Inhibition of NFkappaB activity through maintenance of IkappaBalpha levels contributes to dihydrotestosterone-mediated repression of the interleukin-6 promoter.
Androgens repress expression of many genes, yet the mechanism of this activity has remained elusive. The cytokine, interleukin-6, is active in a variety of biological systems, and its expression is repressed by androgens. Accordingly we dissected the mechanism of androgen's ability to inhibit interleukin-6 expression at the molecular level. In a series of co-transfection assays, we found that 5alpha-dihydrotestosterone, through the androgen receptor, repressed activation of the interleukin-6 promoter, in part, by inhibiting NFkappaB activity. It did not appear that 5alpha-dihydrotestosterone inhibited NFkappaB by activating the androgen receptor to compete for the NFkappaB response element as we could not detect androgen receptor binding to the IL-6 promoter by DNase I footprinting assay. However, by electrophoretic mobility shift assay we found that 5alpha-dihydrotestosterone repressed formation of NFkappaB middle dotNFkappaB response element complex formation. In LNCaP prostate carcinoma cells, 5alpha-dihydrotestosterone achieved this effect through maintenance of IkappaBalpha protein levels in the face of phorbol ester, a stimulus that results in IkappaBalpha degradation. Finally, we confirmed that IkappaBalpha inhibits NFkappaB-mediated activation of the interleukin-6 promoter. These data suggest that maintenance of IkappaBalpha levels may represent the first identified mechanism for androgen-mediated repression of a natural androgen-regulated gene.
Keller
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ET
Institute on Aging, Department of Medicine, University of Wisconsin, Madison, Wisconsin 53706, USA.
Chang
C
C
Ershler
W B
WB
D002460:Cell Line; D018983:DNA Footprinting; D004268:DNA-Binding Proteins / Q000378:metabolism*; D013196:Dihydrotestosterone / Q000494:pharmacology*; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D005786:Gene Expression Regulation; D006367:HeLa Cells; D006801:Humans; D020672:I-kappa B Proteins; D015850:Interleukin-6 / Q000235:genetics*; D008297:Male; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000037:antagonists & inhibitors*; D011401:Promoter Regions, Genetic / Q000187:drug effects*; D011471:Prostatic Neoplasms / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011944:Receptors, Androgen / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects
D004268:DNA-Binding Proteins; D020672:I-kappa B Proteins; D015850:Interleukin-6; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D012333:RNA, Messenger; D011944:Receptors, Androgen; D013196:Dihydrotestosterone; D000072000:NF-KappaB Inhibitor alpha