The alkylating agent 4-nitroquinoline 1-oxide (4-NQO) is a powerful carcinogen and induces squamous cell hyperplasia, squamous cell dysplasia, papilloma, and squamous cell carcinoma (SCC) in rat oral epithelia. Oral cancers induced by a single application of 4-NQO develop through a multistage process in a way similar to the development of this cancer in humans. In this study, mutations in exons 1 and 2 of Ki-ras, N-ras, and Ha-ras and exons 4-7 of p53 were examined by polymerase chain reaction (PCR)-single strand conformation polymorphism (SSCP) analysis, followed by PCR-direct sequencing for the confirmation of mutations. Samples for the mutation analysis were obtained from dysplasias, papillomas, and SCCs on the tongue epithelia induced in F344 rats by adding 4-NQO (20 ppm) to their drinking water for 8 wk. The Ha-ras mutations (61A-->T transversions in the second position) were found in five of 29 (17%) samples (one dysplasia and four SCCs). However, no mutations were detected in either Ki-ras, N-ras, or p53 under two different conditions of PCR-SSCP analysis. We suggest that some neoplasms in oral carcinogenesis induced by 4-NQO may involve Ha-ras mutations but not mutations in Ki-ras, N-ras, or p53. The 4-NQO-induced rat oral carcinogenesis model may provide a system for evaluation of the mechanisms of multistage oral carcinogenesis associated with Ha-ras mutation without Ki-ras, N-ras, or p53 mutation.