Inhibition of NFκB Activity through Maintenance of IκBα Levels Contributes to Dihydrotestosterone-mediated Repression of the Interleukin-6 Promoter

Abstract

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 5␣-dihydrotestosterone, through the androgen receptor, repressed activation of the interleukin-6 promoter, in part, by inhibiting NFB activity. It did not appear that 5␣-dihydrotestosterone inhibited NFB by activating the androgen receptor to compete for the NFB 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 5␣-dihydrotestosterone repressed formation of NFB⅐NFB response element complex formation. In LNCaP prostate carcinoma cells, 5␣-dihydrotestosterone achieved this effect through maintenance of IB␣ protein levels in the face of phorbol ester, a stimulus that results in IB␣ degradation. Finally, we confirmed that IB␣ inhibits NFB-mediated activation of the interleukin-6 promoter. These data suggest that maintenance of IB␣ levels may represent the first identified mechanism for androgen-mediated repression of a natural androgen-regulated gene.Androgen hormones modulate expression of many genes, and both androgen-inducible (reviewed in Refs. 1, 2) and androgenrepressed (3-18) genes have been described. Androgen induction of transcription is mediated through the androgen receptor (AR), 1 a member of the steroid hormone receptor family. The AR is a 110-kDa nuclear protein (19,20) that consists of transactivation, DNA binding, nuclear localization, dimerization, and ligand binding domains (reviewed in Ref. 21). The AR activates gene transcription by specific binding to a DNA sequence, the androgen response element (ARE), in a liganddependent manner. The consensus ARE is similar to the glucocorticoid response element (GRE) (22) and as such can be stimulated by glucocorticoid receptor (GR). However, there are AREs that favor AR-induced transcriptional activation over that of . Although the ARE confers AR-mediated transcriptional activation on a gene, to date there has been no documentation of its ability to play a role in androgen-mediated repression of transcription.Transcriptional repression of genes is a vital component for modulation of gene expression in eukaryotes. Major mechanisms responsible for negative regulation of eukaryotic gene expression include cytoplasmic sequestration of transcription factors, blocking of transcription factor response elements, direct inhibition of transcription factors by protein-protein interactions, interference with trans-activation of DNA-bound transcription factors, and direct inhibition of transcription by binding to the promoter (reviewed in Refs. 27-33). Whether one of these mechanisms or some other mechanism accounts for AR's ability to repress transcrip...

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