Peroxiredoxin 6 Phosphorylation and Subsequent Phospholipase A2 Activity Are Required for Agonist-mediated Activation of NADPH Oxidase in Mouse Pulmonary Microvascular Endothelium and Alveolar Macrophages
Peroxiredoxin 6 Phosphorylation and Subsequent Phospholipase A2 Activity Are Required for Agonist-mediated Activation of NADPH Oxidase in Mouse Pulmonary Microvascular Endothelium and Alveolar Macrophages
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DOI：
10.1074/jbc.m110.206623
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发表时间：
2011-04-01
影响因子：
4.8
通讯作者：
Fisher, Aron B.
中科院分区：
文献类型：
--
作者：
Chatterjee, Shampa;Feinstein, Sheldon I.;Fisher, Aron B.
过氧化物氧还蛋白6（Peroxiredoxin 6，Prdx 6）是一种具有谷胱甘肽过氧化物酶和磷脂酶A2（phospholipase A2，PLA（2））活性的双功能酶，参与中性粒细胞中NADPH氧化酶2（NADPH oxidase 2，NOX 2）的活化，但其作用机制尚不清楚。我们现在证明，Prdx 6是激动剂诱导的肺微血管内皮细胞（PMVEC）中的NOX 2激活所必需的，并且该作用需要Prdx 6的PLA（2）活性。反应性氧簇（ROS）的产生在血管紧张素II（Ang II）或佛波醇12-肉豆蔻酸酯13-乙酸酯显着减少灌注肺和分离的PMVEC从Prdx 6 null小鼠。Rac 1和p47（phox），胞质组分的NOX 2，易位到内皮细胞膜后，血管紧张素II治疗野生型，但不是Prdx 6无效PMVEC。MJ 33是Prdx 6 PLA（2）活性的抑制剂，可阻断PMVEC中激动剂诱导的PLA（2）活性和ROS生成，阻断率> 80%，而其他PLA（2）的抑制剂无效。用野生型和C47 S突变体Prdx 6转染Prx 6缺失细胞，但不转染PLA（2）活性位点突变体（S32 A、H26 A和D140 A），“拯救”了Ang II诱导的PLA（2）活性和ROS生成。野生型细胞的血管紧张素II治疗导致Prdx 6的磷酸化及其随后的易位从胞质溶胶到细胞膜。MAPK抑制剂U 0126显著降低磷酸化以及PLA（2）活性和ROS生成。因此，激动剂诱导的MAPK活化导致Prdx 6磷酸化并易位至细胞膜，在细胞膜处其PLA（2）活性促进NOX 2复合物的组装和氧化酶的活化。
Peroxiredoxin 6 (Prdx6), a bifunctional enzyme with glutathione peroxidase and phospholipase A2 (PLA(2)) activities, participates in the activation of NADPH oxidase 2 (NOX2) in neutrophils, but the mechanism for this effect is not known. We now demonstrate that Prdx6 is required for agonist-induced NOX2 activation in pulmonary microvascular endothelial cells (PMVEC) and that the effect requires the PLA(2) activity of Prdx6. Generation of reactive oxygen species (ROS) in response to angiotensin II (Ang II) or phorbol 12-myristate 13-acetate was markedly reduced in perfused lungs and isolated PMVEC from Prdx6 null mice. Rac1 and p47(phox), cytosolic components of NOX2, translocated to the endothelial cell membrane after Ang II treatment in wild-type but not Prdx6 null PMVEC. MJ33, an inhibitor of Prdx6 PLA(2) activity, blocked agonist-induced PLA(2) activity and ROS generation in PMVEC by > 80%, whereas inhibitors of other PLA(2)s were ineffective. Transfection of Prx6 null cells with wild-type and C47S mutant Prdx6, but not with mutants of the PLA(2) active site (S32A, H26A, and D140A), "rescued" Ang II-induced PLA(2) activity and ROS generation. Ang II treatment of wild-type cells resulted in phosphorylation of Prdx6 and its subsequent translocation from the cytosol to the cell membrane. Phosphorylation as well as PLA(2) activity and ROS generation were markedly reduced by the MAPK inhibitor, U0126. Thus, agonist-induced MAPK activation leads to Prdx6 phosphorylation and translocation to the cell membrane, where its PLA(2) activity facilitates assembly of the NOX2 complex and activation of the oxidase.