Determining Integrin Molecular Tension for the Recruitment and the Activation of Focal Adhesion Kinase

Abstract

Focal adhesion kinase (FAK), a member of protein tyrosine kinase family, is an important mediator of signals transduced by integrins (major mechanosensitive transmembrane receptors transmitting cellular forces at cell matrix interface). FAK regulates various cellular functions such as cell adhesion, proliferation, differentiation and spreading. Although the biochemical role of FAK in integrin signaling is relatively more understood, the biomechanical processes for FAK's recruitment and activation by integrins remains less clear. To study the mechanobiology of FAK, Our group adopted two molecular tension tools, tension gauge tether (TGT) and integrative tension sensor (ITS), both having a designable tension threshold, selectable in the range of 10-60 pN. TGT globally knocks down integrin tensions in live cells to a designed force level, while ITS visualizes integrin tension by fluorescence imaging with high resolution. With TGT, ITS and a FRET-based biosensor reporting FAK activation, we manipulated and mapped integrin tension in live cells, and studied the consequent FAK aggregation and activation. ITS and FAK live cell imaging revealed that integrin tension higher than 12 pN was produced after FAK aggregation, indicating that FAK recruitment to focal adhesions does not require integrin tension higher than 12 pN. TGT and FAK biosensor revealed that FAK was not activated on TGT surfaces with tension tolerance (Ttol) of 12, 23 and 33 pN, but on the surfaces with Ttol of 43 pN and 54 pN. These results suggest that FAK activation by integrin signaling requires integrin tension higher than 33 pN. Overall, our study revealed that FAK recruitment and FAK activation have distinct requirement for integrin tensions at separate force levels.