Cells respond to genotoxic stress by activation of many genes, including the tumor suppressor p53. p53 activates transcriptionally target genes, such as p21waf1 and gadd45, which can lead to cell cycle arrest, or bax, which can lead to cell death. We examined the response to genotoxic stress in two hematopoietic cell lines that harbor either wild-type (MOLT-4) or a mutant p53 with a codon 161 mutation (U266). We adapted a multiprobe RNase protection assay (RPA) to determine the steady-state RNA levels, and in combination with nuclear runoff assays, transcriptional rates of multiple stress-induced genes. We found a differential activation of growth arrest and cell death-specific p53 target genes in cells with wild-type or mutant p53. Our results show that genotoxic stress can activate the p21waf1 and gadd45 genes in both cell lines. However, the bax gene was not induced in U266 cells. Bax and gadd45 gene induction could be efficiently blocked by pretreating the cells with the antioxidant compound pyrrolidine dithiocarbamate, suggesting that oxidative stress was involved in these responses. Induction of all three genes in MOLT-4 cells was clearly at the transcriptional level, because we detected transcriptional activity by nuclear runoff RPA assays, and transfection with a consensus p53 binding sequence. U266 cells did not activate the same reporter, in spite of the upregulation of p21waf1 and gadd45 RNA levels. However, the p21waf1-reporter constructs containing 0.9 to 2.4 kb of the native p21 promoter were potently activated in U266 cells. These results indicate a differential regulation of p53 target genes in cells containing wild-type or codon 161 mutant p53.