Fig. 3

SB-699551 signals via canonical Gα_i/o coupling and through the PI3K/AKT/mTOR pathway. a Schematic outlining the procedure followed for the PPA. b The dot blots from X-ray film that correspond to the most notable phosphoproteins whose abundance changed after treatment with SB-699551. The full-length X-ray film is shown in Supplementary Figure 2. c Mean pixel densities of candidate phosphoproteins in MCF-7 tumor cells grown under each condition. d Western blotting validation of temporal phosphoproteomic changes in the MCF-7 and MDA-MB-157 breast tumor cell lines for up to a 60-min exposure to the vehicle (0.1% DMSO) or SB-699551 (SB; 4 μM). Phosphoprotein abundance (pixel density) was calculated by the ratio of phosphoprotein to that of the total protein, each normalized to an alpha (α)-Tubulin loading control. e Heatmap showing fold increase (green) or decrease (red) of phosphoprotein abundance in SB-699551-treated cells relative to those treated with the vehicle. f Western blotting validation of phosphoproteomic changes in MCF-7 and MDA-MB-157 breast tumor cells lines after a 24-h exposure to the vehicle (V) or SB-699551 (3 μM, 4 μM or 5 μM). g Relative phosphoprotein abundance after a 24-h exposure of MCF-7 and MDA-MB-157 cells to SB-699551. Western blot frames (outlined in black) were cropped; full-length blots are shown in Supplementary Figure 8. Blots were imaged using the LI-COR Biosciences Odyssey Platform and analysed using ImageStudio software