The product of the tumor suppressor gene p53 binds to DNA and activates transcription from promoters containing its consensus binding site. This activity has been hypothesized to be responsible for its biological effects. However, the total number and nature of human genomic sites with which p53 can functionally interact is unknown. In this paper, we have used a Saccharomyces cerevisiae-based screen to identify human genomic sequences that activate transcription from an adjacent reporter gene in a p53-dependent manner (p53-tagged sites, PTS). Fifty-seven different PTS were identified, and the total number of such sites in the human genome was predicted to be between 200 and 300. Almost all contained two adjacent copies of the previously defined consensus 5'-PuPuPuC(A/T)(T/A)GPyPyPy-3'. Spacing between the copies was found to be critical for sequence-specific transcriptional activation in vivo. These results further refine the nature of the genomic sequences likely to be most important for p53-mediated tumor suppression.