Stimulation of protein kinase C pathway mediates endocytosis of human nongastric H+-K+-ATPase, ATP1AL1.
The human nongastric H+-K+-ATPase, ATP1AL1, shown to reabsorb K+ in exchange for H+ or Na+, is localized in the luminal plasma membrane of renal epithelial cells. It is presumed that renal H+-K+-ATPases can be regulated by endocytosis. However, little is known about the molecular mechanisms that control plasma membrane expression of renal H+-K+-ATPases. In our study, activation of protein kinase C (PKC) using phorbol esters (phorbol 12-myristate 13-acetate) leads to clathrin-dependent internalization and intracellular accumulation of the ion pump in stably transfected Madin-Darby canine kidney cells. Functional inactivation of the H+-K+-ATPase by PKC activation is shown by intracellular pH measurements. Proton extrusion capacity of ATP1AL1-transfected cells is drastically reduced after phorbol 12-myristate 13-acetate incubation and can be prevented with the PKC blocker bisindolylmaleimide. Ion pump internalization and inactivation are specifically mediated by the PKC pathway, whereas activation of the protein kinase A pathway has no influence. Our results show that the nongastric H+-K+-ATPase is a specific target for the PKC pathway. Therefore, PKC-mediated phosphorylation is a potential regulatory mechanism for apical nongastric H+-K+-ATPase plasma membrane expression.
Reinhardt
J
J
Institute of Physiology, University of Münster, D-48149 Münster, Germany. Juergen.Reinhardt@pharma.novartis.com
Kosch
M
M
Lerner
M
M
Bertram
H
H
Lemke
D
D
Oberleithner
H
H
D000818:Animals; D019921:Biotinylation; D002273:Carcinogens / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D004705:Endocytosis / Q000187:drug effects / Q000502:physiology*; D005455:Fluorescent Antibody Technique; D017506:H(+)-K(+)-Exchanging ATPase / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007668:Kidney / Q000166:cytology*; D018613:Microscopy, Confocal; D011493:Protein Kinase C / Q000378:metabolism*; D000644:Quaternary Ammonium Compounds / Q000494:pharmacology; D000254:Sodium-Potassium-Exchanging ATPase / Q000235:genetics / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014162:Transfection
D002273:Carcinogens; D000644:Quaternary Ammonium Compounds; D011493:Protein Kinase C; C088641:ATP12A protein, human; D017506:H(+)-K(+)-Exchanging ATPase; D000254:Sodium-Potassium-Exchanging ATPase; D013755:Tetradecanoylphorbol Acetate