Tissue organization and maintenance within multicellular organisms is in part dependent on the ability of cells to undergo programmed cell death or apoptosis. Conversely, disruption of cell death pathways often is associated with tumor development. At present, the molecular control of apoptosis in epithelial cells is poorly understood. Here we describe evidence linking epidermal growth factor-receptor (EGF-R) activation to survival of normal human keratinocytes in culture. Inhibition of EGF-R activation by an anti-EGF-R antagonistic monoclonal antibody (mAb 425), followed by detachment of keratinocytes from the substratum, induced extensive death with several features of apoptosis in keratinocyte cultures. Other, non-epithelial normal human cells including melanocytes and fibroblasts, did not show this effect. Similar to EGF-R blockade by mAb 425, inhibition of the EGF-R tyrosine kinase activity using tyrphostin AG1478 resulted in lack of attachment and extensive cell death upon passaging. Attachment to keratinocyte-derived ECM partially resuced mAb 425-treated keratinocytes from cell death, indicating that adhesion-dependent and EGF-R-dependent signal transduction pathways serve partially overlapping but not redundant roles in supporting keratinocyte survival.