Essential role of ER stress-induced apoptosis in the anti-cancer effect of prodigiosin on human breast cancer cells

• Main Authors: Mu-Yun Pan, 潘慕芸
• Other Authors: Chia-Che Chang 張嘉哲
• Format: Others
• Language: zh-TW
• Published: 2012
• Online Access: http://ndltd.ncl.edu.tw/handle/72210762380006491467

碩士 === 國立中興大學 === 生物醫學研究所 === 100 === Endoplasmic Reticulum (ER) is the site for synthesis and folding of secretory proteins. Disturbance in the homeostasis of protein folding triggers unfolding protein response (UPR) and causes ER stress, ultimately leading to apoptosis if unresolved. Prodigiosin (PG) is a secondary metabolite of Serratia and Streptomyces and has been identified as a potential anti-cancer agent, mostly owing to induction of apoptosis selectively in some malignant cell lines. The mechanisms of the proapoptotic effect of prodigiosin remain elusive. In this study, we presented evidence supporting the role of ER stress in the anti-cancer effect of PG on human breast cancer cell lines. Quantitative real-time RT-PCR analysis and immunoblot analysis indicated that ER stress markers GRP78 and CHOP are dramatically induced by PG in MCF-7, MDA-MB-231 and T-47D cell lines, suggesting that PG induces ER stress. Furthermore, we found that the three pathway initiated by ER stress IRE1, PERK and ATF6 have been activated upon PG treatment, as evidenced by Ser724 phosphorylation of IRE1, XBP1 mRNA splicing, Thr183/Tyr185 dual phosphorylation of JNK, Ser51 phosphorylation of eIF2α and induction of ATF6 transcriptional activity, confirming that PG is an ER stresser. Notably, knockdown of CHOP by RNAi evidently decreased levels of PARP cleavage and cytotoxicity induced by PG, illustrating CHOP is the essential role of ER stress-induced apoptosis in PG’s anti-cancer effect. In addition, functional blockade of eIF2α and JNK by overexpession dominant-negative eIF2α and JNK-specific inhibitor SP600125 in MCF-7 cell, respectively, markedly suppressed PG-induced CHOP up-regulation. Moreover, we found prodigiosin-induced BCL2 suppression in a CHOP-dependent manner, and BCL2 overexpression greatly enhanced the survival of PG-treated cells. Taken together, we conclude that induction of ER stress-induced apoptosis is essential for the anti-cancer effect of PG, and PG engages PERK and JNK signaling pathways to up-regulate CHOP expression, leading to the induction of apoptosis by reducing BCL-2.