Abstract 5574: Gli2 protein and the AR operational switch to the castration resistant prostate cancer transcription program

Abstract

Abstract Gli proteins are transcription factors that mediate Hedgehog (HH) signaling in cells. Previously, we reported that Gli2 binds to the androgen receptor (AR) protein in prostate cancer (PCa) cells and further upregulates expression of genes under AR control. This “co-activator” effect allows the growth of Gli2 overexpressing LNCaP cells in androgen-depleted medium, mimicking the behavior of castrate resistant variants of PCa. We report here our success in localizing the specific binding domains between Gli2 and AR proteins. Outcomes of GST-pulldown and co-immunoprecipitation studies showed that Gli2 binds to AR via a limited domain (aa628-aa897) within the C-terminal region and this recognizes the tau5/AF5 domain (aa 392-558) within the N-terminal region of the AR protein. Because tau5/AF5 is conserved in truncated (splice variant) ARs, we also showed that Gli2 recognizes and co-activates the transcriptional activity of a truncated variant AR (AR-V7). To further define Gli2 regulatory functions on AR transcriptional activity in CRPC, we overexpressed Gli2ΔN (a constitutive-active form of Gli2) in LNCaP-AI (androgen-independent) cells and examined its effects on AR target gene expressions. Surprisingly, we found that Gli2Δ overexpression coincided with significant downregulated expression of genes considered “traditional” AR targets (i.e., KLK3 and KLK2), while increasing the expression of M-phase check-point genes, such as UBE2C, CDK1 and CDC20. In contrast, treatment of unmodified LNCaP-AI cells with the Gli-inhibitor, GANT61, significantly increased expression of KLK2 and KLK3 while downregulating UBE2C, CDC20 and CDK1. Our outcomes now imply that Gli2 differentially affects the AR transcriptional program in androgen-dependent (LNCaP) vs androgen-independent (LNCaP-AI) cells. In androgen-dependent cells, Gli2 co-activates expression of the traditional AR target genes while suppressing expression of genes involved in the CRPC transcriptional program whereas in androgen-independent cells, Gli2 co-activates expression of CRPC program genes while downregulating expression of traditional AR target genes. This dichotomy suggests that Gli2 participates in the decisions through which AR transcriptional activity is redirected in castration resistant disease. Supported by the DOD PCRP (W81XH-10-1-0493, to RB) Citation Format: Na Li, Ralph Buttyan, Sarah Truong, Yue Yu, Mengqian Chen. Gli2 protein and the AR operational switch to the castration resistant prostate cancer transcription program. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5574. doi:10.1158/1538-7445.AM2014-5574