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J. Biol. Chem., Vol. 277, Issue 10, 7945-7954, March 8, 2002
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From the Division of Reproductive Biology, Department of Gynecology
and Obstetrics, Stanford University School of Medicine,
Stanford, California 94305
The enzyme 3 The nucleotide sequence(s) reported in this paper for the mouse
3
AP-2
and the Homeodomain Protein Distal-less 3 Are
Required for Placental-specific Expression of the Murine
3
-Hydroxysteroid Dehydrogenase VI Gene, Hsd3b6*
-hydroxysteroid
dehydrogenase/isomerase (3-HSD) is essential for the biosynthesis of
all active steroid hormones. It exists as multiple isoforms in humans
and rodents, each the product of a distinct gene. Human 3-HSD I in
placenta is essential for placental progesterone biosynthesis and thus
is essential for the maintenance of pregnancy. The murine ortholog,
3-HSD VI, is the only isoform expressed in giant trophoblast cells
during the first half of mouse pregnancy. This study was designed to identify the cis-acting element(s) and the associated
transcription factors required for trophoblast-specific expression of
3-HSD VI. Transfection studies in placental and
nonplacental cells identified a novel 66-bp trophoblast-specific
enhancer element located between 
2896 and 2831 of the 3-HSD VI
promoter. DNase protection analysis of the enhancer element identified
three trophoblast-specific binding sites, FPI, FPII, and FPIII.
Electrophoretic mobility shift assays with oligonucleotides
representing the protected sequences, FPI and FPIII, and nuclear
extracts isolated from human JEG-3 cells and from mouse trophoblast
cells, demonstrated the same binding pattern that was distinct from the
binding pattern with mouse Leydig cell nuclear proteins. Further
electrophoretic mobility shift assays identified AP-2
and the
homeodomain protein, Dlx 3, as the transcription factors that
specifically bind to FPI and FPIII, respectively. Site-specific
mutations in each of the binding sites eliminated enhancer activity
indicating that AP-2 and Dlx 3, together with an additional
transcription factor(s) that are conserved between humans and mice, are
required for trophoblast-specific expression of 3-HSD VI.
*
This work was supported by NICHD, National Institutes of
Health cooperative agreement U54 HD 31398 as part of the Specialized Cooperative Centers Program in Reproductive Research.The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
-HSD VI gene has been submitted to the
GenBankTM/EBI Data Bank with accession number(s)
AY046511 (3256 bp promoter and exon 1) and AY046512 (1792 bp complete
cDNA).
To whom correspondence should be addressed: Div. of Reproductive
Biology, Dept. of Gynecology and Obstetrics, Stanford University School
of Medicine, 300 Pasteur Dr., Stanford, CA 94305-5317. Tel.:
650-725-6802; Fax: 650-725-7102; E-mail:
anita.payne@stanford.edu.
Copyright © 2002 by The American Society for Biochemistry and Molecular Biology, Inc.
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