Overexpression of GRB2 Enhances Epithelial to Mesenchymal Transition of A549 Cells by Upregulating SNAIL Expression

GRB2 is an adaptor protein which interacts with phosphorylated TGF-beta receptor and is critical for mammary tumour growth. We found that TGF-beta 1-induced EMT increased GRB2 expression in A549 cells (non-small cell lung cancer). Overexpression of GRB2 (A549(GRB2)) enhanced cell invasion while knocking down GRB2 (A549(GRB2KD)) reduced cell migration and invasion, probably due to increased vinculin and reduced Paxillin patches in A549(GRB2KD) cell. TGF-beta 1-induced EMT was more pronounced in A549(GRB2) cells and attenuated in A549(GRB2KD) cells. This could be due to the reduced expression of E-cadherin in A549(GRB2) and increased expression of E-cadherin in A549(GRB2KD) cells, even before TGF-beta 1 stimulation. Expression of SNAIL was elevated in A549(GRB2) cells and was further enhanced by TGF-beta 1 stimulation, suggesting that GRB2 down-regulates E-cadherin by enhancing the expression of SNAIL. The N-SH3 domain of GRB2 was critical for suppressing E-cadherin expression, while the C-SH3 domain of GRB2 mediating interaction with proteins such as N-WASP was critical for promoting invasion, and the SH2 domain was critical for suppressing E-cadherin expression and invasion. Thus, our data suggests that GRB2 enhances EMT by suppressing E-cadherin expression and promoting invasion probably through N-WASP to promote metastasis.