ARSENIC TRIOXIDE INDUCES A BECLIN-1 INDEPENDENT AUTOPHAGIC PATHWAY VIA MODULATION OF SNON/SKIL EXPRESSION IN OVARIAN CARCINOMA CELLS

Abstract

Arsenic trioxide (As ₂O ₃), used to treat promyelocytic leukemia, triggers cell death via unknown mechanisms. To further our understanding of As ₂O ₃-induced death, we investigated its effects on transforming growth factor-β (TGFβ) signaling mediators in ovarian cells. Dysregulated TGFβ signaling is a characteristic of ovarian cancers. As ₂O ₃ reduced the protein expression of EVI1, TAK1, SMAD2/3, and TGFβRII while increasing SnoN/SkiL. EVI1 protein was modulated by treatment with the proteosome inhibitors, MG132 and PS-341/Velcade, suggesting that degradation occurs via the ubiquitin-proteosome pathway. The sensitivity of ovarian cells to As ₂O ₃–induced apoptosis correlated with expression of multidrug resistance protein 1. Interestingly, expression of SnoN was similar to LC3-II (autophagy marker) which increased with induction of cytoplasmic vacuolation preceding apoptosis. These vesicles were identified as autophagosomes based on transmission electron microscopy and immunofluorescence staining with EGFP-LC3. The addition of N-acetyl-L-cysteine (ROS scavenger) to As ₂O ₃-treated cells reversed changes in SnoN protein and the autophagic/apoptotic response. In contrast to Beclin-1 knockdown, siRNA targeting ATG5, ATG7, and hVps34 markedly reduced autophagy in As ₂O ₃-treated ovarian carcinoma cells. Further, treatment with SnoN siRNA markedly decreased LC3-II levels and increased PARP degradation (an apoptosis marker). Collectively, these findings suggest that As ₂O ₃ induces a Beclin-1 independent autophagic pathway in ovarian carcinoma cells and implicates SnoN in promoting As ₂O ₃-mediated autophagic cell survival.