Identification and Functional Characterization of the Human Glutathione S-Transferase P1 Gene as a Novel Transcriptional Target of the p53 Tumor Suppressor Gene

Abstract

The glutathione S-transferase P1 (GSTP1) is involved in multiple cellular functions, including phase II metabolism, stress response, signaling, and apoptosis. The mechanisms underlying the significantly high GSTP1 expression in many human tumors are, however, currently not well understood. We report here that the GSTP1 gene is a heretofore unrecognized downstream transcriptional target of the tumor suppressor p53. We identified a p53-binding motif comprising two consecutive half-sites located in intron 4 of the GSTP1 gene and is highly homologous to consensus p53-binding motifs in other p53-responsive genes. Using a combination of electrophoretic mobility shift assay and DNase I footprinting analyses, we showed that wild-type p53 protein binds to the GSTP1 p53 motif and luciferase reporter assays showed the motif to be transcriptionally functional in human tumor cells. In a temperature-sensitive p53-mutant cells, levels of both p21/WAF1 and GSTP1 gene transcripts increased time dependently when cells were switched from the inactive mutant state to the wild-type p53 state. Small interfering RNA-mediated reduction of p53 expression resulted in a specific decrease in GSTP1 expression and in tumor cells with mutated p53; adenovirally mediated expression of wild-type p53 increased GSTP1 expression significantly. In a panel of early-passage brain tumor cultures from patients, high levels of GSTP1 transcripts and protein were associated with wild-type p53 and, conversely, low GSTP1 levels with mutant p53. p53 expression knockdown by small interfering RNA increased cisplatin sensitivity. The ability of wild-type p53 to transcriptionally activate the human GSTP1 gene defines a novel mechanism of protecting the genome and, potentially, of tumor drug resistance.