Mitogen-activated protein kinase-mediated phosphorylation of peroxiredoxin 6 regulates its phospholipase A2 activity
Mitogen-activated protein kinase-mediated phosphorylation of peroxiredoxin 6 regulates its phospholipase A2 activity
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DOI：
10.1042/bj20082061
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发表时间：
2009-05-01
影响因子：
4.1
通讯作者：
Fisher, Aron B.
中科院分区：
文献类型：
--
作者：
Wu, Yongzheng;Feinstein, Sheldon I.;Fisher, Aron B.
Prdx 6（peroxiredoxin 6）是一种具有GSH过氧化物酶和PLA（2）（磷脂酶A（2））[aiPLA（2）（酸性钙非依赖性PLA（2））]活性的双功能蛋白，负责肺表面活性物质磷脂的代谢。我们认为该酶的aiPLA（2）活性是通过磷酸化来调节的。将分离的大鼠肺泡II型细胞（AECII）与PMA（一种PKC（蛋白激酶C）激动剂）孵育，对Prdx 6表达没有影响，但导致aiPLA（2）活性增加约75%，而这种活性可通过用MAPK（丝裂原活化蛋白激酶）抑制剂SB 202190或PD 98059预处理细胞而消除。用PMA孵育AECII后的Prdx 6磷酸化通过用抗磷酸苏氨酸或磷酸丝氨酸抗体免疫沉淀后的放射自显影来证明。在体外，ERK（细胞外信号调节激酶）和p38的几种活性亚型使Prdx 6磷酸化，导致aiPLA（2）活性增加11倍。增加的活性是钙非依赖性的，并且被aiPLA 2抑制剂、表面活性剂蛋白A和十六烷基-3-三氟乙基甘油基-sn-2-磷酸-甲醇（MJ 33）消除。Prdx 6的过氧化物酶活性不受磷酸化的影响。在体外磷酸化的Prdx 6的质谱分析表明，一个独特的磷酸化位点在Thr-177和突变的这个残基废除蛋白质磷酸化和MAPK介导的活性的增加。这些结果表明MAPKs可以介导Prdx 6在Thr-177处的磷酸化，从而显著增加其aiPLA（2）活性。
Prdx6 (peroxiredoxin 6), a bifunctional protein with both GSH peroxidase and PLA(2) (phospholipase A(2)) [aiPLA(2) (acidic calcium-independent PLA(2))] activities, is responsible for the metabolism of lung surfactant phospholipids. We propose that the aiPLA(2) activity of the enzyme is regulated through phosphorylation. Incubation of isolated rat alveolar type II cells (AECII) with PMA, a PKC (protein kinase C) agonist, had no effect on Prdx6 expression but led to similar to 75% increase in aiPLA(2) activity that was abolished by pretreatment of cells with the MAPK (mitogen-activated protein kinase) inhibitors, SB202190 or PD98059. Prdx6 phosphorylation after incubation of AECII with PMA was demonstrated by autoradiography after immunoprecipitation with either anti-phosphothreonine or -phosphoserine antibodies. In vitro, several active isoforms of ERK (extracellular-signal-regulated kinase) and p38 phos-phorylated Prdx6, resulting in an 11-fold increase in aiPLA(2) activity. The increased activity was calcium-independent and was abolished by the aiPLA2 inhibitors, surfactant protein A and hexadecyl-3-trifluorethylglycero-sn-2-phospho-methanol (MJ33). The peroxidase activity of Prdx6 was unaffected by phosphorylation. Mass spectroscopic analysis of in vitro phosphorylated Prdx6 showed a unique phosphorylation site at Thr-177 and mutation of this residue abolished protein phosphorylation and the increase in MAPK-mediated activity. These results show that the MAPKs can mediate phosphorylation of Prdx6 at Thr-177 with a consequent marked increase in its aiPLA(2) activity.