P68/DdX5 Supports β-Catenin &amp; RNAP II during Androgen Receptor Mediated Transcription in Prostate Cancer

Emma L Clark,Frances V Fuller-Pace,Richard Temperley,Christiana Hadjimichael,Craig N Robson,Amy Barnard,Jean-Marc A Lobaccaro

doi:10.1371/journal.pone.0054150

Abstract

The DEAD box RNA helicase p68 (Ddx5) is an important androgen receptor (AR) transcriptional co-activator in prostate cancer (PCa) and is over-expressed in late stage disease. β-Catenin is a multifunctional protein with important structural and signalling functions which is up-regulated in PCa and similar to p68, interacts with the AR to co-activate expression of AR target genes. Importantly, p68 forms complexes with nuclear β-Catenin and promotes gene transcription in colon cancer indicating a functional interplay between these two proteins in cancer progression. In this study, we explore the relationship of p68 and β-Catenin in PCa to assess their potential co-operation in AR-dependent gene expression, which may be of importance in the development of castrate resistant prostate cancer (CRPCa). We use immunoprecipitation to demonstrate a novel interaction between p68 and β-Catenin in the nucleus of PCa cells, which is androgen dependent in LNCaP cells but androgen independent in a hormone refractory derivative of the same cell line (representative of the CRPCa disease type). Enhanced AR activity is seen in androgen-dependent luciferase reporter assays upon transient co-transfection of p68 and β-Catenin as an additive effect, and p68-depleted Chromatin-Immunoprecipitation (ChIP) showed a decrease in the recruitment of the AR and β-Catenin to androgen responsive promoter regions. In addition, we found p68 immunoprecipitated with the processive and non-processive form of RNA polymerase II (RNAP II) and show p68 recruited to elongating regions of the AR mediated PSA gene, suggesting a role for p68 in facilitating RNAP II transcription of AR mediated genes. These results suggest p68 is important in facilitating β-Catenin and AR transcriptional activity in PCa cells.

Highlights

The onset and progression of prostate cancer (PCa) is driven by the transcriptional function of the androgen receptor (AR), and ablation of androgens is an effective strategy at early stages of the disease [1]
We describe a functional interaction between p68, b-Catenin and the AR in the nucleus of PCa cells
The AR has been shown to signal through the Wnt/b-Catenin pathway in PCa as an adaptation to castrate levels of androgens [12], and bCatenin is known to interact with other co-activators of the AR

Summary

Introduction

The onset and progression of prostate cancer (PCa) is driven by the transcriptional function of the androgen receptor (AR), and ablation of androgens is an effective strategy at early stages of the disease [1]. Further evidence suggests that the growth of metastatic prostate tumour cells in the bone is via androgen mediated Wnt activation [13], and increased nuclear b-Catenin levels have been correlated with prostate cancer disease progression [14,15]. P68 and the highly homologous protein p72 (Ddx17), are over-expressed and form complexes with bCatenin in the nucleus of colon cancer cells to activate gene transcription and promote cell proliferation [22,23,24]. Endogenous protein immunoprecipitated from nuclear LNCaP extracts showed the b-Catenin-p68 interaction was facilitated in the presence of androgens (R1881, 10 nM), but in the LNCaP-AI cell line the interaction was facilitated in the absence of androgens (R1881, 10 nM) (Figure 1C).

Results

Discussion

Materials and Methods