbinding sites on F-actin are predicted. These simulations demonstrate that indeed both talin and vinculin can exhibit molecular mechanosensing