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J Biol Chem, Vol. 273, Issue 34, 21856-21866, August 21, 1998
From the Dehydroepiandrosterone
sulfotransferase (Std) catalyzes sulfonation of androgenic steroids and
certain aromatic procarcinogens. In rats, this enzyme is selectively
expressed in the liver, and its expression is strongly repressed by
androgens. DNase I footprinting and electrophoretic mobility shift
analyses revealed two hepatocyte nuclear factor-1 (HNF1), three
CCAAT/enhancer-binding protein (C/EBP), and one consensus palindromic
thyroid hormone response elements within the first 215 base pairs (bp)
of the promoter sequence of rat Std. This promoter is
normally inactive in fibroblast-derived NIH 3T3 cells. However,
overexpression of HNF1 and C/EBP resulted in synergistic activation of
the Std promoter in this cell type, indicating essential
roles of these two trans-regulators in liver-selective expression of
the rat Std gene. On the other hand, point mutations at any
one of five cis elements proximal to the
Tissue-specific and Androgen-repressible Regulation of the Rat
Dehydroepiandrosterone Sulfotransferase Gene Promoter
,,, and§
Department of Cellular and Structural
Biology,
215 bp region markedly
reduced reporter gene expression, suggesting that all of these sites
are important for overall promoter function. Androgenic repression of
the Std gene in rat liver can be recapitulated in androgen
receptor (AR)-negative HepG2 hepatoma cells after cotransfection with
an AR expression plasmid. Functional assay of a nested set of
5'-deleted promoters mapped the negative androgen response region
between positions 235 and 310. Antibody supershift and oligonucleotide competition identified three OCT-1 and two C/EBP elements between bp 231 and 292. An additional OCT-1 site was found
to overlap with a C/EBP element at the 262/252 position. Mutational
inactivation of any one of five cis elements within the 231/292
region abolished negative androgen response. However, none of these cis
elements showed DNase I protection by recombinant AR in footprinting
assay, suggesting the absence of a direct AR-DNA interaction. Thus,
these studies on rat Std promoter function indicate that
(i) HNF1 and C/EBP are responsible for liver specificity of the rat
Std gene; (ii) androgenic repression of the gene requires the presence of all of the OCT-1 and C/EBP elements between positions 231 and 292; and (iii) AR may exert its negative regulatory effect
indirectly through transcriptional interference of OCT-1 and C/EBP
rather than through a direct DNA-AR interaction.
Copyright © 1998 by The American Society for Biochemistry and Molecular Biology, Inc.
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