Okadaic acid stimulates ouabain‐sensitive 86Rb+‐uptake and phosphorylation of the Na+/K+‐ATPase α‐subunit in rat hepatocytes

FEBS Letters

Mader, Annie Cielo L.¹  Lynch, Christopher J.¹  Hazen, Stacy A.¹  Ng, Yuk-Chow²  McCall, Kenneth M.¹ 

[1]Department of Cellular and Molecular Physiology and Pharmacology, College of Medicine, The Pennsylvania State University, Hershey, PA 17033, USA

[2]The Milton S. Hershey Medical Center and Pennsylvania State University, College of Medicine, Hershey, PA 17033, USA

关键词: Na+/K+-ATPase;    Protein phosphatase;    Insulin;    Liver;    Okadaic acid;   

DOI  :  10.1016/0014-5793(94)80085-5

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Ca²⁺-mobilizing and cAMP-dependent hormones rapidly increase sodium, potassium-dependent adenosine triphosphatase (Na⁺/K⁺-ATPase)-mediated transport in rat hepatocytes. To explore the possible role of protein phosphatases in these responses we used a protein phosphatase inhibitor, okadaic acid. Okadaic acid stimulation of ouabain-sensitive ⁸⁶Rb⁺-uptake was maximal between two and three minutes and displayed an EC₅₀ of 41 ± 1 nM. Inhibition of Na⁺/H⁺ exchange with an amiloride analog abolished the response to insulin, but had no effect on okadaic acid-mediated stimulation of Na⁺/K⁺-ATPase transport. In hepatocytes metabolically-radiolabeled with ³²P_i, okadaic acid stimulated the incorporation of radioactivity into several 95 kDa peptides, one of which reacted with anti-LEAVE peptide antisera, that recognizes Na⁺/K⁺-ATPase α-subunits. In other experiments Na⁺/K⁺-ATPase was immunoprecipitated from detergent-solubilized membrane fractions of metabolically-radiolabeled cells with an antisera to purified rat kidney Na⁺/K⁺-ATPase. A 95 kDa phosphoprotein was immunoprecipitated using anti-Na⁺/K⁺-ATPase antisera, but not by preimmune serum. Okadaic acid stimulated incorporation of radioactivity into this band by 220 ± 28%. These findings provide support for the hypothesis that rapid stimulation of hepatic Na⁺/K⁺-ATPase by hormones may be related to protein kinase/phosphatase-mediated changes in the phosphorylation state of the Na⁺/K⁺-ATPase α-subunit.

【 授权许可】

Unknown