Figure 6

Rac3 regulates TGFβ1-induced E-cadherin downregulation - A. OE19 cells were treated with TGFβ1 (0–10.0 ng/ml) for 4 days or OE33 cells were treated with TGFβ1 (5 ng/ml) for 4 days, and then the cell lysates were analyzed by immunoblotting with E-cadherin and β-actin antibodies. E-cadherin levels were quantified by Image J software. B. OE19 cells were transfected with V5 tagged Rac3 inactive form (Rac3N17-V5) plasmid for 24 h followed by treatment of TGF-β1 (0–5 ng/ml) for 4 days. The cell lysates were analyzed by immunoblotting with E-cadherin, V5 tag, and β-actin antibodies. E-cadherin levels were quantified by Image J software. C. OE19 cells were transfected with Cont shRNA or Rac3 shRNA plasmid for 48 h followed by the treatment with TGF-β1 (0–5 ng/ml) for 4 days. The cell lysates were analyzed by immunoblotting with E-cadherin, Rac3, and β-actin antibodies. E-cadherin levels were quantified by Image J software. D. OE19 cells were transfected with Rac3-V5 plasmid for 48 h and then cell lysates were analyzed by immunoblotting with Snail, V5 tag, and β-actin antibodies. Snail levels were quantified by Image J software. E. OE19 cells were transfected with Rac3N17-V5 or vector plasmid for 24 h and then incubated with TGFβ1 (5 ng/ml) for 4 days. Cells were fixed and immunostained with E-cadherin (green). Nuclei were stained with DAPI (red). F. Cell length was measured by Image J software. All the blots and images are representative of three independent experiments.