Downstream signal transduction defects that suppress transformation in two revertant cell lines expressing activated...

The neu gene encodes the transmembrane tyrosine kinase growth factor receptor, p185. To study neu-induced cellular transformation, we developed revertant cells from the neu-transformed NIH 3T3 cell line, B104-1-1, by treating the cells with the chemical mutagen ethyl methanesulfonate. The morphologically normal revertant cells were first selected by their ability either to attach to culture plates or to survive in the presence of the cytotoxic reagents colchicine or 5-fluoro-2-deoxyuridine. Two of the 21 candidate revertant cell lines isolated were further characterized and were found to lose their anchorage independence and ability to grow in 1% calf serum, indicating that they were nontransformed even though they still expressed p185 oncoprotein. The tyrosine residues of p185 in these two revertants were underphosphorylated, which may have contributed to their nontransformed status. In addition, these revertants also resisted transformation by neu and several additional oncogenes (H-ras, N-ras, v-mos, v-abl, and v-fos) as determined by focus forming assays. These results indicated that we had successfully developed, from neu-transformed cells, revertants exhibiting defective tyrosine phosphorylation of the neu oncoprotein. The results also suggested that neu and several other oncogenes may share common elements in their pathways for the induction of cellular transformation.