502 IDENTIFICATION OF THE BINDING DOMAINS THAT ENABLE THE LIGAND-INDEPENDENT CO-ACTIVATION OF ANDROGEN RECEPTOR BY GLI2

You have accessJournal of UrologyProstate Cancer: Basic Research (III)1 Apr 2013502 IDENTIFICATION OF THE BINDING DOMAINS THAT ENABLE THE LIGAND-INDEPENDENT CO-ACTIVATION OF ANDROGEN RECEPTOR BY GLI2 Na Li, Mengqian Chen, and Ralph Buttyan Na LiNa Li Vancouver, Canada More articles by this author , Mengqian ChenMengqian Chen Columbia, SC More articles by this author , and Ralph ButtyanRalph Buttyan Vancouver, Canada More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2013.02.1896AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Gli proteins are transcription factors that mediate Hedgehog signaling. Gli activity guides cell fate decisions during development and is required for the proper morphogenesis of embryonic tissues, including the prostate gland. Previously, we showed that exogenous overexpression of Gli1 or Gli2 in androgen-deprived LNCaP cells increased the expressions of androgen-dependent genes in prostate cancer (PCa) cells and conferred androgen-independent growth (Chen et al., Mol Cancer, 9: 89-101, 2010). We also showed that Gli1 or Gli2 proteins co-immunoprecipitate with AR, suggesting that AR interacts with Gli proteins to mediate these effects. Based upon this data, we proposed that Gli1/2 proteins are ligand-independent co-activators of AR in PCa. To better understand the Gli2-AR interaction and its effects on PCa responsiveness to androgen ablation, we fine mapped the specific interaction domains between Gli2 and AR. METHODS Complete or partial recombinant Gli2 cDNAs were tagged with GST and the GST-tagged proteins were purified on glutathione beads prior to incubation with in vitro translated human AR protein fragments. Domain-specific Gli2 polypeptides were tested for their ability to co-activate AR in transient expression assays using luciferase reporter vectors under the control of AR-dependent gene promoters and enhancers in 293T cells. Gli2 was tested for its ability to co-activate natural truncated variants of the AR. RESULTS GST pulldown assays showed that GLI2 specifically recognizes and binds the Tau5 region within the AR N-terminal domain (AA 391-555). Likewise, AR binds to Gli2 within an extended C-terminal domain (AA 628-1091) that may involve 2 distinct AR binding sites. AR co-activation studies in 293T cells showed that the presence of both the AR binding site(s) and the Gli2 activation domain are required for Gli2 co-activation of AR. The interaction of Gli2 with the Tau4 domain of AR was consistent with the ability of Gli2 to co-activate truncated variants of the AR that lack the DNA binding domain. CONCLUSIONS Gli2 co-activation of AR requires direct interaction between the tau5 domain in the AR N-terminus and the AR binding domains in the Gli2 C-terminus as well as the Gli2 activation domain. Gli2 binding at tau5 supports co-activation of the wildtype AR (in both the presence or absence of androgen) as well as co-activation of truncated receptors that do not bind androgens. As such, the evidence that Gli2 expression is increased in PCa by androgen ablation suggests that it may be a factor in progression to castration resistant PCa. © 2013 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 189Issue 4SApril 2013Page: e206 Advertisement Copyright & Permissions© 2013 by American Urological Association Education and Research, Inc.MetricsAuthor Information Na Li Vancouver, Canada More articles by this author Mengqian Chen Columbia, SC More articles by this author Ralph Buttyan Vancouver, Canada More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

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