The influence of zinc status on p21 gene expression was examined in human hepatoblastoma (HepG2) cells. Cells were cultured for one passage in a basal medium depleted of zinc to induce severe zinc-deficient (ZD) cells or in the basal medium supplemented with 0.4, 4.0, 16, or 32 µM zinc to represent mild zinc deficiency (ZD0.4), the amount of zinc in most normal media (ZN), the normal human plasma zinc level (zinc-adequate, ZA), or the high end of plasma zinc attainable by oral supplementation (ZS), respectively. In ZD and ZD0.4 cells, the nuclear p21 protein level, mRNA abundance and promoter activity were reduced to 40%, 70%, and 65%, respectively, of ZN cells. However, p21 protein and mRNA levels as well as p21 promoter activity were not altered in ZA and ZS cells as compared to ZN cells. Moreover, the amounts of acetylated histone 4 associated with the proximal and distal p21 promoter regions, as a measure of p21 promoter accessibility, were decreased in ZD (73 and 64 %, respectively) and ZD0.4 (82 and 77 %, respectively) cells than in ZN cells (100 and 100 %, respectively). Thus, multiple lines of evidence indicate that the transcriptional process of p21 is down-regulated by depressed zinc status in HepG2 cells. Furthermore, the transfection of 5µg of PCMV-p21 plasmid, which constitutively expressed p21, was able to normalize the reduction in p21 protein level and cyclin D1- cdk4 complex activity but not the inhibition of cell growth and G1/S cell cycle progression in ZD cells.