Involvement of Na/K-ATPase in hydrogen peroxide-induced activation of the Src/ERK pathway in LLC-PK1 cells.
We have shown that Na/K-ATPase interacts with Src. Here, we test the role of this interaction in H2O2-induced activation of Src and ERK. We found that exposure of LLC-PK1 cells to H2O2 generated by the addition of glucose oxidase into the culture medium activated Src and ERK1/2. It also caused a modest reduction in the number of surface Na/K-ATPases and in ouabain-sensitive Rb(+) uptake. These effects of H2O2 seem similar to those induced by ouabain, a specific ligand of Na/K-ATPase, in LLC-PK1 cells. In accordance, we found that the effects of H2O2 on Src and ERK1/2 were inhibited in α1 Na/K-ATPase-knockdown PY-17 cells. Whereas expression of wild-type α1 or the A420P mutant α1 defective in Src regulation rescued the pumping activity in PY-17 cells, only α1, and not the A420P mutant, was able to restore the H2O2-induced activation of protein kinases. Consistent with this, disrupting the formation of the Na/K-ATPase/Src complex with pNaKtide attenuated the effects of H2O2 on the kinases. Moreover, a direct effect of H2O2 on Na/K-ATPase-mediated regulation of Src was demonstrated. Finally, H2O2 reduced the expression of E-cadherin through the Na/K-ATPase/Src-mediated signaling pathway. Taken together, the data suggest that the Na/K-ATPase/Src complex may serve as one of the receptor mechanisms for H2O2 to regulate Src/ERK protein kinases and consequently the phenotype of renal epithelial cells.
Renal Division, Department of Medicine, Peking University First Hospital, Institute of Nephrology, Peking University, Beijing, China.
Yu
Y
Wang
Department of Physiology and Department of Pharmacology and Medicine, College of Medicine, University of Toledo, Toledo, OH 43614, USA.
Qiqi
Q
Ye
Renal Division, Department of Medicine, Peking University First Hospital, Institute of Nephrology, Peking University, Beijing, China.
Changxuan
C
Liu
Department of Physiology and Department of Pharmacology and Medicine, College of Medicine, University of Toledo, Toledo, OH 43614, USA.
Jeffrey X
JX
Xie
Department of Physiology and Department of Pharmacology and Medicine, College of Medicine, University of Toledo, Toledo, OH 43614, USA.
Yanling
Y
Yan
Department of Physiology and Department of Pharmacology and Medicine, College of Medicine, University of Toledo, Toledo, OH 43614, USA.
Fangfang
F
Lai
Department of Physiology and Department of Pharmacology and Medicine, College of Medicine, University of Toledo, Toledo, OH 43614, USA.
Qiming
Q
Duan
Renal Division, Department of Medicine, Peking University First Hospital, Institute of Nephrology, Peking University, Beijing, China.
Xiaomei
X
Li
Department of Physiology and Department of Pharmacology and Medicine, College of Medicine, University of Toledo, Toledo, OH 43614, USA.
Jiang
J
Tian
Department of Physiology and Department of Pharmacology and Medicine, College of Medicine, University of Toledo, Toledo, OH 43614, USA. Electronic address: Zi-jian.Xie@utoledo.edu.
Zijian
Z
Xie
D000818:Animals; D015820:Cadherins / Q000235:genetics / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation; D005947:Glucose / Q000378:metabolism / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D018374:LLC-PK1 Cells; D019950:Mitogen-Activated Protein Kinase 1 / Q000235:genetics* / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000235:genetics* / Q000378:metabolism; D010042:Ouabain / Q000494:pharmacology; D010641:Phenotype; D021122:Protein Subunits / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction; D000254:Sodium-Potassium-Exchanging ATPase / Q000235:genetics* / Q000378:metabolism; D013552:Swine; D019061:src-Family Kinases / Q000235:genetics* / Q000378:metabolism
D015820:Cadherins; D004791:Enzyme Inhibitors; D021122:Protein Subunits; D010042:Ouabain; D006861:Hydrogen Peroxide; D019061:src-Family Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D000254:Sodium-Potassium-Exchanging ATPase; D005947:Glucose
ERK; Free radicals; Hydrogen peroxide; Mutant; Na/K-ATPase; Src