Abstract 1704: The co-chaperone, SGTA, acts as a regulator of androgen receptor activity in prostate cancer

Abstract

Abstract Introduction: Androgen signalling is essential for the initiation and progression of prostate cancer (PCa), which is exemplified by the response to hormonal based treatment strategies such as androgen ablation therapy, and the central role of the androgen receptor (AR) in tumor progression following biochemical relapse. Understanding the cellular components that regulate AR throughout the cell may lead to the identification of novel therapeutic targets. We recently described the biological actions of the co-chaperone SGTA on AR activity in PCa cells, and specifically that SGTA acts in the cytoplasm to limit or control AR response to agonist. The SGTA protein consists of a central tetratricopeptide (TPR) domain and a glutamine-rich (QRD) carboxyl terminus separated by a linker region (LR), and is thought to form a homodimer. However, the precise regions of SGTA that mediates its effects on AR signalling are currently unknown. The aim of this study was to map the SGTA residues required for homodimerisation, identify the requirement of different SGTA domains for an affect AR on activity and to determine the global effect of SGTA knockdown on androgen regulated genes in PCa cells. Methodology: Mammalian two hybrid assays, Co-IP, and native PAGE were used to identify amino acid residues required for SGTA homodimerisation. Transactivation assays using multiple AR responsive promoters were used to define the role of SGTA homodimerisation and individual SGTA structural domains on AR transcriptional output. Transient siRNA knockdown of SGTA combined with microarray analysis were used to assess the global effect of SGTA knockdown on androgen regulated genes. Results: Native PAGE analysis revealed that full length SGTA can exist as both a monomer (∼20%) and homodimer (∼80%) in mammalian cells. Deletion of the first 80 amino acids of SGTA abrogated homodimerisation, whereas deletion of the TPR and QRD regions had no effect. Although, the first 80 amino acids of SGTA were not sufficient to self dimerise or to form a homodimer with full length SGTA, its co-expression with full length SGTA was able to potently block dimerisation (p<0.001) in a dominant manner. Disruption of SGTA homodimerisation or deletion of the LR or QRD significantly affected the capacity of SGTA to regulate AR activity at low (0.1nM), but not at higher agonist concentrations. Moreover, only a subset of androgen regulated genes appear to be affected by SGTA siRNA knockdown. Conclusion: These analyses have identified regions of SGTA necessary for dimerisation and functional effects on AR transcriptional output, and support the underlying hypothesis that SGTA could be directly targeted by small molecular compounds or peptides as a novel means of modulating AR action in disease. This work was funded by the NHMRC (#45366 to WDT). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1704.