Monitoring in real-time focal adhesion protein dynamics in response to a discrete mechanical stimulus

VON BILDERLING CATALINA; CALDAROLA, MARTÍN; MASIP, MARTÍN E.; BRAGAS, ANDREA V.; PIETRASANTA, LÍA I.

Congreso; XLIII Reunión Anual SAB 2014; 2014

Sociedad Argentina de Biofísica

The adhesion of cells to the extracellular matrix is a hierarchical,force-dependent, multistage process that evolves at severaltemporal scales [1]. An understanding of this complex processrequires a precise measurement of forces and its correlation withprotein responses in living cells. We present a method toquantitatively assess live cell responses to a local and specificmechanical stimulus. Our approach combines atomic forcemicroscopy (AFM) with fluorescence imaging. Using this approach,we evaluated the recruitment of adhesion proteins such as vinculin,FAK and zyxin triggered by applying to live cells forces in the nNregime. We observed the development of a nascent adhesion site,which was evident from the accumulation of vinculin at the positionwhere the force was applied. In addition, we quantified therecruitment time for FAK in the formation of a new adhesion site,and analyzed the zyxin spatial distribution remodeling in maturefocal adhesion as a function of the applied force. We havedemonstrated that this method is a useful tool for the study of avariety of complex biological processes involved in cellularmechanotransduction