Fig. 2

The interactome and phosphorylation status of Gab2 is differentially affected by sorafenib and axitinib. a Differentially SILAC labeled K562tet/Gab2-HA cells were exposed to 1 μg/ml doxycycline (to induce Gab2-HA expression) prior to treatment with either 1 μM imatinib, 10 μM sorafenib or 1 μM axitinib, and DMSO as control, respectively for 4 h. Purified Gab2 protein complexes were combined 1:1:1 and analyzed by LC-MS/MS. A biological replicate with reversed labels was performed and results of replicates correlated well. Protein interactions dependent on inhibitor sensitive phosphorylation sites will be reduced. b Venn diagram of imatinib, sorafenib and axitinib treatment showing TKI-sensitive Gab2 interactors. c/d TKI-sensitive changes in the Gab2 interactome (c) and the phosphorylation of Gab2 (d). Each bar represents an independent experiment (e) K562tet Vector and Gab2 cells were exposed to 1 μg/ml doxycycline prior to the treatment with the indicated inhibitors. Purified Gab2 complexes were analyzed using the indicated antibodies. f Schematic model of TKI action on the Bcr-Abl/Grb2/Gab2 signaling complex. Axitinib acts like imatinib, dasatinb, nilotinib and ponatinib mainly through the Bcr-Abl/Grb2/Gab2 axis, whereas sorafenib seems to act independently and most likely by affecting signaling pathways up- and downstream of Gab2. Due to the effects of axitinib on Gab2 mediated resistance, axitinib might act additionally also on other kinases, similar to sorafenib. g Diagram showing the potency of sorafenib and axitinib in all tested TKI resistances