Endoplasmic reticulum stress mediates withaferin A-induced apoptosis in human renal carcinoma cells

Abstract

The accumulation ofmisfolded proteins in the lumen of the endoplasmic reticulum(ER) results in cellular stress that initiates a specialized response designated as the unfolded protein response. ER stress has been implicated in a variety of common diseases, such as diabetes, ischemia and neurodegenerative dis- orders. Withaferin A, a major chemical constituent of Withania somnifera, has been reported to inhibit tumor cell growth. We show that withaferin A induced a dose-dependent apoptotic cell death in several types of human cancer cells, as measured by FACS analysis and PARP cleavage. Treatment of Caki cells with withaferin A induced a number of signature ER stress markers, including phosphorylation of eukary- otic initiation factor-2a (eIF-2 a), ER stress-specific XBP1 splicing, and up-regulation of glucose-regulated protein (GRP)-78. In addition, withaferin A caused up-regulation of CAAT/enhancer-binding protein- homologous protein (CHOP), suggesting the induction of ER stress. Pretreatment with N-acetyl cysteine (NAC) significantly inhibited withaferin A-mediated ER stress proteins and cell death, suggesting that reactive oxygen species (ROS) mediate withaferin A-induced ER stress. Furthermore, CHOP siRNA or inhi- bition of caspase-4 activity attenuated withaferin A-induced apoptosis. Taken together, the present study provides strong evidence supporting an important role of the ER stress response in mediating withaferin A-induced apoptosis. Keywords: Withaferin A Endoplasmic reticulum stress Apoptosis CHOP Unfolded protein response