CdGAP is required for transforming growth factor β- and Neu/ErbB-2-induced breast cancer cell motility and invasion

RhoA, Rac1 and Cdc42, the best-characterized members of the Rho family of small GTPases, are critical regulators of many cellular activities. Cdc42 GTPase-activating protein (CdGAP) is a serine-and proline-rich RhoGAP protein showing GAP activity against both Cdc42 and Rac1 but not RhoA. CdGAP is phosphorylated downstream of the MEK-ERK (extracellular signal-regulated kinase) pathway in response to serum and is required for normal cell spreading and polarized lamellipodia formation. In this study, we found that CdGAP protein and mRNA levels are highly increased in mammary tumor explants expressing an activated Neu/ErbB-2 (Neu-NT) receptor. In response to transforming growth factor-beta (TGF beta) stimulation, Neu-NT-expressing mammary tumor explants demonstrate a clear induction in cell motility and invasion. We show that downregulation of CdGAP expression by small interfering RNA abrogates the ability of TGF beta to induce cell motility and invasion of Neu-NT-expressing mammary tumor explants. However, it has no effect on TGF beta-mediated cell adhesion on type 1 collagen and fibronectin. Interestingly, protein expression of E-Cadherin is highly increased in Neu-NT-expressing mammary tumor explants depleted of CdGAP. In addition, complete loss of E-Cadherin expression is not observed in CdGAP-depleted cells during TGF beta-mediated epithelial to mesenchymal transition. Downregulation of the CdGAP expression also decreases cell proliferation of Neu-NT-expressing mammary tumor explants independently of TGF beta. Rescue analysis using re-expression of various CdGAP deletion-mutant proteins revealed that the proline-rich domain (PRD) but not the GAP domain of CdGAP is essential to mediate TGF beta-induced cell motility and invasion. Finally, we found that TGF beta induces the expression and phosphorylation of CdGAP in mammary epithelial NMuMG cells. Taken together, these studies identify CdGAP as a novel molecular target in TGF beta signaling and implicate CdGAP as an essential component in the synergistic interaction between TGF beta and Neu/ErbB-2 signaling pathways in breast cancer cells. Oncogene (2011) 30, 1032-1045; doi:10.1038/onc.2010.477; published online 1 November 2010