FIGURE 3:
Gal-8 binds α5β1 integrin and activates FAK in the pathway of EGFR transactivation. (A) GST-Gal-8 pulls down α-5 integrin, β1 integrin, and EGFR (lane 1) from 500 μg of cell extract, which is blocked by lactose (50 mM) (lane 2), indicating dependency of Gal-8/glycan interaction. Lane 3 is empty to avoid spill-off from lane 4 showing the presence of analyzed proteins in 50 μg of total cell extract. (B) MDCK-Gal-8H cells have activated FAK as shown by immunoblot with anti-FAK (Tyr-397). (C) Gal-8 activates EGFR in a FAK-dependent manner. MDCK cells incubated with Gal-8 show an increased level of Tyr phosphorylation of EGFR (pEGFR), which is reduced by FAK inhibitor Y15.