Low salt concentrations activate AMP-activated protein kinase in mouse macula densa cells.
The energy-sensing kinase AMP-activated protein kinase (AMPK) is associated with the sodium-potassium-chloride cotransporter NKCC2 in the kidney and phosphorylates it on a regulatory site in vitro. To identify a potential role for AMPK in salt sensing at the macula densa, we have used the murine macula densa cell line MMDD1. In this cell line, AMPK was rapidly activated by isosmolar low-salt conditions. In contrast to the known salt-sensing pathway in the macula densa, AMPK activation occurred in the presence of either low sodium or low chloride and was unaffected by inhibition of NKCC2 with bumetanide. Assays using recombinant AMPK demonstrated activation of an upstream kinase by isosmolar low salt. The specific calcium/calmodulin-dependent kinase kinase inhibitor STO-609 failed to suppress AMPK activation, suggesting that it was not part of the signal pathway. AMPK activation was associated with increased phosphorylation of the specific substrate acetyl-CoA carboxylase (ACC) at Ser(79), as well as increased NKCC2 phosphorylation at Ser(126). AMPK activation due to low salt concentrations was inhibited by an adenovirus construct encoding a kinase dead mutant of AMPK, leading to reduced ACC Ser(79) and NKCC2 Ser(126) phosphorylation. This work demonstrates that AMPK activation in macula densa-like cells occurs via isosmolar changes in sodium or chloride concentration, leading to phosphorylation of ACC and NKCC2. Phosphorylation of these substrates in vivo is predicted to increase intracellular chloride and so reduce the effect of salt restriction on tubuloglomerular feedback and renin secretion.
Cook
Natasha
N
Dept. of Nephrology, Austin Health, Studley Rd., Heidelberg 3084, Victoria, Australia. natasha.cook@austin.org.au
Fraser
Scott A
SA
Katerelos
Marina
M
Katsis
Frosa
F
Gleich
Kurt
K
Mount
Peter F
PF
Steinberg
Gregory R
GR
Levidiotis
Vicki
V
Kemp
Bruce E
BE
Power
David A
DA
D055372:AMP-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism*; D000103:Acetyl-CoA Carboxylase / Q000378:metabolism; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001562:Benzimidazoles / Q000494:pharmacology; D002034:Bumetanide / Q000494:pharmacology; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D002460:Cell Line; D004789:Enzyme Activation; D007684:Kidney Tubules / Q000187:drug effects / Q000201:enzymology* / Q000473:pathology; D051379:Mice; D009154:Mutation; D053644:Naphthalimides / Q000494:pharmacology; D009336:Necrosis; D009994:Osmolar Concentration; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011994:Recombinant Proteins / Q000378:metabolism; D012694:Serine; D012965:Sodium Chloride / Q000378:metabolism*; D049994:Sodium Potassium Chloride Symporter Inhibitors / Q000494:pharmacology; D028021:Sodium-Potassium-Chloride Symporters / Q000378:metabolism; D064466:Solute Carrier Family 12, Member 1; D013997:Time Factors
D001562:Benzimidazoles; D053644:Naphthalimides; D047428:Protein Kinase Inhibitors; D011994:Recombinant Proteins; C458525:STO 609; C578715:Slc12a1 protein, mouse; D049994:Sodium Potassium Chloride Symporter Inhibitors; D028021:Sodium-Potassium-Chloride Symporters; D064466:Solute Carrier Family 12, Member 1; D002034:Bumetanide; D012965:Sodium Chloride; D012694:Serine; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D055372:AMP-Activated Protein Kinases; D000103:Acetyl-CoA Carboxylase