Cytosolic phospholipase A2 is required for optimal ATP activation of BK channels in GH(3) cells.
To test the hypothesis that ATP activation of BK channels in GH(3) cells involves cytosolic phospholipase A(2) (cPLA(2)) as a potential protein target for phosphorylation, we first inhibited the activity of cPLA(2) by both pharmacologic and molecular biologic approaches. Both approaches resulted in a decrease rather than an increase in BK channel activity by ATP, suggesting that in the absence of cPLA(2), phosphorylation of other regulatory elements, possibly the BK channel protein itself, results in inactivation rather than activation of the channel. The absence of changes in activity in the presence of the non-substrate ATP analog 5'-adenylyl-beta,gamma-imidodiphosphate verified that ATP hydrolysis was required for channel activation by ATP. Experiments with an activator and inhibitor of protein kinase C (PKC) support the hypothesis that PKC can be involved in the activation of BK channels by ATP; and in the absence of PKC, other kinases appear to phosphorylate additional elements in the regulatory pathway that reduce channel activity. Our data point to cPLA(2)-alpha (but not cPLA(2)-gamma) as one target protein for phosphorylation that is intimately associated with the BK channel protein.
Denson
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DD
Departments of Anesthesiology and Physiology and the Center for Cellular and Molecular Signaling, Emory University School of Medicine, Atlanta, Georgia 30322, USA. ddenson@emory.org
Wang
X
X
Worrell
R T
RT
AlKhalili
O
O
Eaton
D C
DC
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D000266:Adenylyl Imidodiphosphate / Q000494:pharmacology; D000818:Animals; D034341:Aristolochic Acids; D015153:Blotting, Western; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D003600:Cytosol / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006868:Hydrolysis; D051036:Large-Conductance Calcium-Activated Potassium Channels; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D010616:Phenanthrenes / Q000494:pharmacology; D010741:Phospholipases A / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology*; D054467:Phospholipases A2; D010766:Phosphorylation; D015221:Potassium Channels / Q000378:metabolism*; D024681:Potassium Channels, Calcium-Activated; D020033:Protein Isoforms; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D051381:Rats
D034341:Aristolochic Acids; D004791:Enzyme Inhibitors; D051036:Large-Conductance Calcium-Activated Potassium Channels; D016376:Oligonucleotides, Antisense; D010616:Phenanthrenes; D015221:Potassium Channels; D024681:Potassium Channels, Calcium-Activated; D020033:Protein Isoforms; D000266:Adenylyl Imidodiphosphate; C022571:adenosine 5'-O-(3-thiotriphosphate); D000255:Adenosine Triphosphate; C000228:aristolochic acid I; D011493:Protein Kinase C; D010741:Phospholipases A; D054467:Phospholipases A2