Changes that occur during tumor promotion, the rate-limiting phase of multistep carcinogenesis, may offer the best targets for prevention of cancer or reversal of early disease. The murine epidermal JB6 promotion-sensitive (P+) and -resistant (P−) cell lines provide a cell culture model for tumor promoter-induced neoplastic transformation ideally suited to the identification of molecular events that mediate or inhibit transformation. A differential display comparison of P+ and P− cell mRNAs yielded seven differentially expressed sequences. One of the sequences preferentially expressed in P− cells identified an ∼3.6-kb message that was induced to higher levels in P− cells following exposure to the tumor promoter 12-O-tetradecanoylphorbol acetate than in P+ cells. The message was detected in mRNA from heart, lung, and spleen. cDNA cloning of the P− preferential sequence revealed a high degree of identity to human pleckstrin (PLEK), the major PKC substrate in platelets (Tyers et al., 1988, Nature 333: 470). We report the complete mouse cDNA sequence of pleckstrin and the localization of the gene to chromosome 11, its expression in a nonhematopoetic cell line, and its potential role in blocking neoplastic transformation.