Phosphorylation regulates an inwardly rectifying ATP-sensitive K(+)- conductance in proximal tubule cells of frog kidney.
K(+) channels in the renal proximal tubule play an important role in salt reabsorption. Cells of the frog proximal tubule demonstrate an inwardly rectifying, ATP-sensitive K(+) conductance that is inhibited by Ba(2+), G(Ba). In this paper we have investigated the importance of phosphorylation state on the activity of G(Ba) in whole-cell patches. In the absence of ATP, G(Ba) decreased over time; this fall in G(Ba) involved phosphorylation, as rundown was inhibited by alkaline phosphatase and was accelerated by the phosphatase inhibitor F(-)(10 mM: ). Activation of PKC using the phorbol ester PMA accelerated rundown via a mechanism that was dependent on phosphorylation. In contrast, the inactive phorbol ester PDC slowed rundown. Inclusion of the PKC inhibitor PKC-ps in the pipette inhibited rundown. These data indicate that PKC-mediated phosphorylation promotes channel rundown. Rundown was prevented by the inclusion of PIP-2 in the pipette. PIP-2 also abrogated the PMA-mediated increase in rundown, suggesting that regulation of G(Ba) by PIP-2 occurred downstream of PKC-mediated phosphorylation. G-protein activation inhibited G(Ba), with initial currents markedly reduced in the presence of GTPgammas. These properties are consistent with G(Ba) being a member of the ATP-sensitive K(+) channel family.
Department of Biomedical Science, University of Sheffield, Alfred Denny Building, Sheffield, UK S10 2TN. l.robson@sheffield.ac.uk
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D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D004553:Electric Conductivity; D019204:GTP-Binding Proteins / Q000378:metabolism*; D015640:Ion Channel Gating / Q000502:physiology*; D007668:Kidney / Q000502:physiology; D007687:Kidney Tubules, Proximal / Q000378:metabolism*; D010766:Phosphorylation; D011188:Potassium / Q000378:metabolism*; D024661:Potassium Channels, Inwardly Rectifying / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D011896:Rana temporaria / Q000502:physiology
D024661:Potassium Channels, Inwardly Rectifying; D000255:Adenosine Triphosphate; D011493:Protein Kinase C; D019204:GTP-Binding Proteins; D011188:Potassium