Fig. 2

Colocalization analysis of the wild type Gαi₃, Gαs and their muteins with various Gβγ dimers with the cell membrane or ER represented as Pearson correlation coefficient (PCC). A-E HEK293 cells transfected with Gαi₃/Gαs-Citrine encoding vector alone or together with different Gβγ vectors without fluorescent protein were imaged after cell membrane staining (Deep Red dye). Images were collected on living cells subsequently for Citrine and Deep Red dye fluorescence. Cells transfected only with Gαi₃/Gαs-Citrine vector represent the membrane localization of the Gαi₃/Gαs without overexpression of the Gβγ dimer. F Similarly, colocalization of the Gαi₃ C3A-Citrine with ER was performed. ER was stained using the CellLight™ system, and images were acquired two days after infection. Data were collected from at least two independent experiments (n = 2) and are presented as mean ± 95% confidence interval (CI). All obtained mean PCC were compared with the WT Gαi₃(A-D)/Gαs (E) PCC with unpaired t-test (black color, * – p < 0.05, ** – p < 0.01, *** – p < 0.005, **** – p < 0.001).Additionally differences in the PCC values between Gαi₃ muteins G2A vs C3A (red color ****—p < 0.001) or Gαi₃ C3A β₁γ₂ vs Gαi₃β₁γ₈ (black color) all with unpaired t-test (F)