Angiotensin II increases the amount of Na,K‐ATPase in the plasma membrane of proximal tubule cells by phosphorylation of the α‐1 subunit at Ser‐943.

Abstract

Our results and those of others have demonstrated that direct stimulation of Na,K-ATPase activity in rat proximal tubules (PCT) by angiotensin II (Ang II) is associated with an altered pattern of phosphorylation of the α-1 subunit of the Na,K-ATPase. The stimulatory effect of Ang II is mediated by the AT1 receptor and appears to involve recruitment of more Na,K-ATPase to the plasma membrane (PM). Also, in rat PCT this stimulation by Ang II can be blocked by pertussis toxin, implicating Gαi. To examine if the increase in the amount of α-1 subunit in the PM in response to Ang II is mediated by changes in phosphorylation of Ser-943, the only known PKA site on the α-1 subunit, we used site directed mutagenesis to develop lines of opossum kidney (OK) cells that stably express the AT1a receptor and either the wild-type rat α-1 subunit or an α-1 subunit with Ser-943 mutated to alanine. Using Western blot analysis of biotinylated PM proteins isolated on immobilized streptavidin, Ang II significantly increased the amount of α-1 subunit in the PM of OK cells expressing the wild-type α-1 subunit by ~45% but effected no change in the amount of S943A mutant α-1 subunits in the PM. We conclude that the influence of Ang II on Na,K-ATPase activity includes increasing the number of copies of the α-1 subunit in the PM and this effect is mediated, in part, by phosphorylation of the α-1 subunit at Ser-943. Supported by NIH DK60752.