Identification of phosphorylated proteins involved in the oncogenesis of prostate cancer via Pin1-proteomic analysis

BACKGROUND The peptidyl-prolyl isomerase Pin1 regulates a subset of phosphorylated proteins by catalyzing the cis-trans isomerization of their specific phosphorylated Ser/Thr-Pro motifs. Although Pin1 has been shown to be involved in cell transformation and the maintenance of the malignant phenotype in prostate cancer, its specific substrates during these processes have not yet been determined. METHODS. Cancer-specific phosphorylated proteins were isolated from two human prostate cancer cell lines (PC-3, LNCaP) and the Dunning rat prostate cancer cell lines by GST-pull down analysis with recombinant GST-Pin1 protein. These proteins were then identified by the LC-MS/MS analysis using a Q-Tof micro mass spectrometer and processed for further functional analysis. RESULTS. We newly identified five prostate cancer-specific Pin1 binding proteins (PINBPs) in this screen. Among these, TRK-fused gene (TFG) was found to be preferentially up-regulated in prostate cancer cell lines and tissues. The targeted inhibition of TFG by specific siRNA resulted in the reduced cell proliferation and the induction of premature senescence in PC3 prostate cancer cells. We further found that TFG can facilitate the cell signaling mediated by NF-kappaB and androgen receptor (AR). Tissue micro-dissection based quantitative RT-PCR analysis of prostate cancer tissues following radical prostatectomy further revealed that TFG expression is closely associated with both a higher probability and shorter period of tumor recurrence following surgery. CONCLUSIONS. Pin1-based proteomics analysis is a useful tool for the identification of prostate cancer-specific phosphorylated proteins. TFG could be a potential diagnostic and/or prognostic marker and therapeutic target in prostate cancer. Prostate 72:626-637, 2012. (C) 2011 Wiley Periodicals, Inc.