Abstract 4073: Proteasomal degradation of JunD is required for TGF-β induced inhibition of proliferation in prostate cancer cells

Transforming growth factor-β (TGF-β) inhibits proliferation of prostate epithelial cells. However, prostate cancer cells in advanced stages become resistant to TGF-β effects on proliferation. In this study, we have investigated the role of AP-1 family of transcription factors, specifically the Jun family members (c-Jun, JunB and JunD) in TGF-β1 effects on prostate cancer cell proliferation. Basal expression of individual Jun family members was determined by RT-PCR and Western blot analysis in four prostate cell lines: RWPE1, LNCaP, DU145 and PC3 cells. Jun levels were also examined in response to TGF-β1. c-Jun was expressed in all cell lines and its expression was up-regulated by TGF-β1. Levels of JunB were higher in normal cells compared to those in cancer cells and TGF-β1caused an increase in JunB levels. JunD levels were higher in all cell lines; TGF-β1 had differential effects on JunD protein levels. While it caused a significant decrease in JunD levels in RWPE-1 and DU145 cells, it did not affect JunD levels in PC3 cells. These differential effects on JunD levels correlated with differential effects of TGF-β on cell proliferation. Individual Jun family members were overexpressed in DU145 cells and effects on cell proliferation were determined. Over-expression of c-Jun and JunB decreased proliferation rate in DU145 cells; however, overexpression of JunD increased proliferation in these cells. Interestingly, cells over-expressing JunD still succumb to inhibitory effect of TGF-β1 after six to eight days of incubation. Furthermore, silencing JunD by siRNA decreased proliferation in both DU145 and PC3 cells. To determine the molecular mechanism of TGF-β-induced down-regulation of JunD, we pretreated cells over-expressing JunD with a proteosomal inhibitor, MG132. Pretreatment of MG132 blocked the degradation of JunD in in DU145 cells over-expressing JunD. In conclusion, our studies show that specific Jun family exerts differential effects on proliferation in prostate cancer cells in response to TGF-β1. While c-Jun and Jun B mediate the inhibitory effects of TGF-β1 on proliferation, JunD counteracts these effects. TGF-β1 causes inhibition in proliferation by degrading JunD via proteasomal degradation. Acknowledgements: These studies were supported by the NIH/NIMHD/RCMI grant #G12MD007590, NIH/NIMHD #5P20MD002285, and DOD grant # W8I-08-1-0077. Citation Format: Ana C. Millena, BaoHan T. Vo, Nicole Strong, Lindsey Walker, Yang Cao, Natalya Klueva, Curt Pfarr,