Peroxiredoxin 6 (Prdx6) supports NADPH oxidase1 (Nox1)-based superoxide generation and cell migration.
Peroxiredoxin 6 (Prdx6) supports NADPH oxidase1 (Nox1)-based superoxide generation and cell migration.
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DOI：
10.1016/j.freeradbiomed.2016.04.009
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发表时间：
2016-07
影响因子：
7.4
通讯作者：
Leto TL
中科院分区：
文献类型：
--
作者：
Kwon J;Wang A;Burke DJ;Boudreau HE;Lekstrom KJ;Korzeniowska A;Sugamata R;Kim YS;Yi L;Ersoy I;Jaeger S;Palaniappan K;Ambruso DR;Jackson SH;Leto TL
Nox 1是结肠上皮中活性氧（ROS）的丰富来源，最近显示在伤口愈合和上皮稳态中起作用。在酵母双杂交筛选实验中，我们以Noxa 1的SH 3结构域为诱饵，鉴定了过氧化物氧还蛋白6（Prdx 6）作为Noxa 1的新型结合伴侣。Prdx 6是Prdx抗氧化酶家族的独特成员，表现出谷胱甘肽过氧化物酶和磷脂酶A2活性。我们在过表达两种蛋白质的细胞中证实了这种相互作用，显示Prdx 6结合并稳定野生型Noxa 1，但不结合并稳定SH 3结构域突变形式Noxa 1 W 436 R。我们在几种细胞模型中证明，Prdx 6敲低抑制Nox 1活性，而增强的Prdx 6表达支持更高的Nox 1衍生的超氧化物产生。过氧化物酶和脂肪酶缺陷的突变形式的Prdx 6（Prdx 6 C47 S和S32 A，分别）未能结合或稳定的Nox 1组分或支持Nox 1介导的超氧化物生成。此外，Prdx 6磷脂酶A2活性的过渡态底物类似物抑制剂（MJ-33）显示抑制Nox 1活性，表明Nox 1活性受Prdx 6的磷脂酶活性调节。最后，野生型Prdx 6，而不是脂肪酶或过氧化物酶突变形式，支持Nox 1介导的细胞迁移的HCT-116结肠上皮细胞模型的伤口闭合。这些发现强调了一种新的途径，其中这种抗氧化酶积极调节氧化剂生成系统，以支持细胞迁移和伤口愈合。
Nox1 is an abundant source of reactive oxygen species (ROS) in colon epithelium recently shown to function in wound healing and epithelial homeostasis. We identified Peroxiredoxin 6 (Prdx6) as a novel binding partner of Nox activator 1 (Noxa1) in yeast two-hybrid screening experiments using the Noxa1 SH3 domain as bait. Prdx6 is a unique member of the Prdx antioxidant enzyme family exhibiting both glutathione peroxidase and phospholipase A2 activities. We confirmed this interaction in cells overexpressing both proteins, showing Prdx6 binds to and stabilizes wild type Noxa1, but not the SH3 domain mutant form, Noxa1 W436R. We demonstrated in several cell models that Prdx6 knockdown suppresses Nox1 activity, whereas enhanced Prdx6 expression supports higher Nox1-derived superoxide production. Both peroxidase- and lipase-deficient mutant forms of Prdx6 (Prdx6 C47S and S32A, respectively) failed to bind to or stabilize Nox1 components or support Nox1-mediated superoxide generation. Furthermore, the transition-state substrate analogue inhibitor of Prdx6 phospholipase A2 activity (MJ-33) was shown to suppress Nox1 activity, suggesting Nox1 activity is regulated by the phospholipase activity of Prdx6. Finally, wild type Prdx6, but not lipase or peroxidase mutant forms, supports Nox1-mediated cell migration in the HCT-116 colon epithelial cell model of wound closure. These findings highlight a novel pathway in which this antioxidant enzyme positively regulates an oxidant-generating system to support cell migration and wound healing.