Androgen receptor signalling: comparative analysis of androgen response elements and implication of heat-shock protein 90 and 14-3-3eta.
Androgen receptor (AR) signalling was analysed using as models the cysteine-rich secretory protein-1 (CRISP-1) and CRISP-3 gene promoters, which are differentially regulated by androgen in vivo and contain multiple potential androgen response elements. Using electrophoretic mobility shift assay, we identified several elements with differing affinities for the AR at positions -3706, -1270, -1253 and -350 of the CRISP-1 promoter and at positions -369 and -349 of the CRISP-3 promoter. The strongest binding was observed for the -1253 element of CRISP-1. In transactivation assays using a PC-3 cell line stably transfected with the AR (PC-3/AR), the -1253 element placed as two or four copies upstream of the TK minimal promoter yielded a strong induction of luciferase reporter gene activity in the presence of the androgen methyltrienolone (R1881). In the context of the CRISP promoters a 2-fold induction by R1881 was measured for the CRISP-3 upstream region whereas only limited effects were noted for the CRISP-1 upstream region. The androgenic stimulation of the p(-1253 ARE)(4x)-TK-luciferase reporter construct was dose-dependently inhibited by geldanamycin and radicicol, two compounds that selectively interact with the chaperone protein, heat-shock protein 90. Cotransfection with an expression vector for the 14-3-3eta protein markedly enhanced the androgen-dependent stimulation. These results emphasize the influence of promoter context on androgen regulation and the importance of AR-associated proteins.
Research Laboratories of Schering AG, D-13342, Berlin, Germany. bernard.haendler@schering.de
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Haendler
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Schüttke
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Schleuning
D048948:14-3-3 Proteins; D016227:Benzoquinones; D002460:Cell Line; D004791:Enzyme Inhibitors / Q000494:pharmacology; D017930:Genes, Reporter / Q000235:genetics; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism*; D006801:Humans; D047029:Lactams, Macrocyclic; D007783:Lactones / Q000494:pharmacology; D018942:Macrolides; D008297:Male; D008562:Membrane Glycoproteins; D015741:Metribolone / Q000494:pharmacology; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics; D011467:Prostate / Q000187:drug effects / Q000378:metabolism; D011809:Quinones / Q000494:pharmacology; D011944:Receptors, Androgen / Q000378:metabolism*; D020218:Response Elements / Q000235:genetics*; D012471:Salivary Proteins and Peptides / Q000235:genetics; D029589:Seminal Plasma Proteins; D015398:Signal Transduction / Q000187:drug effects; D045165:Testosterone Congeners / Q000494:pharmacology; D015533:Transcriptional Activation / Q000187:drug effects; D014446:Tyrosine 3-Monooxygenase / Q000235:genetics / Q000378:metabolism*
D048948:14-3-3 Proteins; D016227:Benzoquinones; C103538:CRISP1 protein, human; C489926:CRISP3 protein, human; D004791:Enzyme Inhibitors; D018841:HSP90 Heat-Shock Proteins; D047029:Lactams, Macrocyclic; D007783:Lactones; D018942:Macrolides; D008562:Membrane Glycoproteins; D011809:Quinones; D011944:Receptors, Androgen; D012471:Salivary Proteins and Peptides; D029589:Seminal Plasma Proteins; D045165:Testosterone Congeners; C482984:YWHAH protein, human; D015741:Metribolone; D014446:Tyrosine 3-Monooxygenase; C035359:monorden; C001277:geldanamycin