ROS-dependent Phosphorylation of Bax by Wortmannin Sensitizes Melanoma Cells for TRAIL-induced Apoptosis

Cell Death Dis. 2013 Oct 10;4(10):e839. doi: 10.1038/cddis.2013.344.

Abstract

The pathways of reactive oxygen species (ROS)-mediated apoptosis induction, of Bax activation and the sensitization of tumor cells for TRAIL (TNF-related apoptosis-inducing ligand)-induced apoptosis are still largely elusive. Here, sensitization of melanoma cells for TRAIL by the PI3-kinase inhibitor wortmannin correlated to the activation of mitochondrial apoptosis pathways. Apoptosis was dependent on Bax and abrogated by Bcl-2 overexpression. The synergistic enhancement was explained by Bax activation through wortmannin, which tightly correlated to the characteristic Bax phosphorylation patterns. Thus, wortmannin resulted in early reduction of the Bax-inactivating phosphorylation at serine-184, whereas the Bax-activating phosphorylation at threonine-167 was enhanced. Proving the responsibility of the pathway, comparable effects were obtained with an Akt inhibitor (MK-2206); while suppressed phosphorylation of serine-184 may be attributed to reduced Akt activity itself, the causes of enhanced threonine-167 phosphorylation were addressed here. Characteristically, production of ROS was seen early in response to wortmannin and MK-2206. Providing the link between ROS and Bax, we show that abrogated ROS production by α-tocopherol or by NADPH oxidase 4 (NOX4) siRNA suppressed apoptosis and Bax activation. This correlated with reduced Bax phosphorylation at threonine-167. The data unraveled a mechanism by which NOX4-dependent ROS production controls apoptosis via Bax phosphorylation. The pathway may be considered for proapoptotic, anticancer strategies.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Androstadienes / pharmacology*
  • Apoptosis / drug effects*
  • Caspases / metabolism
  • Cell Line, Tumor
  • Enzyme Activation / drug effects
  • Heterocyclic Compounds, 3-Ring
  • Humans
  • Melanoma / enzymology
  • Melanoma / metabolism*
  • Melanoma / pathology*
  • Mitochondria / drug effects
  • Mitochondria / metabolism
  • NADPH Oxidase 4
  • NADPH Oxidases / metabolism
  • Phosphorylation / drug effects
  • Protein Kinase Inhibitors / pharmacology
  • Proto-Oncogene Proteins c-akt / antagonists & inhibitors
  • Proto-Oncogene Proteins c-akt / metabolism
  • Reactive Oxygen Species / metabolism*
  • TNF-Related Apoptosis-Inducing Ligand / pharmacology*
  • Wortmannin
  • bcl-2-Associated X Protein / metabolism*

Substances

  • Androstadienes
  • Heterocyclic Compounds, 3-Ring
  • MK 2206
  • Protein Kinase Inhibitors
  • Reactive Oxygen Species
  • TNF-Related Apoptosis-Inducing Ligand
  • bcl-2-Associated X Protein
  • NADPH Oxidase 4
  • NADPH Oxidases
  • NOX4 protein, human
  • Proto-Oncogene Proteins c-akt
  • Caspases
  • Wortmannin