Parathyroid hormone-mediated regulation of Na+-K+-ATPase requires ERK-dependent translocation of protein kinase Calpha.
Parathyroid hormone (PTH) inhibits Na+-K+-ATPase activity by serine phosphorylation of the alpha1 subunit through protein kinase C (PKC)- and extracellular signal-regulated kinase (ERK)-dependent pathways. Based on previous studies we postulated that PTH regulates sodium pump activity through isoform-specific PKC-dependent activation of ERK. In the present work utilizing opossum kidney cells, a model of renal proximal tubule, PTH stimulated membrane translocation of PKCalpha by 102 +/- 16% and PKCbetaI by 41 +/- 7% but had no effect on PKCbetaII and PKCzeta. Both PKCalpha and PKCbetaI phosphorylated the Na+-K+-ATPase alpha1 subunit in vitro. PTH increased the activity of PKCalpha but not PKCbetaI. Coimmunoprecipitation assays demonstrated that treatment with PTH enhanced the association between Na+-K+-ATPase alpha1 subunit and PKCalpha, whereas the association between Na+-K+-ATPase alpha1 subunit and PKCbetaI remained unchanged. A PKCalpha inhibitory peptide blocked PTH-stimulated serine phosphorylation of the Na+-K+-ATPase alpha1 subunit and inhibition of Na+-K+-ATPase activity. Pharmacologic inhibition of MEK-1 blocked PTH-stimulated translocation of PKCalpha, whereas transfection of constitutively active MEK-1 cDNA induced translocation of PKCalpha and increased phosphorylation of the Na+-K+-ATPase alpha1 subunit. In contrast, PTH-stimulated ERK activation was not inhibited by pretreatment with the PKCalpha inhibitory peptide. Inhibition of PKCalpha expression by siRNA did not inhibit PTH-mediated ERK activation but significantly reduced PTH-mediated phosphorylation of the Na+-K+-ATPase alpha1 subunit. Pharmacologic inhibition of phosphoinositide 3-kinase blocked PTH-stimulated ERK activation, translocation of PKCalpha, and phosphorylation of the Na+-K+-ATPase alpha1 subunit. We conclude that PTH stimulates Na+-K+-ATPase phosphorylation and decreases the activity of Na+-K+-ATPase by ERK-dependent activation of PKCalpha.
Khundmiri
Syed J
SJ
Department of Medicine, University of Louisville and Veterans Affairs Medical Center, Louisville, Kentucky 40202, USA. syed.khundmiri@louisvile.edu
Dean
William L
WL
McLeish
Kenneth R
KR
Lederer
Eleanor D
ED
D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D020935:MAP Kinase Signaling System; D008969:Molecular Sequence Data; D009893:Opossums; D010281:Parathyroid Hormone / Q000494:pharmacology*; D010446:Peptide Fragments / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D021381:Protein Transport; D011994:Recombinant Proteins / Q000378:metabolism; D012413:Rubidium / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D014162:Transfection
D010281:Parathyroid Hormone; D010446:Peptide Fragments; D011994:Recombinant Proteins; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D048049:Extracellular Signal-Regulated MAP Kinases; D000254:Sodium-Potassium-Exchanging ATPase; D012413:Rubidium