The tumor suppressor p53 plays a crucial role in the cellular response to DNA damage by transcriptional activation of numerous downstream genes. Although a considerable number of p53 target genes have been reported, the precise mechanism of p53-regulated tumor suppression still remains to be elucidated. Here, we report a novel role of the DFNA5 gene in p53-mediated etoposide-induced cell death. The DFNA5 gene has been previously reported to be responsible for autosomal-dominant, nonsyndromic hearing impairment. The expression of the DFNA5 gene was strongly induced by exogenous and endogenous p53. The chromatin immunoprecipitation assay indicated that a potential p53-binding sequence is located in intron 1 of the DFNA5 gene. Furthermore, the reporter gene assay revealed that the sequence displays p53-dependent transcriptional activity. The ectopic expression of DFNA5 enhanced etoposide-induced cell death in the presence of p53; however, it was inhibited in the absence of p53. Finally, the expression of DFNA5 mRNA was remarkably induced by gamma-ray irradiation in the colon of p53(+/+) mice but not in that of p53(-/-) mice. These results suggest that DFNA5 plays a role in the p53-regulated cellular response to genotoxic stress probably by cooperating with p53.