Rhabdomyosarcoma is the most commonly occurring soft-tissue sarcoma in children. Some reports have discussed the altered expression and molecular abnormalities of cell-cycle-regulatory proteins in rhabdomyosarcoma; however, variable frequencies of occurrence have been noted. In the current study, among 72 cases of rhabdomyosarcoma, the authors evaluated for the expression of p53, MDM2, p16, p21/WAF1, p27, cyclin D1, cyclin E, pRb and E2F-1 protein immunohistochemically and assessed for proliferative activities using MIB-1. We also analyzed the mutation of the p53 gene in 45 cases, the amplification of the MDM2 gene in 18 cases and the mutation of the H-ras gene in 29 cases, using formalin-fixed paraffin-embedded materials. Furthermore, we assessed the correlation between clinicopathologic factors and the results of both immunohistochemical and molecular analyses. Alveolar type affected older patients, and it had a significantly higher mitotic rate compared with the embryonal type (P=0.0226). p53 overexpression was detected in 22 (30.6%) of 72 cases, and 10 (22.2%) of 45 cases had p53 gene abnormalities. As for MDM2, its overexpression was found in nine (12.5%) of 72 cases, and three (16.7%) of 18 cases showed MDM2 amplification. A statistically significant association was observed between immunoreaction for MDM2 and p53 overexpression (P=0.0002), and p53 and MDM2 overexpression was significantly correlated with high MIB-1 labeling indices. E2F-1 labeling indices showed a significantly higher score in alveolar type compared with that seen in embryonal type (P=0.0334), but MIB-1 did not. In conclusion, our study suggests that p53 overexpression may be related to tumor progression because tumors with p53 overexpression have a high proliferative activity in the current study. Alveolar type had a significantly higher both mitotic rate and E2F-1 labeling indices when compared with the embryonal type. The current study is the first report of the correlation of E2F-1 with alveolar rhabdomyosarcoma.