Hydrogen peroxide inducible clone-5/Androgen Receptor Activator 55 (Hic-5/ARA55) is a group III LIM domain protein that functions at focal adhesion complexes as well as in the nucleus as a nuclear receptor coactivator. Because the interaction of the androgen receptor (AR) with Hic-5/ARA55 results in enhanced androgen-induced transcription, we analyzed Hic-5/ARA55 expression in prostate tissue sections from normal human donors and prostate cancer patients. In each sample, Hic-5/ARA55 expression was confined to the stromal compartment of the prostate. Furthermore, in a human prostate stromal cell line (i.e. WPMY-1 cells) Hic-5/ARA55 was localized both at focal adhesion complexes and within the soluble cytoplasmic compartment. The ability of Hic-5/ARA55 to shuttle between the nuclear and cytoplasmic compartments within WPMY-1 cells was revealed upon inhibition of nuclear export with leptomycin B (LMB). siRNA ablation experiments established endogenous Hic-5/ARA55 as a coactivator for both viral and endogenous cellular AR-regulated genes. Furthermore, chromatin immunoprecipitation (ChIP) analysis showed androgen-dependent recruitment of Hic-/ARA55 to the promoter of the stromal androgen-responsive KGF gene. Using the A1-2 derivative of T47D breast cancer cells, we examined the mechanism by which Hic-5/ARA55 potentiates nuclear receptor transactivation. Hic-5/ARA55 was found to be an important component of glucocorticoid receptor (GR)-coactivator complexes in A1-2 cells since ablation of Hic-5/ARA55 expression by RNA interference-mediated silencing reduced GR transactivation. As shown by ChIP assays, Hic-5/ARA55 is recruited to glucocorticoid-responsive promoters of the MMTV, c-fos, and p21 genes in response to glucocorticoid treatment. Results from sequential ChIP assays established that Hic-5/ARA55 associates with the corepressor, NCoR, in the absence of glucocorticoids. However, upon glucocorticoid stimulation, Hic-5/ARA55 interacts with GR-coactivator containing complexes at these promoters. Ablation of Hic-5/ARA55 expression resulted in reduction of both TIF-2 and p300 recruitment to glucocorticoid-responsive promoters. These data provide the first demonstration of a stromal-specific AR coactivator that has an impact on an androgen regulated growth factor that is essential for stromal/epithelial cell communication in the prostate. Furthermore, these results suggest that Hic-5/ARA55 is required for optimal GR-mediated gene expression possibly by providing a scaffold that organizes or stabilizes coactivator complexes at some hormone-responsive promoters.
| Identifer | oai:union.ndltd.org:PITT/oai:PITTETD:etd-12192005-155456 |
| Date | 20 December 2005 |
| Creators | Heitzer, Marjet Danteel |
| Contributors | Mark Nichols, Donald DeFranco, Anthony Zeleznik, Alan Wells, William Walker |
| Publisher | University of Pittsburgh |
| Source Sets | University of Pittsburgh |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.library.pitt.edu/ETD/available/etd-12192005-155456/ |
| Rights | unrestricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to University of Pittsburgh or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
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