Reversible Oxidative Modification A Key Mechanism of Na+-K+ Pump Regulation
Reversible Oxidative Modification A Key Mechanism of Na+-K+ Pump Regulation
复制标题
DOI：
10.1161/circresaha.109.199547
复制
发表时间：
2009-07-17
影响因子：
20.1
通讯作者：
Rasmussen, Helge H.
中科院分区：
文献类型：
--
作者：
Figtree, Gemma A.;Liu, Chia-Chi;Rasmussen, Helge H.
血管紧张素 II (Ang II) 通过蛋白激酶 (PK)C 依赖性 NADPH 氧化酶激活来抑制心脏肌膜 Na+-K+ 泵。我们检查了这是否是由泵亚基的氧化修饰介导的。我们在基线时检测到兔心室肌细胞中β(1)亚基的谷胱甘肽化，但未检测到α(1)亚基。过氧亚硝酸盐 (ONOO-)、百草枯或 Ang II 激活 NADPH 氧化酶可增加 beta(1) 亚基谷胱甘肽化。谷胱甘肽化的增加与α(1)/β(1)亚基免疫共沉淀的减少相关。添加超氧化物歧化酶后，谷胱甘肽作用被逆转。谷氧还蛋白 1 催化去谷胱甘肽化，与 β(1) 亚基发生共免疫沉淀，当包含在贴片移液器溶液中时，可消除百草枯诱导的肌细胞 Na+-K+ 泵电流 (I-p) 抑制。 beta(1) 亚基的半胱氨酸 (Cys46) 可能是谷胱甘肽化的候选者。我们在非洲爪蟾卵母细胞中表达 Na+-K+ 泵 α(1) 亚基和野生型或 Cys46 突变的 β(1) 亚基。 ONOO 诱导 β(1) 亚基的谷胱甘肽化和 Na+-K+ 泵周转数减少。 Cys46 的突变消除了这一点。 ONOO- 还诱导猪肾 Na+-K+ ATP 酶 beta(1) 亚基的谷胱甘肽化。根据 RH421 荧光测定，这与泵的限速 E-2 -> E-1 构象变化大约减少 2 倍相关。我们认为 Na+-K+ 泵的激酶依赖性调节是通过其 Cys46 处的 β(1) 亚基的谷胱甘肽化来实现的。这些发现对以神经激素失调、心肌氧化应激和心肌细胞 Na+ 水平升高为特征的病理生理状况具有影响。 （Circ Res. 2009；105:185-193。）
Angiotensin II (Ang II) inhibits the cardiac sarcolemmal Na+-K+ pump via protein kinase (PK)C-dependent activation of NADPH oxidase. We examined whether this is mediated by oxidative modification of the pump subunits. We detected glutathionylation of beta(1), but not alpha(1), subunits in rabbit ventricular myocytes at baseline. beta(1) Subunit glutathionylation was increased by peroxynitrite (ONOO-), paraquat, or activation of NADPH oxidase by Ang II. Increased glutathionylation was associated with decreased alpha(1)/beta(1) subunit coimmunoprecipitation. Glutathionylation was reversed after addition of superoxide dismutase. Glutaredoxin 1, which catalyzes deglutathionylation, coimmunoprecipitated with beta(1) subunit and, when included in patch pipette solutions, abolished paraquat-induced inhibition of myocyte Na+-K+ pump current (I-p). Cysteine (Cys46) of the beta(1) subunit was the likely candidate for glutathionylation. We expressed Na+-K+ pump alpha(1) subunits with wild-type or Cys46-mutated beta(1) subunits in Xenopus oocytes. ONOO- induced glutathionylation of beta(1) subunit and a decrease in Na+-K+ pump turnover number. This was eliminated by mutation of Cys46. ONOO- also induced glutathionylation of the Na+-K+ ATPase beta(1) subunit from pig kidney. This was associated with a approximate to 2-fold decrease in the rate-limiting E-2 -> E-1 conformational change of the pump, as determined by RH421 fluorescence. We propose that kinase-dependent regulation of the Na+-K+ pump occurs via glutathionylation of its beta(1) subunit at Cys46. These findings have implications for pathophysiological conditions characterized by neurohormonal dysregulation, myocardial oxidative stress and raised myocyte Na+ levels. (Circ Res. 2009;105:185-193.)