Human GSTA1 regulation and function: GSTA1 repression by HNF-1 and JNK inhibition via GSTA1

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Romero, Laura

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University of Guelph

Abstract

The glutathione S-transferases (GSTs) represent a major group of detoxification enzymes that play an important role in the clearing of reactive electrophiles and products of lipid peroxidation. GSTs have been recently identified as redox sensitive inhibitors of stress-activated kinase activity. This study investigated the molecular mechanisms underlying repression of human GST alpha class 1-1 (GSTA1-1) in cultured human colonic epithelial cells by proinflammatory cytokines and chemical inducers of rodent GSTs. In addition, the inhibitory effect of GSTA1-1 on stress-mediated Jun NH2-terminal kinase (JNK) activation was examined. Recombinant human IL-1[beta] caused dose-dependent reductions in GSTA1/A2 mRNA, protein and activity in Caco-2 cells. In luciferase reporter gene experiments, GSTA1 promoter activity was decreased in Caco-2 cells treated with IL-1[beta]. The IL-1[beta]-responsive region was mapped to an area 286 base pairs upstream to the coding region. Deletion of a hepatic nuclear factor 1 (HNF-1) responsive element (HRE) within this region abolished the IL-1[beta]-mediated suppression of GSTA1 luciferase promoter activity. We also examined the involvement of HNF-1[alpha] in the transcriptional repression of GSTA1 by some archetypal inducers of rodent GSTs, such as 12-O-teradecanoyl phorbol-13-acetate (TPA), 3-methylcholanthrene (3-MC), 2-tert-butyl-4-hydroxy-anisol (BHA) and Phenobarbital (PB). Using truncated reporter constructs and site directed mutagenesis we showed that the repressive effect of xenobiotics was also dependent on the presence of a functional HRE. Finally, to determine the influence of GSTA1-1 in inhibiting JNK stress signaling, we compared the degree of JNK activation by IL-[beta], H2 O2 or ultraviolet (UV) irradiation in cells with differential GSTA1 expression. JNK activation was significantly reduced in post-confluent Caco-2 cells with elevated GSTA1-1 and in MEF 3T3 cells with tet-off-inducible GSTA1-1 expression. Moreover, the frequency of N-butryrate/TNF[alpha] apoptosis was significantly reduced in postconfluent Caco-2 cells that over-express GSTA1-1. These results demonstrate a profound inhibitory effect of IL-1[beta] and xenobiotics on human GSTA1-1 transcription via a repressive mechanism mediated by HNF-1. Furthermore, a novel role was identified for GSTA1-1 suppression of JNK activation by inflammatory cytokines and oxidative stress.

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glutathione S-transferases, detoxification enzymes, reactive electrophiles, lipid peroxidation, stress-activated kinase

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