Received for publication September 19, 2007.
Revised November 20, 2007.
Accepted for publication November 21, 2007.

REGULATION OF THE RAT UDP-GLYCOSYLTRANFERASE 1A6 BY GLUCOCORTICOIDS INVOLVES A CRYPTIC GLUCOCORTICOID RESPONSE ELEMENT

Abstract

Glucocorticoids precociously induce fetal rat UDP-glycosyltransferase 1A6 (UGT1A6) and potentiate polycyclic aromatic hydrocarbon (PAH)-dependent induction of this enzyme in vivo and in isolated rat hepatocytes. To establish whether induction was due to glucocorticoid receptor (GR), luciferase reporter vectors were tested in transfection assays with HepG2 cells. Using a reporter construct containing approximately 2.26 kb of the 5'-flanking region of the UGT1A6 non-coding leader exon (A1*), dexamethasone increased basal activity 3-to-7-fold in cells co-transfected with an expression plasmid for GR. PAH increased gene expression 23-fold, but the presence of DEX only induced PAH-dependent expression by 1.5-fold, suggesting interaction between GR and Ah receptor. Further, the GR antagonist RU 38 486 was a partial agonist increasing, rather than inhibiting basal activity 3-fold. 5’-deletion analysis defined the 5’-boundary for a functional glucocorticoid responsive unit between base pairs –141 and –118 relative to the transcription start site. This region contains the Ah receptor (AhRE) response element and both PAH and glucocorticoid-dependent gene activation were lost when this area was deleted. Mutation of a single base pair located in the AhRE region simultaneously reduced induction by PAH and increased glucocorticoid induction. Thus the sequences of both the AhRE and glucocorticoid response elements appear to overlap, suggesting Ah receptor binding may decrease glucocorticoid-dependent induction due to interactions of these two cis-acting elements. Mutation of a putative GRE located between base pair -81 and -95 reduced, but did not completely eliminate glucocorticoid-dependent induction of the reporter, suggesting a non-classical mechanism of induction is involved in this response.

Key words: Ah receptor, enzyme induction, gene regulation, glucuronidation, nuclear receptors, UDP glucuronyltransferases