Hydrogen peroxide-induced activation of SAPK/JNK regulated by phosphatidylinositol 3-kinase in Chinese hamster V79 cells.
To clarify activation mechanisms of stress-activated protein kinase/C-Jun N-terminal kinase (SAPK/JNK) during oxidative stress, the roles of phosphatidylinositol 3-kinase (PI 3-kinase), concentration of intracellular calcium ([Ca2+]i), and cyclic AMP-dependent kinase (PKA) in hydrogen peroxide (H2O2)-induced SAPK/JNK activation were examined in Chinese hamster V79 cells. SAPK/JNK was dose-dependently activated after H2O2 treatment (from 10 microM to 1 mM), and a PI 3-kinase inhibitor (wortmaninn), intracellular calcium chelator (BAPTA-AM), and PKA activator (dibutyl cyclic AMP and forskolin) inhibited this activation. An increase in [Ca2+], was observed after treatment with H2O2. Immunoprecipitation revealed that a PI 3-kinase regulatory subunit, p85alpha, was associated with insulin receptor substance 1 (IRS-1) phosphorylated by H2O2 treatment. Furthermore, the formation of this complex of p85alpha and phospho-IRS-1 was abolished by the presence of BAPTA-AM but not forskolin. These results indicated that the PI 3-kinase activated through phosphorylation of IRS-1 upstream of SAPK/JNK after H2O2 treatment of V79 cells and that [Ca2+]i was a regulation factor for phosphorylation of IRS-1.
Department of Environmental Veterinary Medicine, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo, Japan.
O
O
Inanami
K
K
Takahashi
A
A
Yoshito
M
M
Kuwabara
D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D003994:Bucladesine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D002614:Chelating Agents / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D006224:Cricetinae; D003412:Cricetulus; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004789:Enzyme Activation; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D015151:Immunoblotting; D055504:Insulin Receptor Substrate Proteins; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000276:immunology / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011233:Precipitin Tests; D000077191:Wortmannin
D000730:Androstadienes; D003994:Bucladesine; D002118:Calcium; D002614:Chelating Agents; D005576:Colforsin; D017868:Cyclic AMP-Dependent Protein Kinases; D004533:Egtazic Acid; D006861:Hydrogen Peroxide; D055504:Insulin Receptor Substrate Proteins; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010750:Phosphoproteins; D000077191:Wortmannin; C526292:IRS1 protein, human; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester