Development of a Ratiometric Tension Sensor Exclusively Responding to Integrin Tension Magnitude in Live Cells

Integrin tensions are critical for cell mechanotransduction. By converting force to fluorescence, molecular tension sensors image integrin tensions in live cells with a high resolution. However, the fluorescence signal intensity results collectively from integrin tension magnitude, tension dwell time, integrin density, sensor accessibility, and so forth, making it highly challenging to specifically monitor the molecular force level of integrin tensions. Here, a ratiometric tension sensor (RTS) was developed to exclusively monitor the integrin tension magnitude. The RTS consists of two tension-sensing units that are coupled in series and always subject to the same integrin tension. These two units are activated by tension to fluoresce in separate spectra and with different activation rates. The ratio of their activation probabilities, reported by fluorescence ratiometric measurement, is solely determined by the local integrin tension magnitude. RTS responded sensitively to the variation of integrin tension magnitude in platelets and focal adhesions due to different cell plating times, actomyosin inhibition, or vinculin knockout. At last, RTS confirmed that integrin tension magnitude in platelets and focal adhesions decreases monotonically with the substrate rigidity, verifying the rigidity dependence of integrin tensions in live cells and suggesting that integrin tension magnitude could be a key biomechanical factor in cell rigidity sensing.