Angiotensin II-dependent phosphorylation at Ser(11)/Ser(18) and Ser(938)TI Angiotensin II-dependent phosphorylation at Ser(11)/Ser(18) and Ser(938) shifts the E-2 conformations of rat kidney Na+/K+-ATPase

摘要

Kidney plasma membranes, which contain a single alpha-1 isoform of Na+/K+-ATPase, simultaneously contain two sub-conformations of E2P, differing in their rate of digoxin release in response to Na+ and ATP. Treating cells with Ang II (angiotensin II) somehow changes the conformation of both, because it differentially inhibits the rate of digoxin release. In the present study we tested whether Ang II regulates release by increasing phosphorylation at Se-11/Ser(18) and Ser(938). Opossum kidney cells co-expressing the AT(1a) receptor and either alpha-1.wild-type, alpha-1.S11A/S18A or alpha-1.S938A were treated with or without 10 nM Ang H for 5 min, increasing phosphorylation at the three sites. Na+/K+-ATPase was bound to digoxin-affinity columns in the presence of Na+, ATP and Mg2+. A solution containing 30 mM NaCl and 3 mM ATP eluted similar to 20% of bound untreated Na+/K+-ATPase (Population #1). Pre-treating cells with Ang II slowed the elution of Population #1 in alpha-1.wild-type and alpha-1.S938A, but not alpha-1.S11A/S18A cells. Another 50% of bound Na+/K+-ATPase (Population #2) was subsequently eluted in two phases by a solution containing 150 mM NaCl and 3 nM ATP. Ang II increased the initial rate and slowed the second phase in alpha-1.wild-type, but not alpha-1.5938A, cells. Thus Ang II changes the conformation of two forms of EP2 via differential phosphorylation.