Proteasome inhibitor-I enhances tunicamycin-induced chemosensitization of prostate cancer cells through regulation of NF-κB and CHOP expression Pham Thi Thu Huong a , Dong-Oh Moon a , Sun Ok Kim a , Kyoon Eon Kim b , Sook Jung Jeong a , Ki Won Lee c,f , Kyung Sang Lee d , Jae Hyuk Jang a , Raymond Leo Erikson e , Jong Seog Ahn a, ⁎, Bo Yeon Kim a,f, ⁎⁎ a Chemical Biology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), 685-1 Yangcheonri, Ochangeup, Cheongwongun, 363-883, Republic of Korea b Department of Biochemistry, College of Natural Sciences, ChungNam National University, 220 Gung-dong, Yuseong-gu, Daejeon, 305-764, Republic of Korea c Department of Agricultural Biotechnology, Center for Agricultural Biomaterials, Seoul National University, Seoul 151-921, Republic of Korea d Laboratory of Metabolism, National Cancer Institute, NIH, Rockville Pike, Bethesda, MD 20892-4258, USA e Department of Molecular and Cellular Biology, Harvard University, Cambridge, MA 02138, USA f World Class Institute, Korea Research Institute of Bioscience and Biotechnology, Republic of Korea abstract article info Article history: Received 8 December 2010 Accepted 13 January 2011 Available online 27 January 2011 Keywords: Proteasome inhibitor-1 Tunicamycin ER-stress NF-κB CHOP Although endoplasmic reticulum (ER) stress induction by some anticancer drugs can lead to apoptotic death of cancer cells, combination therapy with other chemicals would be much more efficient. It has been reported that proteasome inhibitors could induce cancer cell death through ER-stress. Our study, however, showed a differential mechanism of proteasome inhibitor-I (Pro-I)-induced cell death. Pro-I significantly enhanced apoptotic death of PC3 prostate cancer cells pretreated with tunicamycin (TM) while other signaling inhibitors against p38, mitogen activated kinase (MEK) and phosphatidyl-inositol 3-kinase (PI3K) did not, as evidenced by cell proliferation and cell cycle analyses. NF-κB inhibition by Pro-I, without direct effect on ER-stress, was found to be responsible for the TM-induced chemosensitization of PC3 cells. Moreover, TM-induced/enhancer-binding protein (C/EBP) homologous protein (CHOP) expression was enhanced by Pro-I without change in GRP78 expression. CHOP knockdown by siRNA also showed a significant decrease in Pro-I chemosensitization. All these data suggest that although TM could induce both NF-κB activation and CHOP expression through ER-stress, both NF-κB inhibition and increased CHOP level by Pro-I are required for enhanced chemosensitization of PC3 prostate cancer cells. Thus, our study might contribute to the identification of anticancer targets against prostate cancer cells. © 2011 Elsevier Inc. All rights reserved. 1. Introduction NF-κB is activated by a variety of signals through mechanisms that result in phosphorylation and proteasomic degradation of the inhibitory IκBα protein [1]. It has been reported that IκBα mutant- mediated inhibition of NF-κB nuclear translocation enhanced apo- ptotic killing of human fibro-sarcoma and pancreatic carcinoma cells by certain chemotherapeutic drugs [2,3]. Regarding the IκBα regulation, many chemical agents, referred to as biological response or resistance modifiers, have been demonstrated to alter chemo- sensitivity in refractory tumor cells and are potentially useful in clinical cancer therapeutics [4]. Although ubiquitin–proteasome pathway is an efficient therapeutic target for cancer treatment [5,6], and recently developed proteasome inhibitors have been potent anti- cancer agents for various cancers, their underlying molecular mechanism remains to be elucidated. Recently, endoplasmic reticulum (ER) has emerged a prominent target for the development of chemotherapeutics against diverse diseases including diabetes and cancer. Tunicamycin (TM), a nucleoside antibiotic, inhibits the first step in the biosynthesis of N-linked oligosaccharides in cells, resulting in the ER stress induction and apoptotic cell death by unfolded protein response (UPR) in certain cancer cells [7–11]. CCAAT/enhancer-binding protein (C/EBP) homolo- gous protein (CHOP), also called GADD153, implicated in the regulation of processes relevant to energy metabolism, cellular proliferation, differentiation, and expression of cell-type-specific genes, is primarily pro-apoptotic and is one of the highly inducible genes during ER stress [12,13]. In many cases of ER stress, induction or over-expression of CHOP sensitizes cells to ER stress-induced apoptosis, whereas CHOP deletion protects the cells from apoptosis. The pro-apoptotic effect of CHOP in ER stress may be mediated by the induction of growth arrest and DNA damage gene 34 (GADD34) that dephosphorylates eukaryotic translation initiation factor 2α (eIF2α) [12]. These results suggested that CHOP regulation could be critical for the treatment of cancer. Cellular Signalling 23 (2011) 857–865 ⁎ Correspondence to: J.S. Ahn, Korea Research Institute of Bioscience and Biotech- nology (KRIBB), 685-1 Yangcheonri, Ochangeup, Cheongwongun, 363-883, Republic of Korea. Tel.: +82 43 240 6160. ⁎⁎ Correspondence to: B.Y. Kim, World Class Institute, Korea Research Institute of Bioscience and Biotechnology, Ochangeup, Republic of Korea. Tel.: + 82 43 240 6163. E-mail addresses: jsahn@kribb.re.kr (J.S. Ahn), bykim@kribb.re.kr (B.Y. Kim). 0898-6568/$ – see front matter © 2011 Elsevier Inc. All rights reserved. doi:10.1016/j.cellsig.2011.01.010 Contents lists available at ScienceDirect Cellular Signalling journal homepage: www.elsevier.com/locate/cellsig