Simultaneous phosphorylation of Ser11 and Ser18 in the alpha-subunit promotes the recruitment of Na+,K+-ATPase molecules to the plasma membrane

Citation
R. Efendiev et al., Simultaneous phosphorylation of Ser11 and Ser18 in the alpha-subunit promotes the recruitment of Na+,K+-ATPase molecules to the plasma membrane, BIOCHEM, 39(32), 2000, pp. 9884-9892
Citations number
44
Categorie Soggetti
Biochemistry & Biophysics
Journal title
BIOCHEMISTRY
ISSN journal
00062960 → ACNP
Volume
39
Issue
32
Year of publication
2000
Pages
9884 - 9892
Database
ISI
SICI code
0006-2960(20000815)39:32<9884:SPOSAS>2.0.ZU;2-J
Abstract
Renal sodium homeostasis is a major determinant of blood pressure and is re gulated by several natriuretic and antinatriuretic hormones. These hormones , acting through intracellular second messengers, either activate or inhibi t proximal tubule Na+,K+-ATPase. We have shown previously that phorbol este r (PMA) stimulation of endogenous PKC leads to activation of Na+,K+-ATPase in cultured proximal tubule cells (OK cells) expressing the rodent Na+,K+-A TPase or-subunit. We have now demonstrated that the treatment with PMA lead s to an increased amount of Na+,K+-ATPase molecules in the plasmalemma, whi ch is proportional to the increased enzyme activity. Colchicine, dinitrophe nol, and potassium cyanide prevented the PMA-dependent stimulation of activ ity without affecting the increased level of phosphorylation of the Na+,K+- ATPase alpha-subunit. This suggests that phosphorylation does not directly stimulate Na+,K+-ATPase activity; instead, phosphorylation may be the trigg ering mechanism for recruitment of Na+,K+-ATPase molecules to the plasma me mbrane. Transfected cells expressing either an S11A or S18A mutant had the same basal Na+,K+-ATPase activity as cells expressing the wild-type rodent alpha-subunit, but PMA stimulation of Na+,K+-ATPase activity was completely abolished in either mutant. PMA treatment led to phosphorylation of the al pha-subunit by stimulation of PKC-beta, and the extent of this phosphorylat ion was greatly reduced in the S11A and S18A mutants. These results indicat e that both Ser11 and Ser18 of the alpha-subunit are essential for PMA stim ulation of Na+,K+-ATPase activity, and that these amino acids are phosphory lated during this process. The results presented here support the hypothesi s that PMA regulation of Na+,K+-ATPase is the result of an increased number of Na+,K+-ATPase molecules in the plasma membrane.