Abstract 1722: The outer nuclear membrane population of Bcl-2 contributes to drug resistance in human prostate cancer cells.

Abstract

Abstract Bcl-2 is the archetypal gene and first characterized member of the family bearing its name. Prostate cancer was the first solid tumor malignancy in which an elevation of cellular Bcl-2 levels was associated with advanced disease. It is thought that the anti-apoptotic activity of Bcl-2 contributes to the uniformly lethal nature of castration-resistant advanced prostate cancer. Bcl-2 is membrane-bound and located on the cytosolic face of the mitochondrial outer membrane (MOM), endoplasmic reticulum (ER), and outer nuclear membrane (ONM). The interest in functions of Bcl-2 have centered on its maintenance of MOM integrity. Recently, the Bcl-2 population on the ER has been shown to control calcium ion release into the cytosol through its interactions with the inositol 1,4,5-trisphosphate receptor (IP3R). In contrast to the well known and growing functions of MOM and ER Bcl-2 populations respectively, the role of Bcl-2 on the ONM has yet to be defined. While progress in our understanding of MOM and ER Bcl-2 has been made, in part, by the targeted overexpression of the protein to the MOM and ER, no such model systems have been utilized to answer questions of Bcl-2 on the ONM. We believe this has limited our understanding of the ONM Bcl-2 population. With this deficit in our understanding in mind, we sought to generate a Bcl-2 fusion gene construct that would, for the first time, enable the targeted expression of the protein to the ONM. The Bcl-2 cDNA was mutagenized so that the sequence representing its C-terminal transmembrane (TM) domain was replaced by a consensus sequence derived from a family of recently characterized genes known to specifically localize to the ONM. The ONM Bcl-2 mutant, housed in a eukaryotic expression construct, was stably expressed in LNCaP and CWR22Rv1 human prostate cancer cell lines. Clones verified to stably express ONM Bcl-2, in addition to other clones expressing the WT gene, a TM-deleted variant, and previously described MOM- and ER-specific variants, were challenged in vitro by a variety of apoptotic stimuli. We successfully showed the stable targeted expression of Bcl-2 to the ONM, in addition to the MOM and ER in human prostate cancer cell lines. Treatment of these clones with several agents resulted in enhanced resistance to death that was location dependant. We have developed a useful system to help enhance our understanding of Bcl-2. The ONM population appears to contribute to drug resistance in prostate cancer cells. The mechanism responsible for ONM Bcl-2-induced apoptotic resistance may differ from its role on other membranes, and warrants further investigation. Citation Format: Theodore E. Ewachiw, Jennifer R. Cohen, Simon A. Williams. The outer nuclear membrane population of Bcl-2 contributes to drug resistance in human prostate cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1722. doi:10.1158/1538-7445.AM2013-1722