Most of the p53 target genes, all except MDM2, COP1 and PIRH2, perform functions in apoptosis, differentiation and cell cycle arrest. The aforementioned oncogenes downregulate p53 through a negative feedback mechanism, and thus contribute to tumor development. In this study, we report a new p53 target, PRL-1, which is believed to be a significant regulator in the development and metastasis of a variety of cancer types. Phosphatase of regenerating liver 1 (PRL-1) overexpression reduced the levels of endogenous and exogenous p53 proteins, and inhibited p53-mediated apoptosis. On the other hand, the ablation of PRL-1 by small interfering RNA (siRNA) increased p53 protein levels. The p53 downregulation was mediated by p53 ubiquitination and subsequent proteasomal degradation. Furthermore, p53 ubiquitination by PRL-1 was achieved through two independent pathways, by inducing PIRH2 transcription and by inducing MDM2 phosphorylation through Akt signaling. In addition, we showed that the PRL-1 gene harbors a p53 response element in the first intron, and its transcription is regulated by the p53 protein. These findings imply that the new oncogenic p53 target, PRL-1, may contribute to tumor development by the downregulation of p53 by a negative feedback mechanism.