Identification of androgen-selective androgen-response elements in the human aquaporin-5 and Rad9 genes.
The AR (androgen receptor) is known to influence the expression of its target genes by binding to different sets of AREs (androgen-response elements) in the DNA. One set consists of the classical steroid-response elements which are partial palindromic repeats of the 5'-TGTTCT-3' steroid-receptor monomer-binding element. The second set contains motifs that are AR-specific and that are proposed to be partial direct repeats of the same motif. On the basis of this assumption, we used an in silico approach to identify new androgen-selective AREs in the regulatory regions of known androgen-responsive genes. We have used an extension of the NUBIScan algorithm to screen a collection of 85 known human androgen-responsive genes compiled from literature and database searches. We report the evaluation of the most promising hits resulting from this computational search by in vitro DNA-binding assays using full-size ARs and GRs (glucocorticoid receptors) as well as their isolated DBDs (DNA-binding domains). We also describe the ability of some of these motifs to confer androgen-, but not glucocorticoid-, responsiveness to reporter-gene expression. The elements found in the aquaporin-5 and the Rad9 (radiation-sensitive 9) genes showed selective AR versus GR binding in band-shift assays and a strong activity and selectivity in functional assays, both as isolated elements and in their original contexts. Our data indicate the validity of the hypothesis that selective AREs are recognizable as direct 5'-TGTTCT-3' repeats, and extend the list of currently known selective elements.
Moehren
Udo
U
Laboratory of Molecular Endocrinology, Department of Molecular Cell Biology, Faculty of Medicine, Catholic University of Leuven, Campus Gasthuisberg, Onderwijs en Navorsing I, Box 901, Herestraat 49, B-3000 Leuven, Belgium.
Denayer
Sarah
S
Podvinec
Michael
M
Verrijdt
Guy
G
Claessens
Frank
F
D000728:Androgens / Q000378:metabolism*; D051402:Aquaporin 5 / Q000235:genetics / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D015894:Genome, Human / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000378:metabolism; D020218:Response Elements / Q000235:genetics*
D000728:Androgens; D051402:Aquaporin 5; D018797:Cell Cycle Proteins; D004247:DNA; D027425:ATP-Binding Cassette, Sub-Family C Proteins; C069930:rad9 protein; C421109:multidrug resistance-associated protein 1