In vitro studies suggest that effective tumor suppression by p53 requires multiple domains to execute transcription-dependent and transcription-independent functions. We generated a mutant p53 allele in mice, p53(W25QL26S) (p53(QS)), containing an inactive transactivation domain to evaluate the importance of transactivation for p53-mediated tumor suppression. Recently, we discovered that the allele also contains a valine substitution for alanine at codon 135, which borders the DNA-binding domain. We found that p53(QSval135) bound to chromatin albeit less well than p53(QSala135), but both were equally deficient in transcriptional regulation, apoptosis induction in mouse embryo fibroblasts (MEFs), and suppression of tumor formation by E1A, Ha-Ras transformed MEFs. p53(QSval135) mice and p53-null mice exhibited identical tumor development kinetics and spectra in spontaneous and oncogene-initiated tumorigenicity assays, when tested in a homo- and heterozygous configuration. The p53(QSval135) allele did not have dominant negative functions and behaved as a null allele. Taken together, these data indicate that effective tumor suppression requires the transcriptional regulation function of p53, and they suggest that transactivation independent functions of p53 are unlikely to contribute significantly to tumor suppression in vivo.