Regulation of oxidative stress-induced calcium release by phosphatidylinositol 3-kinase and Bruton's tyrosine kinase in B cells.
Hydrogen peroxide stimulates a tyrosine kinase-dependent calcium release from intracellular stores, which is assumed to be achieved through the activation of phospholipase Cgamma2 (PLCgamma2) via a tyrosine phosphorylation mechanism in B cells. Here we show that H(2)O(2) induces both tyrosine phosphorylation on PLCgamma2 and the activation of phosphatidylinositol 3-kinase (PI3K) in B cells, and that the phosphatidylinositol 3-kinase inhibitor, Wortmannin, partially inhibited the H(2)O(2)-induced calcium release without affecting tyrosine phosphorylation on PLCgamma2. Overexpression of human Bruton's tyrosine kinase (Btk), which was activated by H(2)O(2), almost completely overcame the inhibition of calcium release by Wortmannin. The reversal of Wortmannin's inhibition by enhancing Btk concentration seemed unique to the H(2)O(2)-mediated effect, because Btk failed to overcome the inhibition of Wortmannin on B cell receptor-triggered calcium mobilization. Immunoblot analysis revealed that Btk formed stable complexes with several tyrosine-phosphorylated proteins, including PLCgamma2, only in Btk-overexpressed cells on H(2)O(2) stimulation. Together, our data are consistent with the notion that PIP3 and/or a high concentration of Btk target the activated PLCgamma2 to its substrate site for maximal catalytic efficiency.
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Laboratory of Biochemistry, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892-0342, USA.
Stadtman
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Chock
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PB
D000077329:Agammaglobulinaemia Tyrosine Kinase; D001402:B-Lymphocytes / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D018384:Oxidative Stress; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects
D006861:Hydrogen Peroxide; D019869:Phosphatidylinositol 3-Kinases; D011505:Protein-Tyrosine Kinases; D000077329:Agammaglobulinaemia Tyrosine Kinase; C000625949:BTK protein, human; D002118:Calcium