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. 1996 Jan 1;313(Pt 1):155–161. doi: 10.1042/bj3130155

The organization of the human GSTP1-1 gene promoter and its response to retinoic acid and cellular redox status.

C Xia 1, J Hu 1, B Ketterer 1, J B Taylor 1
PMCID: PMC1216876  PMID: 8546677

Abstract

High levels of expression of GSTP1-1 are associated with cell proliferation, embryogenesis and malignancy. Given the role of glutathione S-transferase (GST) in detoxication, it is possible that GSTP1-1 evolved specifically to protect proliferating cells and share regulatory mechanisms with other cellular genes which are involved in cell division and tumorigenesis. We have previously shown that the expression of GSTP1 is suppressed by retinoic acid (RA) in the presence of the retinoic acid receptor (RAR) as a result of decreased transcription from its promoter. Through deletion analysis, we show here that the RA-RAR-dependent repression is mediated by the region -73 to +8. Further mutation analysis of this region indicates that the DNA sequence required for RA-RAR-dependent repression co-localizes with a consensus activator protein-1 (AP1) site essential for the promoter activity. The degree of repression correlates with the residual activity of the AP1 site. There are two adjacent G/C boxes. The one immediately downstream from the AP1 site is not essential for the promoter activity, but mutation of the second, further downstream, impairs the promoter. On the other hand, mutation of either of these two G/C boxes has little effect on RA-RAR suppression. We also show that the expression of GSTP1 is regulated by the redox status of the cell. Using the chloramphenicol acetyltransferase assay system, we have demonstrated that treatment with H2O2 induced transcription from the promoter and that this effect can be blocked by pre-incubation with N-acetylcysteine (NAC). It was shown that the induction by H2O2 is mediated by trans-acting factor NF-kappa B (nuclear factor kappa B), via a putative NF-kappa B site, 'GGGACCCTCC', located from -96 to -86. Co-transfection with an NF-kappa B (p65) expression construct increased the promoter activity, an effect which could be blocked by co-transfection with an I kappa B (MAD-3) expression construct. Deletion of the NF-kappa B site abolished the effect of both H2O2 and co-transfection of NF-kappa B. Interestingly, NAC is also an inducer for GSTP1. The effect of NAC was shown to be mediated largely by the AP1 site, since mutation of this site abolished the induction by NAC.

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