Fig. 2.

LIPUS stimulates rapid GTPase activation and endocytosis. (A,B) B16 cells expressing (A) Rac1 or (B) Cdc42 FRET-based activity reporters were imaged before, during and after LIPUS stimulation. Images and quantifications of ratiometric FRET (normalized between 0 and 100 over time) analysis show that LIPUS stimulation leads to rapid activation of both GTPases, and that elevated GTPase activation persists after the stimulation ends. Scale bars: 10 µm. Data are mean±s.e.m. from three independent experiments; n=26 in A and n=17 in B. (C) Pretreatment of B16 cells with the Rac1 inhibitor EHT1864 (10 µM), but not with the Cdc42 inhibitor ML141 (10 µM), blocked LIPUS-induced CDR formation. Measurements are from the indicated numbers of XY positions, representative of three independent experiments. (D) Pretreatment with EHT1864 (10 µM) also blocked LIPUS-induced motility increases in both B16 and MEF cells. Cells were tracked over 16 h to assess cell motility (results are representative of three independent experiments and are presented as described in the Materials and Methods). *P<0.05, ***P<0.001.