Fig. 3

TMEM52B ECD-derived peptides suppress EGFR activation by interfering with the interaction between soluble E-cadherin and EGFR. a Cells were treated for 48 h with recombinant extracellular E-cadherin protein fused to Fc (sE-cad-Fc; 25 µg/mL) and with the peptides. Recombinant Fc protein (25 µg/mL) was used as a negative control. Cells were lysed prior to immunoblot analysis. Densitometric quantification was performed using GAPDH as a loading control; phosphorylated proteins were normalized against the corresponding total protein. b Co-immunoprecipitation analysis was performed to examine the interaction between EGFR and E-cadherin in HCT-15 cells. c Co-immunoprecipitation was performed using lysates from HCT-15 cells treated with the peptides (100 µg/mL) for 48 h. d E-cadherin-binding activities of the peptides were determined by ELISA. Immunoplate wells were coated with the recombinant human E-cadherin ECD protein (100 ng/well), and then incubated with varying amounts of peptide-Fc fusion proteins. Bound peptide-Fc fusion proteins were detected by using HRP-conjugated anti-human Fc antibodies. Mock Fc protein was used as a negative control