p53 is a transcription factor that is activated by genotoxic stress and mediates cell cycle arrest and apoptosis. Here we demonstrate that infection of mouse liver with recombinant E1/E3-deleted adenovirus leads to p53 activation and simultaneously to the down-regulation of albumin gene expression. In vitro transcription assays indicate that transcriptional mechanisms mediated through the albumin promoter are responsible for reduced albumin mRNA levels during viral infection. Albumin expression is maintained in the liver by a combination of liver-enriched transcription factors such as CAAT enhancer-binding protein (C/EBP)alpha and C/EBPbeta. We show that p53 wild type and tumor-derived p53 mutations repress C/EBP-mediated transactivation of the albumin promoter. The binding of C/EBPalpha or -beta to its cognate sequence in the albumin promoter is not inhibited by p53 expression. Deletion analysis and domain swapping experiments show that repression of C/EBPbeta-mediated transactivation is dependent on the N-terminal domain of p53 and the transactivation domain, leucine zipper domain, and the inhibitory domain II (amino acids 163-191) of C/EBPbeta. Our results provide a molecular explanation for the p53-mediated down-regulation of liver-specific gene expression after viral infection. Additionally, as overexpression of p53 mutants is frequently found in undifferentiated hepatocellular carcinomas, the same mechanisms may contribute to the lack of liver-specific gene transcription in these tumors.