Generation and Evaluation of Transgenic Mice Expressing FRET-based Tension Sensor

<p>In order to visualize forces at molecular level, several tension sensors have been developed based on Förster resonance energy transfer (FRET) and applied in cultured cells. Recently, transgenic mice expressing a tension sensor were also developed to visualize forces at tissue level. However, their fluorescence was so weak that a high-cost FLIM (Fluorescence Lifetime Imaging) system was necessary for observation. Furthermore, the fluorescent protein pair adopted in the sensor cannot be observed with widely-used 488 nm laser system. In a previous study, we developed an actinin tension sensor by inserting a FRET cassette sstFRET-GR into actinin. This sensor can be observed with the 488 nm laser system. In this study, we introduced the gene of this sensor engineered with the Cre/loxP system into the ROSA26 locus of the C57BL/6N mice, and crossbred them with Cre mice to obtain mice expressing the tension sensor. We excised various tissues from them including aorta, heart, tendon, skin, diaphragm, and intestine, and isolated cells from each tissue. The fluorescence was so bright that the change in FRET ratio was observable with a general confocal microscope for all tissues and cells. We performed tensile tests of these tissues and cells, and confirmed that the sensor is working properly. The ratio of FRET ratio change to unit tensile strain was almost similar among tissues, however it is smaller than that in their cells. The present FRET mice may become a powerful tool in mechanobiology of cells and tissues.</p>