Role of reactive oxygen species and Cr(VI) in Ras-mediated signal transduction

Suwei Wang, Stephen S. Leonard, Jianping Ye, Ning Gao, Liying Wang, Xianglin Shi

Research output: Contribution to journalReview articlepeer-review

30 Scopus citations

Abstract

Previous studies have shown that a constitutively active isoform of Ras is able to produce superoxide radical (O2-). The present study investigate the mechanisms by which O2- radical mediates signals from Ras protein to the nucleus, leading to cellular responses such as apoptosis in Cr(VI) -stimulated cells. Two human prostate tumor cell lines, Ras(+), which overexpresses Ras, and Ras(-), which has a normal Ras level, were utilized. Compared to Ras(-) cells, Ras(+) cells exhibited higher susceptibility to apoptosis induced by Cr(VI). Catalase, sodium formate, and deferoxamine inhibited Cr(VI)-induced apoptosis. Similar differences were observed in both cellular DNA damage and the activation of p53 protein. The differences in Cr(VI)-induced cell responses in Ras(+) and Ras(-) cells were due to differences in the generation of free radicals between these two cells. ESR spin trapping measurements showed that Ras(+) cells generated more hydroxyl radical (•OH), O2- radical, and Cr(V) than Ras(-) cells following Cr(VI) stimulation. The generation of the reactive oxygen species (ROS) can be abolished by the addition of superoxide dismutase (SOD) or if the experiment were carried out in an argon atmosphere. Catalase inhibited spin adduct signals but was much less potent than SOD. The mechanism of ROS generation in Cr(VI)-stimulated Ras(+) cells involves the reduction of molecular oxygen to O2- radical by a flavoenzyme-containing NADPH oxidase complex as shown by oxygen consumption and diphenylene iodonium (DPI) inhibition. Results shown above support the following conclusions: (a) Ras protein mediates O2- radical generation through reduction of molecular oxygen by NADPH oxidase in Cr(VI)-stimulated cells. (b) The O2- radical and Cr(VI) produce other reactive species, including H2O2, •OH radical, and Cr(V) through O2- dismutation and Haber-Weiss type of reactions. (c) Among these reactive species, •OH radical is responsible for the further transduction of signals from Ras to the nucleus, leading to various cell responses.

Original languageEnglish
Pages (from-to)119-127
Number of pages9
JournalMolecular and Cellular Biochemistry
Volume255
Issue number1-2
DOIs
StatePublished - Jan 2004

Bibliographical note

Funding Information:
Molecular oxygen is the original source of ROS generation in Ras(+) cells under Cr(VI) stimulation. Reduction of molecular oxygen and generation of O2·– was demonstrated by elimination of ESR signals under an argon atmosphere. The major reductase involved is the flavoprotein-containing NADPH oxidase complex rather than the mitochondrial electron transfer chain. This conclusion was supported by the inhibition of ROS generation by DPI, a highly selective flavoprotein inhibitor. Rotenone, an inhibitor of mitochondrial electron transport chain, only exhibited slight inhibition.

Keywords

  • Apoptosis
  • Cr(VI)
  • DNA damage
  • NADPH oxidase
  • Ras oncoprotein
  • Reactive oxygen species
  • p53 activation

ASJC Scopus subject areas

  • Molecular Biology
  • Clinical Biochemistry
  • Cell Biology

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