%0 Journal Article
%T Celastrol targets mitochondrial respiratory chain complex I to induce reactive oxygen species-dependent cytotoxicity in tumor cells
%A Guozhu Chen
%A Xuhui Zhang
%A Ming Zhao
%A Yan Wang
%A Xiang Cheng
%A Di Wang
%A Yuanji Xu
%A Zhiyan Du
%A Xiaodan Yu
%J BMC Cancer
%D 2011
%I BioMed Central
%R 10.1186/1471-2407-11-170
%X The downregulation of heat shock protein 90 (HSP90) client proteins, phosphorylation of c-Jun NH2-terminal kinase (JNK), and cleavage of PARP, caspase 9 and caspase 3 were detected by western blotting. The accumulation of reactive oxygen species (ROS) was analyzed by flow cytometry and fluorescence microscopy. Cell cycle progression, mitochondrial membrane potential (MMP) and apoptosis were determined by flow cytometry. Absorption spectroscopy was used to determine the activity of mitochondrial respiratory chain (MRC) complexes.Celastrol induced ROS accumulation, G2-M phase blockage, apoptosis and necrosis in H1299 and HepG2 cells in a dose-dependent manner. N-acetylcysteine (NAC), an antioxidative agent, inhibited celastrol-induced ROS accumulation and cytotoxicity. JNK phosphorylation induced by celastrol was suppressed by NAC and JNK inhibitor SP600125 (SP). Moreover, SP significantly inhibited celastrol-induced loss of MMP, cleavage of PARP, caspase 9 and caspase 3, mitochondrial translocation of Bad, cytoplasmic release of cytochrome c, and cell death. However, SP did not inhibit celastrol-induced ROS accumulation. Celastrol downregulated HSP90 client proteins but did not disrupt the interaction between HSP90 and cdc37. NAC completely inhibited celastrol-induced decrease of HSP90 client proteins, catalase and thioredoxin. The activity of MRC complex I was completely inhibited in H1299 cells treated with 6 ¦ÌM celastrol in the absence and presence of NAC. Moreover, the inhibition of MRC complex I activity preceded ROS accumulation in H1299 cells after celastrol treatment.We identified ROS as the key intermediate for celastrol-induced cytotoxicity. JNK was activated by celastrol-induced ROS accumulation and then initiated mitochondrial-mediated apoptosis. Celastrol induced the downregulation of HSP90 client proteins through ROS accumulation and facilitated ROS accumulation by inhibiting MRC complex I activity. These results identify a novel target for celastrol-in
%U http://www.biomedcentral.com/1471-2407/11/170