Force meets chemistry: Analysis of mechanochemical conversion in focal adhesions using fluorescence recovery after photobleaching

Abstract

Mechanotransduction—the process by which mechanical forces are converted into changes of intracellular biochemistry—is critical for normal cell and tissue function. Integrins facilitate mechanochemical conversion by transferring physical forces from the extracellular matrix, across the cell surface, and to cytoskeletal‐associated proteins within focal adhesions. It is likely that force alters biochemistry at these sites by altering molecular binding affinities of a subset of focal adhesion proteins, but this has been difficult to quantify within living cells. Here, we describe how the fluorescence recovery after photobleaching (FRAP) technique can be adapted and used in conjunction with mathematical models to directly measure force‐dependent alterations in molecular binding and unbinding rate constants of individual focal adhesion proteins in situ. We review these recent findings, and discuss the strengths and limitations of this approach for analysis of mechanochemical signaling in focal adhesions and other cellular structures. The ability to quantify molecular binding rate constants in the physical context of the living cytoplasm should provide new insight into the molecular basis of cellular mechanotransduction. It also may facilitate future efforts to bridge biological experimentation and mathematical modeling in our quest for a systems biology level description of cell regulation. J. Cell. Biochem. 97: 1175–1183, 2006.