Despite the significance of oxidative damage for carcinogenesis, the molecular mechanisms that lead to increased susceptibility of tissues to oxidative stress are not well-understood. We now report a link between loss of protection against oxidative damage and loss-of-function of Nkx3.1, a homeobox gene that is known to be required for prostatic epithelial differentiation and suppression of prostate cancer. Using gene expression profiling, we find that Nkx3.1 mutant mice display deregulated expression of several antioxidant and prooxidant enzymes, including glutathione peroxidase 2 and 3 (GPx2 and GPx3), peroxiredoxin 6 (Prdx6), and sulfyhydryl oxidase Q6 (Qscn6). Moreover, the formation of prostatic intraepithelial neoplasia in these mutant mice is associated with increased oxidative damage of DNA, as evident by increased levels of 8-hydroxy-2′-deoxyguanosine. We further show that progression to prostate adenocarcinoma, as occurs in compound mutant mice lacking Nkx3.1 as well as the Pten tumor suppressor, is correlated with a further deregulation of antioxidants, including superoxide dismutase enzymes, and more profound accumulations of oxidative damage to DNA and protein, the latter manifested by increased levels of 4-hydroxynonenal. We propose that the essential role of Nkx3.1 in maintaining the terminally differentiated state of the prostate epithelium provides protection against oxidative damage and, thereby, suppression of prostate cancer. Thus, our findings provide a molecular link between a gene whose inactivation is known to be involved in prostate carcinogenesis, namely Nkx3.1, and oxidative damage of the prostatic epithelium.