HER2/ErbB2-induced Breast Cancer Cell Migration and Invasion Require p120 Catenin Activation of Rac1 and Cdc42

Abstract

Breast cancers that overexpress the receptor tyrosine kinase ErbB2/HER2/Neu result in poor patient outcome because of extensive metastatic progression. Herein, we delineate a molecular mechanism that may govern this malignant phenotype. ErbB2 induction of migration requires activation of the small GTPases Rac1 and Cdc42. The ability of ErbB2 to activate these small GTPases necessitated expression of p120 catenin, which is itself up-regulated by signaling through ErbB2 and the tyrosine kinase Src. Silencing p120 in ErbB2-dependent breast cancer cell lines dramatically inhibited migration and invasion as well as activation of Rac1 and Cdc42. In contrast, overexpression of constitutively active mutants of these GTPases reversed the effects of p120 silencing. Lastly, ectopic expression of p120 promoted migration and invasion and potentiated metastatic progression of a weakly metastatic, ErbB2-dependent breast cancer cell line. These results suggest that p120 acts as an obligate intermediate between ErbB2 and Rac1/Cdc42 to modulate the metastatic potential of breast cancer cells.HER2/Neu/ErbB2 (epidermal growth factor receptor 2) is a member of the epidermal growth factor receptor (EGFR) 2 family that is amplified and overexpressed in 20 -30% of breast cancers. HER2/ErbB2-positive breast cancer yields a poor patient prognosis because of a high incidence of metastases and intrinsic resistance to endocrine and conventional chemotherapy (1). ErbB2 is the preferred heterodimerization partner for EGFR, ErbB3, and ErbB4 (2). The large range of downstream signaling targets induced by ErbB2-containing heterodimers permits this receptor and its partners to modulate a wide array of cellular processes. Of these, the nonreceptor tyrosine kinase Src, PI3K, and MAPK pathways are the most studied in mediating ErbB2 responsiveness, which includes increased proliferation, survival, motility, and invasion (3).ErbB2-induced cell migration and invasion, and hence metastatic potential, requires activation of Src and its downstream signaling pathways (4). Src also positively provides feedback and stabilizes competent ErbB2/ErbB3 heterocomplexes to enhance PI3K/Akt signaling (5). ErbB2 overexpression in MDA-MB-435 cells enhances formation of metastases by increasing Src synthesis and decreasing its degradation by calpain (6). Similarly, signaling from ErbB2 to Akt is necessary to promote metastatic potential because attenuation of the Akt pathway by pharmacological inhibitors or RNAi decreases the formation of metastases by the ErbB2-dependent 21T breast cancer cell line (7).

Cdc42 GTPase and Rac1 GTPase act downstream of p120 catenin and require GTP exchange during gastrulation of zebrafish mesoderm

“…Note also that 3 and 5 pg of DN-Cdc42 mRNA alone dramatically impacted the morphology and decreased the survival of the embryos to 20-to 24-somite stages. These results suggest not only that Cdc42 is an essential downstream effecter of p120 catenin d1 for normal cell migration, but also that the ability of Cdc42 to initiate directional polarity of migrating cells by localized GDP to GTP exchange (Etteinne-Manneville, 2004, 2008Etteinne-Manneville and Hall 2001;Yang et al, 2006;Johnson et al, 2010) or to cycle between its active GTP-bound and inactive GDPbound states is an essential feature in this signaling pathway.…”

“…Again we compared the co-injection of p120-catenin d1 splice-MO with wild-type Rac1 mRNA (WTRac1), constitutively active Rac1 (CARac1), or dominant-negative Rac1 (DN-Rac1). CA-Rac1 has typically been used in Xenopus embryo and cell culture experiments (Fang et al, 2004;Deplazes et al, 2009;Johnson et al, 2010). Figure 7A shows that as little as 1 pg of WT-Rac1 mRNA rescued most of the embryos injected at the 1-cell stage with p120 catenin d1 splice-MO.…”

“…RhoA is a downstream effecter of p120 catenin signaling Deplazes et al, 2009;Johnson et al, 2010). In Xenopus embryos, dominant-negative RhoA partially rescues the knockdown of p120 catenin (Fang et al, 2004).…”

“…Our observations that DN-RhoA can rescue the knockdown of p120 catenin d1 show that a release from adhesion also promotes somite cell migration at low concentrations of p120 catenin d1. Since Cdc42 GTPase initiates the polarity of cells (Etteinne-Manneville, 2004, 2008Etteinne-Manneville and Hall, 2001;Yang et al, 2006;Johnson et al, 2010), the inability of CA-Cdc42 GTPase to rescue the knockdown of p120 catenin d1 is not surprising. CACdc42 would compete with endogenous WT-Cdc42 and block the stimulation of actin polymerization by WTCdc42 GTPase.…”

“…ARVCF and p120 catenin d1 diverged from the p120 catenin d2 proteins to bind Rho GTPases during chordate evolution (Zhao et al 2011). Besides stabilizing cadherin, p120 catenin d1 regulates cell motility by activating Cdc42 and Rac1 GTPases and inhibiting RhoA GTPase (Anastasiadis andReynolds., 2000, 2001;Anastasiadis, 2007;Grosheva et al, 2001;Magie et al, 2002;Xiao et al, 2007;McCrea and Park, 2007;Johnson et al, 2010). Consequently, p120 catenin d1 overexpression in cells results in branching filopodia and/or abnormal extension of lamellipodia through the activation of Cdc42 and Rac1 GTPases (Noren et al, 2000).…”