Identification and characterization of proteins that interact with the NH2-Terminus of the Androgen Receptor

The overall goal of the research described in this thesis is to gain a more comprehensive understanding of androgen receptor (AR) action in prostate cancer. Prostate cancer is among the most commonly diagnosed non-skin cancer and the second leading cause of cancer-related death in men. Although prostate cancer is often curable in its early stages, limited treatment options are available for men that present with advanced disease. Androgens are the male sex hormones that are essential for normal growth, development and maintenance of the prostate gland, and may have a role in prostate cancer progression. Androgens carry out their function through the AR and significant effort has been made to gain a better understanding of the accessory proteins that interact with AR to regulate and modify receptor responses. Our laboratory has used conventional and novel yeast two-hybrid assay systems to identify proteins that interact with regions of AR that have traditionally been hard to analyze. Essential elements of these assay systems are discussed in this thesis. Two proteins, RanBPM and cyclin G-associated kinase (GAK), were identified as AR-interacting proteins using these yeast two-hybrid systems and co-immunoprecipitation assays were used to confirm in vivo interactions. Interacting domains between these two proteins and AR were determined by in vitro GST and His-tag pull-down assays. Both RanBPM and GAK were found to interact with multiple domains of AR. Specific questions regarding the role of RanBPM and GAK in AR transactivation were addressed using transcription assays. Both proteins act as co-activators that enhance the NH2-terminal ligand-independent transactivation function of AR. Finally, involvement and prognostic value of RanBPM and GAK in advanced disease were assessed using two prostate cancer progression models. This research provides insight into the role of co-activator function in AR-mediated transcription and the possible role of co-activators in disease progression.Medicine, Faculty ofPathology and Laboratory Medicine, Department ofGraduat