The caudal-related homeodomain protein Cdx2 and hepatocyte nuclear factor 1alpha cooperatively regulate the UDP-glucuronosyltransferase 2B7 gene promoter.
The gastrointestinal tract, contains several UDP-glucuronosyltransferases (UGTs) of the UGT1A and UGT2B subfamilies. UGT2B7 is one particular enzyme expressed throughout the gastrointestinal tract that possesses broad substrate specificity towards orally administered drugs. Because the caudal-related homeodomain protein 2 (Cdx2) regulates many gastrointestinal properties, we sought to determine whether it could regulate the UGT2B7 promoter in the colon-derived cell line Caco-2. Levels of Cdx2 and UGT2B7 were measured in differentiated and non-differentiated Caco-2 cells by the quantitative polymerase chain reaction. The capacity of the UGT2B7 gene promoter to drive expression of the luciferase reporter gene was assessed by transfection into Caco-2 cells, with transcription factor expression plasmids. Mutation of putative transcription factor binding sites and electrophoretic mobility shift assays were used to define important regulatory regions of the UGT2B7 gene promoter. The levels of Cdx2 and UGT2B7 mRNAs were co-ordinately increased in differentiated Caco2 cells compared to non-differentiated cells. Cdx2 activates the UGT2B7 proximal promoter by binding to two adjacent sites. Promoter activation requires Cdx2 binding to both sites wherein these proteins interact to form a putative functional dimer. Dimerization was shown to be dependent on redox state using extracts depleted of dithiothreitol. In addition, Cdx2 was shown to cooperatively activate the UGT2B7 promoter in conjunction with hepatocyte nuclear factor 1alpha (HNF1alpha), a mechanism previously observed to regulate other intestine-specific genes. The present study is the first to define transcription factors involved in the control of intestinal UGT2B expression. The demonstration that Cdx2 and HNF1alpha are important regulators of UGT2B7 expression will aid in defining pathways for coordinate control of drug metabolism in the gastrointestinal tract.
Gregory
Philip A
PA
Department of Clinical Pharmacology, Flinders University School of Medicine, Flinders Medical Centre, Bedford Park, SA 5042, Australia.
Gardner-Stephen
Dione A
DA
Rogers
Anne
A
Michael
Michael Z
MZ
Mackenzie
Peter I
PI
D001711:Biotransformation / Q000235:genetics / Q000502:physiology; D000071616:CDX2 Transcription Factor; D018938:Caco-2 Cells; D005786:Gene Expression Regulation; D014453:Glucuronosyltransferase / Q000096:biosynthesis / Q000235:genetics*; D051538:Hepatocyte Nuclear Factor 1-alpha / Q000502:physiology*; D018398:Homeodomain Proteins / Q000502:physiology*; D006801:Humans; D009154:Mutation; D011401:Promoter Regions, Genetic; D014157:Transcription Factors / Q000235:genetics
D000071616:CDX2 Transcription Factor; C491270:CDX2 protein, human; D051538:Hepatocyte Nuclear Factor 1-alpha; D018398:Homeodomain Proteins; D014157:Transcription Factors; D014453:Glucuronosyltransferase