title: Chemical inducers of rodent glutathione s-transferases down-regulate human GSTA1 transcription through a mechanism involving variant hepatic nuclear factor 1-C.
abstract: The regulation of human GSTA1 by chemical inducers of rodent glutathione S-transferases (GSTs) and the regulatory role of hepatic nuclear factor (HNF) 1 was investigated in Caco-2 cells. Treatment of preconfluent and confluent cells with 12-O-tetra-decanoyl phorbol-13-acetate (TPA), 3-methylcholanthrene (3-MC), 2-tert-butyl-4-hydroxy-anisol (BHA), and phenobarbital (PB) reduced GSTA1 mRNA levels in preconfluent and confluent cells. Constitutive levels of GSTA1 and HNF1alpha mRNA were elevated 6.25- and 50-fold, respectively, in postconfluent cells compared with preconfluent cells. Overexpression of HNF1alpha in cells transfected with a GSTA1 promoter-luciferase construct (pGSTA1-1591-luc) resulted in dose-related increases in reporter activity not observed when an HNF1 response element (HRE) in the proximal promoter was mutated (pGSTA1-DeltaHNF1-luc). TPA, 3-MC, BHA, and PB reduced HNF1alpha mRNA levels in preconfluent and confluent cells and caused marked reductions in luciferase activity in pGSTA1-1591-luc transfectants. Transcriptional repression was abrogated with pGSTA1-DeltaHNF1-luc and with truncated constructs that eliminated a functional HRE. Moreover, cotransfection of pHNF1alpha with pGSTA1-1591-luc partially prevented the reduction in luciferase activity by rodent GST inducers. Immunoblot analysis of DNA binding studies indicate that variant (v)HNF1-C binding to HRE is increased in preconfluent cells treated with 3-MC, BHA, and PB. In addition, overexpression of vHNF1-C repressed GSTA1 transcriptional activity in luciferase reporter assays. Finally, treatment with 3-MC, BHA, and PB increased vHNF1-C mRNA levels in preconfluent cells. These data demonstrate that repression of human GSTA1 transcription by chemical inducers of rodent GSTs occurs, in part, through a mechanism involving the repressive action of vHNF1-C.
authors.affiliation: Department of Biomedical Sciences, University of Guelph, Guelph, ON, Canada N1G 2W1. gkirby@uoguelph.ca.
authors.forename: Laura
authors.initials: L
authors.lastname: Romero
authors.forename: Lorraine
authors.initials: L
authors.lastname: Ng
authors.forename: Gordon M
authors.initials: GM
authors.lastname: Kirby
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002083:Butylated Hydroxyanisole / Q000494:pharmacology; D018938:Caco-2 Cells; D015536:Down-Regulation / Q000187:drug effects / Q000235:genetics; D024202:Electrophoretic Mobility Shift Assay; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism*; D051538:Hepatocyte Nuclear Factor 1-alpha / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008748:Methylcholanthrene / Q000494:pharmacology; D051379:Mice; D010634:Phenobarbital / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D020218:Response Elements / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects; D015262:Xenobiotics / Q000494:pharmacology*
chemical_list: D051538:Hepatocyte Nuclear Factor 1-alpha; D020033:Protein Isoforms; D012333:RNA, Messenger; D015262:Xenobiotics; D002083:Butylated Hydroxyanisole; D008748:Methylcholanthrene; C051361:2-tert-butyl-4-hydroxyanisole; C484555:GSTA1 protein, human; D005982:Glutathione Transferase; D013755:Tetradecanoylphorbol Acetate; D010634:Phenobarbital