1. ¹The Laboratory of Apoptosis and Cancer Biology, State Key Laboratory of Biomembrane and Membrane Biotechnology, Institute of Zoology, The Graduate School of Chinese Academy of Sciences, Chinese Academy of Sciences, Datun Rd, Chao Yang District, Beijing 100101, China
2. ²College of Life Sciences, Nan Kai University, Tianjin 300071, China
3. ³Department of Neuro-Oncology, MD Anderson Cancer Center, 1515 Holcombe Blvd., Houston, TX 77030, USA

Correspondence: Quan Chen, Tel: +86 10 6480 7321 E-mail: chenq@ioz.ac.cn

Received 31 July 2007; Revised 10 October 2007; Accepted 30 October 2007.

Abstract

Intracellular redox homeostasis plays a critical role in determining tumor cells' sensitivity to drug-induced apoptosis. Here we investigated the role of thioredoxin-1 (TRX1), a key component of redox regulation, in arsenic trioxide (As₂O₃)-induced apoptosis. Over-expression of wild-type TRX1 in HepG₂ cells led to the inhibition of As₂O₃-induced cytochrome c (cyto c) release, caspase activation and apoptosis, and down-regulation of TRX1 expression by RNAi sensitized HepG₂ cells to As₂O₃-induced apoptosis. Interestingly, mutation of the active site of TRX1 from Cys^32/35 to Ser^32/35 converted this molecule from an apoptotic protector to an apoptotic promoter. In an effort to understand the mechanisms of this conversion, we used isolated mitochondria from mouse liver and found that recombinant wild-type TRX1 could protect mitochondria from the apoptotic changes. In contrast, the mutant form of TRX1 alone elicited mitochondria-related apoptotic changes, including the mitochondrial permeability transition pore (mPTP) opening, loss of mitochondrial membrane potential, and cyto c release from mitochondria. These apoptotic effects were inhibited by cyclosporine A (CsA), indicating that mutant TRX1 targeted to mPTP. Alteration of TRX1 from its reduced form to oxidized form in vivo by 2,4-dinitrochlorobenzene (DNCB), a specific inhibitor of TRX reductase, also sensitized HepG₂ cells to As₂O₃-induced apoptosis. These data suggest that TRX1 plays a central role in regulating apoptosis by blocking cyto c release, and inactivation of TRX1 by either mutation or oxidization of the active site cysteines may sensitize tumor cells to As₂O₃-induced apoptosis.