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A comprehensive investigation into the molecular mechanism responsible for selective androgen receptor (SARM) tissue-selectivity

Goldberger, Natalie Elizabeth
http://rave.ohiolink.edu/etdc/view?acc_num=osu1202342545

Abstract Details

2008, Doctor of Philosophy, Ohio State University, Biophysics.
The androgen receptor (AR) is a member of the superfamily of nuclear receptors (NRs) that regulate gene expression in a ligand-dependent manner. Selective androgen receptor modulators (SARMs) are ligands that activate AR in a tissue-selective manner to produce unique physiological effects. To investigate the mechanism responsible for SARM tissue-selectivity, the conformation of AR was analyzed using the NHS-biotin labeling technique. Briefly, ligand-bound AR was affinity purified from Sf9 cells and incubated with NHS-biotin to biotinylate surface accessible lysine residues. Unfortunately, no biotinylation was observed for any of the AR samples, thus the possibility that AR chaperone proteins and cofactors were blocking NHS-biotin access to AR was subsequently investigated using native gels, immunoprecipitation (IP) and liquid chromatography coupled with ion trap mass spectroscopy (MS). IP and native gel analysis subsequently revealed multiple ligand-specific bands. The largest native gel protein bands (~ 690 kDa) for both the dihydrotestosterone- and SARM- treated samples were analyzed using tandem MS, and ligand-specific cofactors were identified. Most interesting was the identification of nuclear receptor corepressor (NCoR) in only the SARM-treated sample. This exciting result was subsequently confirmed in LNCaP when a higher degree of AR-SARM/NCoR versus AR-DHT/NCoR association was observed. Lastly, reduced cytoplasmic-nuclear transport and nuclear foci formation were observed by Western blot and immunofluorescence microscopy for AR-SARM versus AR-DHT. In summary, by using a combination of proteomics and immunofluorescence, support for the presence of functionally distinct AR-SARM protein complexes, in terms of AR cellular transport and transactivation, was generated. Most of all, the structural and functional differences revealed herein between AR-DHT and AR-SARM highlight potential avenues of research which may achieve a more comprehensive and detailed picture of the SARM tissue-selective mechanism.
James Dalton (Advisor)
1198 p.
SARM; Selective androgen receptor modulator; Androgen receptor; Proteomics; Mass spectroscopy; Native gel; Conformation; NHS-biotin; NCoR

Recommended Citations

Citations

  • Goldberger, N. E. (2008). A comprehensive investigation into the molecular mechanism responsible for selective androgen receptor (SARM) tissue-selectivity [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1202342545

    APA Style (7th edition)

  • Goldberger, Natalie. A comprehensive investigation into the molecular mechanism responsible for selective androgen receptor (SARM) tissue-selectivity. 2008. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1202342545.

    MLA Style (8th edition)

  • Goldberger, Natalie. "A comprehensive investigation into the molecular mechanism responsible for selective androgen receptor (SARM) tissue-selectivity." Doctoral dissertation, Ohio State University, 2008. http://rave.ohiolink.edu/etdc/view?acc_num=osu1202342545

    Chicago Manual of Style (17th edition)

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© 2008, all rights reserved.
This open access ETD is published by The Ohio State University and OhioLINK.