Figure 3.

TPα-R-stimulated interactions between fluorophore-labeled Gα₁₃ and LARG were resolved by single-cell FRET imaging. A, C) HEK293T cells transfected with 0.5 µg TPα-R, 1 µg Gα₁₃-mTur2, 0.5 µg Gβ, 0.2 Gγ, and 1 µg N-terminally YFP-labeled LARG showed a rapid increase in FRET upon stimulation with 10 nM U-46619 [representative cell in A, and mean ± sem of 30 cells (black trace) in C]. B) YFP-LARG localized mainly to the cytosol, as visualized by confocal microscopy from HEK293T cells transfected with 0.5 µg TPα-R, 1 µg Gα₁₃, 0.5 µg Gβ, 0.2 Gγ, and 1 µg YFP-LARG. C) FRET change upon stimulation with U-46619 was reduced if Gα₁₃ and membrane-bound CFP were transfected instead of Gα₁₃-mTur2 (red trace; mean ± sem of n = 13). D) Also LARG with a YFP inserted between the RH and the DH/PH domain (LARG-insYFP) showed an increase in FRET upon activation of TPα-R (means ± sem; n = 9).