We recently have shown that activated Ras, but not Raf, causes transformation of intestinal (RIE-1, IEC-6) epithelial cells, whereas both activated Ras and Raf transform NIH 3T3 fibroblasts (Oldham, S. M., Clark, G. J., Gangarosa, L. M., Coffey, R. J., and Der, C. J. (1996) Proc. Natl. Acad. Sci. U. S. A. 93, 6924-6928). The observations that conditioned medium from Ras-, but not Raf-, transfected RIE-1 cells, as well as exogenous transforming growth factor alpha (TGFalpha), promoted morphological transformation of parental RIE-1 cells prompted us to identify epidermal growth factor (EGF) receptor (EGFR) ligands produced by Ras-transformed RIE-1 cells responsible for this autocrine effect. Since studies in fibroblasts have shown that v-Src is transforming, we also determined if v-Src could transform RIE-1 cells. H- or K-Ras-transformed cells secreted significant amounts of TGFalpha protein, and mRNA transcripts for TGFalpha, amphiregulin (AR), and heparin-binding EGF-like growth factor (HB-EGF) were induced. Like Ras, v-Src caused morphological and growth transformation of parental RIE-1 cells. However, TGFalpha protein was not secreted by RIE-1 cells stably expressing v-Src or activated Raf, and only minor increases in EGFR ligand mRNA expression were detected in these cells. A selective EGFR tyrosine kinase inhibitor PD153035 attenuated the Ras-, but not Src-, transformed phenotype. Taken together, these observations provide a mechanistic and biochemical basis for the ability of activated Ras, but not activated Raf, to cause transformation of RIE-1 cells. Finally, we suggest that an EGFR-dependent mechanism is necessary for Ras, but not Src, transformation of these intestinal epithelial cells.