title: Regulation of ERK1/2 by ouabain and Na-K-ATPase-dependent energy utilization and AMPK activation in parotid acinar cells.
abstract: We previously found that the phosphorylation of ERK1/2 by submaximal concentrations of the muscarinic receptor ligand carbachol was potentiated in rat parotid acinar cells exposed to ouabain, a cardiac glycoside that inhibits the Na-K-ATPase. We now report that this signaling phenomenon involves the prevention of negative regulation of extracellular signal-regulated kinase-1/2 (ERK1/2) that is normally mediated by AMP-activated protein kinase (AMPK). Carbachol increases the turnover of the ATP-consuming Na-K-ATPase, reducing intracellular ATP and promoting the phosphorylation/activation of the energy sensor AMPK. Ouabain blocks the reduction in ATP and subsequent AMPK phosphorylation, which is regulated by the AMP-to-ATP ratio. The ouabain-promoted enhancement of ERK1/2 phosphorylation was not reproduced in Par-C10 cells, an immortalized rat parotid cell line that did not respond to carbachol with an ATP reduction and that employs an upstream AMPK kinase (Ca(2+)/calmodulin-dependent protein kinase kinase, CaMKK) different from that (LKB1) in native cells. In native parotid cells, inhibitory effects of AMPK on ERK1/2 signaling were examined by activating AMPK with 5-aminoimidazole-4-carboxamide-1-beta-d-ribofuranoside (AICAR), which is converted to an AMP mimetic but does not alter parotid ATP levels. AICAR-treated cells display increases in AMPK phosphorylation and a reduced phosphorylation of ERK1/2 subsequent to activation of muscarinic and P2X(7) receptors, which promote increases in Na-K-ATPase turnover, but not upon epidermal growth factor receptor activation. These results suggest that carbachol-initiated AMPK activation can produce a negative feedback on ERK1/2 signaling in response to submaximal muscarinic receptor activation and that increases in fluid secretion can modulate receptor-initiated signaling events indirectly by producing ion transport-dependent decreases in ATP.
authors.affiliation: Division of Signal Transduction, Department of Medicine, Beth Israel Deaconess Medical Center, Boston, Massachusetts 02111, USA. ssoltoff@bidmc.harvard.edu
authors.forename: Stephen P
authors.initials: SP
authors.lastname: Soltoff
authors.forename: Lee
authors.initials: L
authors.lastname: Hedden
mesh_terms: D055372:AMP-Activated Protein Kinases; D000255:Adenosine Triphosphate / Q000378:metabolism; D000620:Aminoimidazole Carboxamide / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D002217:Carbachol / Q000494:pharmacology; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004734:Energy Metabolism / Q000187:drug effects*; D004789:Enzyme Activation; D020536:Enzyme Activators / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D025461:Feedback, Physiological; D066298:In Vitro Techniques; D008297:Male; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D009097:Multienzyme Complexes / Q000378:metabolism*; D018721:Muscarinic Agonists / Q000494:pharmacology; D010042:Ouabain / Q000494:pharmacology*; D010306:Parotid Gland / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012265:Ribonucleotides / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors* / Q000378:metabolism
chemical_list: D020536:Enzyme Activators; D004791:Enzyme Inhibitors; D009097:Multienzyme Complexes; D018721:Muscarinic Agonists; D012265:Ribonucleotides; D000620:Aminoimidazole Carboxamide; D010042:Ouabain; D000255:Adenosine Triphosphate; D002217:Carbachol; D017346:Protein Serine-Threonine Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D055372:AMP-Activated Protein Kinases; D000254:Sodium-Potassium-Exchanging ATPase; C031143:AICA ribonucleotide