Angiotensin II-dependent phosphorylation at Ser11/Ser18 and Ser938 shifts the E2 conformations of rat kidney Na+/K+-ATPase.
Angiotensin II-dependent phosphorylation at Ser11/Ser18 and Ser938 shifts the E2 conformations of rat kidney Na+/K+-ATPase.
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DOI：
10.1042/bj20111398
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发表时间：
2012-04-01
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影响因子：
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通讯作者：
Yingst DR
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作者：
Massey KJ;Li Q;Rossi NF;Mattingly RR;Yingst DR
肾质膜含有单一α-1亚型的Na+/K+ -ATP酶，同时含有E2P的两个亚构象，它们对Na+和ATP的释放速度不同。用Ang II（血管紧张素II）处理细胞会改变两者的构象，因为它会不同程度地抑制地高辛的释放速度。在本研究中，我们测试了Ang II是否通过增加Ser11/Ser18和Ser938的磷酸化来调节释放。负鼠肾细胞共表达AT1a受体和α-1。野生型,α1。S11A/S18A或α-1。用或不加10 nM Ang II处理S938A 5分钟，三个位点的磷酸化增加。Na+/K+ -ATP酶在Na+、ATP和Mg2+存在下与地高辛亲和柱结合。含有30 mM NaCl和3 mM ATP的溶液洗脱了约20%结合的未处理Na+/K+ -ATP酶（种群#1）。用Ang II预处理细胞后，α-1中1号群体的洗脱速度减慢。野生型和α-1。S938A，而不是α-1。S11A / S18A细胞。另外50%结合的Na+/K+ -ATP酶（种群#2）随后用含有150 mM NaCl和3 mM ATP的溶液分两相洗脱。Ang II增加了α-1的初始速率，减缓了α-1的第二阶段。野生型，但不是α-1型。S938A,细胞。因此，Ang II通过不同的磷酸化改变了两种形式的EP2的构象。
Kidney plasma membranes, which contain a single α-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 Ser11/Ser18 and Ser938. Opossum kidney cells co-expressing the AT1a receptor and either α-1.wild-type, α-1.S11A/S18A or α-1.S938A were treated with or without 10 nM Ang II 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 ~ 20 % of bound untreated Na+/K+ -ATPase (Population #1). Pre-treating cells with Ang II slowed the elution of Population #1 in α-1.wild-type and α-1.S938A, but not α-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 mM ATP. Ang II increased the initial rate and slowed the second phase in α-1.wild-type, but not α-1.S938A, cells. Thus Ang II changes the conformation of two forms of EP2 via differential phosphorylation.