Altered regulation of cell migration plays a major role in development of cancer invasiveness. An N-terminally truncated, constitutively active ErbB2 receptor (DNErbB2), which signals in part through the extracellular signal regulated kinase (ERK)-ribosomal S kinase (RSK) pathway, is common in human breast cancer and is associated with increased malignancy and poor prognosis. It was previously reported that DNErbB2 expression in MCF-7 breast cancer cells leads to sustained ERK1/2 activation and loss of epithelial morphology¹. Migration and invasion require extensive reorganization of cytoskeletal elements, including F-actin, cortactin, and ezrin/radixin/moesin (ERM) proteins. In addition, we and others have demonstrated central roles for ion transport, and specifically the Na⁺/H⁺ exchanger NHE1, in cancer cell migration². Here, we investigate the effect of DNErbB2 expression on the migratory properties of MCF-7 cells, focusing on the possible roles of, and interactions between, NHE1 activity and cytoskeletal reorganization. MCF-7 cells expressing DNErbB2 under control of a tetracycline (tet)-off system (DNErbB2 cells) were grown without tet for 48 h to induce DNErbB2 expression. Two controls were employed: uninduced DNErbB2 cells, and MCF-7 cells expressing the corresponding vector without DNErbB2, grown without tet for 48 h. Tet washout induced strong DNErbB2 expression in DNErbB2 cells concomitant with a change from epithelial-like to elongated, fibroblast-like morphology. Control cells remained epithelial in morphology after wash. NHE1 was evenly distributed in the plasma membrane of control cells, but redistributed to lamellipodia-like protrusions after DNErbB2 expression. NHE1 moreover localized strongly to mm-diameter, round structures between and in the cells which also showed very high levels of F-actin, cortactin, and ERM proteins, which, interestingly, interact directly with NHE1. Localization of phosphorylated, active ERM proteins to these structures increased dramatically after DNErbB2 expression, concomitant with an increase in the net cellular level of these proteins. In 2D migration assays, the migration velocity of serum-starved cells on collagen IV was increased several-fold by DNErbB2 expression. The NHE1 inhibitor EIPA reduced the velocity of control cells by 50%. In contrast, the velocity of DNErbB2 expressing cells was doubled by EIPA, whereas it was decreased by 50% by the RSK inhibitor SL0101. In conclusion, DNErbB2 expression in MCF-7 cells increased migration velocity, at least in part through RSK signaling. Moreover, DNErbB2 expression elicited cytoskeletal reorganization and dramatically altered the role of NHE1 in migration from stimulatory to inhibitory, an effect we hypothesize may reflect NHE1-dependent effects on cell adhesion/invasion.

1. Egeblad M et al. 2001 94:185–91; 2. Schwab A et al. 2007 453:421–32