Synergy between HGF and ErbB2neu promotes epithelial cell invasion

The ErbB-2/neu receptor tyrosine kinase is involved in normal tissue development. However, this receptor has been implicated in the genesis of human breast and renal carcinomas, where ErbB2 is amplified in 20--30% of human breast cancers and correlates with poor prognosis. Using the non-transformed MDCK epithelial cell model, I have established that a deregulated activated ErbB2/Neu receptor (NeuNT) but not overexpression of the wild type (WT) receptor induces cell dispersal and motility, accompanied by the breakdown of cell-cell junctions and E-cadherin internalization, in addition to reorganization of the actin cytoskeleton. This phenotype can be reversed following treatment of the cells with a pharmacological inhibitor of MEK, indicating that MEK dependent pathways are involved in the NeuNT-induced remodeling of cell-cell junctions. In three-dimensional cultures of MDCK cells, NeuNT but not WT ErbB2 triggers a morphogenic response that correlates with recruitment and increased phosphorylation levels of the Shc adapter protein. This demonstrates that the deregulated ErbB2/NT receptor induces a distinct biological response when compared to the wild type receptor and induces the loss of epithelial architecture observed in carcinomas.Invasive morphogenesis downstream from the Met/HGF receptor is modulated through a sustained phosphorylation of the Gab1 docking protein and of downstream kinase (Erk). In contrast, a transient phosphorylation of Gab1 and Erk induced by EGF is not sufficient to promote a morphogenic response. In Chapter III, I demonstrate that NeuNT but not the WT ErbB2 receptor display elevated and sustained levels of Gab1 and Erk phosphorylation which correlates with their ability to promote invasive morphogenesis. In addition, co-immunoprecipitation analyses provide evidence for the recruitment of Gab1 to ErbB2/Neu in a Grb2-dependent and Grb2-independent manner.To identify physiologically relevant factors that synergize with ErbB2, I established that HGF, the Met receptor ligand, promotes the disruption and invasion of NeuNT-induced epithelial structures in three dimensional matrix cultures. Moreover HGF synergizes with NeuNT, enhancing the invasive potential of NeuNT expressing cells ten fold through Matrigel. HGF treatment of NeuNT expressing cells promotes a decrease in E-cadherin protein, and can be blocked or reversed by treatment with the MEK inhibitor, UO126, establishing the involvement of MEK-dependent pathways in this process. These results demonstrate that physiological signals downstream from HGF/Met cooperate with deregulated ErbB2/Neu to enhance the malignant phenotype promoting a more stable epithelial-mesenchymal transition and enhanced cell invasion