The protein product of the rodent neu oncogene, p185neu, is a tyrosine kinase with structural similarity to the epidermal growth factor receptor (EGFR). Pansfection and subsequent oversxpression of the human p185c-erbB2 protein transforms NIH 3T3 cells in vitro. However, NIH 3T3 cells are not transformed by ovetexpressed rodent pi85c-neu. NIH 3T3 transfectants overexpressing EGF receptors are not transformed unless EGF is added to the cultures, and, even then, they are incompletely transformed. Several groups have recently demonstrated EGF-induced, EGFR-mediated phosphorylation of pl85c-neu. During efforts to characterize the interaction of p185c-neu with EGFR further, we created cell lines that simultaneously ovetexpress both pl85c-neu and EGFR and observed that these cells become transformed. These observations demonstrate that two distinct, overexpressed tyrosine kinases can act synergistically to transform NIH 3T3 cells, thus identifying a novel mechanism that can lead to transformation.