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PMID: 15537704 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

The major target of the endogenously generated reactive oxygen species in response to insulin stimulation is phosphatase and tensin homolog and not phosphoinositide-3 kinase (PI-3 kinase) in the PI-3 kinase/Akt pathway.

Molecular biology of the cell ·Vol. 16 ·No. 1 ·2005-01-00 ·Pages 348-57

Seo JH, Ahn Y, Lee SR, Yeol Yeo C, Chung Hur K

Abstract

Phosphoinositide-3 kinase (PI-3 kinase) and its downstream signaling molecules PDK-1 and Akt were analyzed in SK-N-SH and SK-N-BE(2) human neuroblastoma cell lines. When cells were stimulated with insulin, PI-3 kinase was activated in both cell lines, whereas the translocation of PDK-1 to the membrane fraction and phosphorylated Akt were observed only in SK-N-SH cells. Analyses of the insulin-mediated reactive oxygen species (ROS) generation and Phosphatase and Tensin homolog (PTEN) oxidation indicate that PTEN oxidation occurred in SK-N-SH cells, which can produce ROS, but not in SK-N-BE(2) cells, which cannot increase ROS in response to insulin stimulation. When SK-N-SH cells were pretreated with the NADPH oxidase inhibitor diphenyleneiodonium chloride before insulin stimulation, insulin-mediated translocation of PDK-1 to the membrane fraction and phosphorylation of Akt were remarkably reduced, whereas PI-3 kinase activity was not changed significantly. These results indicate that not only PI-3 kinase activation but also inhibition of PTEN by ROS is needed to increase cellular level of phosphatidylinositol 3,4,5-trisphosphate for recruiting downstream signaling molecules such as PDK-1 and Akt in insulin-mediated signaling. Moreover, the ROS generated by insulin stimulation mainly contributes to the inactivation of PTEN and not to the activation of PI-3 kinase in the PI-3 kinase/Akt pathway.

MeSH Terms
3-Phosphoinositide-Dependent Protein Kinases Cell Line, Tumor Cell Membrane/metabolism Electrophoresis, Polyacrylamide Gel Enzyme Inhibitors/pharmacology Humans Hydrogen Peroxide/pharmacology Immunoblotting Immunoprecipitation Insulin/metabolism NADPH Oxidases/metabolism Onium Compounds/pharmacology Oxygen/metabolism PTEN Phosphohydrolase Phosphatidylinositol 3-Kinases/metabolism Phosphoric Monoester Hydrolases/metabolism Phosphorylation Protein Serine-Threonine Kinases/metabolism Protein Transport Proto-Oncogene Proteins/metabolism Proto-Oncogene Proteins c-akt Reactive Oxygen Species Time Factors Tumor Suppressor Proteins/metabolism
Chemicals
Enzyme Inhibitors Insulin Onium Compounds Proto-Oncogene Proteins Reactive Oxygen Species Tumor Suppressor Proteins diphenyleneiodonium Hydrogen Peroxide NADPH Oxidases 3-Phosphoinositide-Dependent Protein Kinases AKT1 protein, human Protein Serine-Threonine Kinases Proto-Oncogene Proteins c-akt Phosphoric Monoester Hydrolases PTEN Phosphohydrolase PTEN protein, human Oxygen
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Seo Ji Hae
Department of Biology, Ewha Women's University, Seoul 120-750, Korea.
Ahn Younghee
Lee Seung-Rock
Yeol Yeo Chang
Chung Hur Kyu
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Article Info
Journal
Molecular biology of the cell
Abbr.
Mol Biol Cell
ISSN
1059-1524
DOI
10.1091/mbc.e04-05-0369
Published
2005-01-00
Epub
2004-00-10
Pages
348-57
Language
English
Region
United States
NLM ID
9201390
PMCID
PMC539178
Subset
IM
External Links
PubMed source DOI: 10.1091/mbc.e04-05-0369 PMC Full Text View journal details
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