The objective of this proposal is to define the functions of novel and critical components of the androgen regulated transcriptome in the patho-biology of prostate gland. Specifically, we will establish expression characteristics and functions of PMEPA1, a novel androgen responsive gene (ARG) discovered in our laboratory by Serial Analysis of Gene Expression in androgen treated LNCaP cells. Background: PMEPA1 expression is prostate-abundant and is restricted to prostatic epithelial cells. PMEPA1 binds to NEDD4 protein (ubiquitin E3 ligase) involved in the ubiquitin-proteasome pathway. PMEPA1 overexpression down-regulates AR protein in LNCaP cells and PMEPA1 negatively regulates prostate cancer cell growth. PMEPA1 also binds to AR. Loss or reduced PMEPA 1 expression is noted in 65% of primary CaP specimens. Our central hypothesis is that PMEPA1, as an androgen responsive-NEDD4 binding protein, down regulates the Androgen Receptor (AR) protein by a feed back loop between PMEPA1 and AR, which plays a critical role in maintaining the AR levels and prostatic epithelial cell growth. We propose to define PMEPA 1 functions and expression in prostate cancer cells by following specific aims: (1) Mechanisms of regulation of PMEPA1 expression by AR: The androgen responsive elements in PMEPA1 promoter will be defined by ChiP assays and PMEPA1 promoter-luciferase assays. Binding of AR to PMEPA1 promoter will also be analyzed in the genome of benign and malignant prostate epithelium; (2) PMEPA1 functions in negative regulation of AR and prostate cancer cell growth: PMEPA1 mediated regulation of AR and the AR-PMEPA1-NEDD4 connection will be established. Ubiquitination of PMEPA1 and AR will be analyzed. Cell growth inhibitory properties of PMEPA1 will be characterized in the AR and NEDD4 context using cell culture and animal models; and (3) PMEPA1 and AR expression features in human prostate pathobiology and their biomarker potential: PMEPA1 and AR expression in prostate cancer cells will be analyzed by real time PCR and IHC and will be correlated with clinico-pathologic features and evaluated for prognostic value. ? Relevance: PMEPA1 is a prostate abundant novel ARG and is a NEDD4 binding protein that inhibits CaP cell growth in vitro. Characterization of the functions and expression profiles of PMEPA 1 will enhance our knowledge of this gene in prostate tumor biology and in androgen signaling pathways. These goals are highly relevant in research defining prostate cancer biology and in the development of potentially new biomarkers and therapeutic targets for CaP. ? ?
