Withaferin A induced impaired autophagy and unfolded protein response in human breast cancer cell-lines MCF-7 and MDA-MB-231

• Published in: Toxicology in vitro 2017-10, Vol.44, p.330-338
• Main Authors: Ghosh, Kamalini, De, Soumasree, Mukherjee, Srimoyee, Das, Sayantani, Ghosh, Amar Nath, Sengupta, Sumita (Bandyopadhyay)
• Format: Article
• Language: English
• Subjects: Accumulation; Acetylation; Autophagy; Autophagy - drug effects; Ayurvedic medicine; Biodegradation; Biological activity; Breast cancer; Breast Neoplasms - metabolism; Cancer; Caspase; Caspase-4; Cell cycle; Cell Line, Tumor; Computer networks; Degradation; Endoplasmic reticulum; Endoplasmic Reticulum Stress - drug effects; ER-stress; Heat-Shock Proteins - metabolism; Human behavior; Humans; Incomplete autophagy; Inhibition; Intracellular; Lysosomes; Lysosomes - metabolism; Microtubule-associated protein 2; Microtubule-Associated Proteins - metabolism; Phagocytosis; Phagosomes; Polymerization; Proteasomes; Protein biosynthesis; Protein folding; Protein synthesis; Proteins; Stress; Stresses; Substrates; Transcription Factor CHOP - metabolism; Tubulin; Ubiquitin; Ubiquitine proteasome system; Unfolded protein response; Unfolded Protein Response - drug effects; Withanolides - toxicity; X-Box Binding Protein 1 - metabolism
• ISSN: 0887-2333; 1879-3177
• DOI: 10.1016/j.tiv.2017.07.025

The autophagy-lysosome pathway and the ubiquitin-proteasome systems are the two major routes for eukaryotic intracellular protein clearance. Cancerous cells often display elevated protein synthesis and byproduct disposal, thus, inhibition of the protein degradation pathways became an emerging approach for cancer therapy. The present study revealed that withaferin-A (WA), the biologically active withanolide derived from Withania somnifera, initially induced formation of autophagosomes in human breast cancer cell-lines, MCF-7 and MDA-MB-231. WA treatment elevated the levels of autophagic substrate p62/SQSTM1 (p62) and both LC3-II and LC3-I (microtubule-associated protein 2 light chain 3) and simultaneously reduced the upstream autophagy markers like beclin-1 and ATG5-ATG12 complex, which indicate accumulation of autophagosomes in the cells. WA induced disruption of microtubular network through inhibition of tubulin polymerization and its hyper-acetylation, thus prevent the formation of autolysosome (by merging of autophagosomes with lysosomes) and its recycling process, leading to incomplete autophagy. Further, WA caused ER (Endoplasmic Reticulum) stress, which is evident from the activation of ER-related caspase-4 and increased levels of ER stress marker proteins. Thus, these findings altogether indicate that WA mediated inhibition of proteasomal degradation system and perturbation of autophagy, i.e. suppression of both the intracellular degradation systems caused accumulation of ubiquitinated proteins, which in turn led to unfolded protein response and ER stress mediated proteotoxicity in human breast cancer cell-lines, MCF-7 and MDA-MB-231. •WA treatment leads to onset of autophagsome formation.•The autophagy, however, is impaired in the later phases.•Wa induces disruption of microtubular network.•WA evokes ER stress mediated proteotoxicity.