The p53 gene is the most frequently mutated gene in human cancers. Loss of functional p53 leads to impaired responses of cancer cells to apoptosis induction and to poor prognosis in patients with certain types of cancer. Cancer gene therapies using ectopic expression of wild-type p53 to force cancer cells through the apoptotic pathway have been tested extensively preclinically and clinically, and genes in various cell lines have been reported to be regulated upon ectopic p53 overexpression. However, the effect of p53 on many other p53-dependent and apoptosis-related genes remains unclear, as does the mechanism of p53-induced apoptosis in human cancers. In this study, we used real-time reverse transcription polymerase chain reaction to evaluate the changes in expression of various p53-dependent and apoptosis-related genes in five human non-small-cell lung cancer cell lines with varying p53 statuses after adenoviral p53 treatment. We found that Ad/p53 induced the expression of the proapoptotic genes PUMA, Bak, Bax, and Fas in a cell type- and time-dependent manner. Among these genes, PUMA was upregulated the most dramatically and broadly. However, when a specific siRNA construct against PUMA was employed, we observed no attenuation of apoptosis in H1299 cells. Our data suggest that Ad-p53 induces the expression of a variety of proapoptotic genes and that lack of induction in one of these genes does not block Ad/p53-mediated cell killing in human lung cancer cells.