Phosphatidylinositol (4,5)-bisphosphate Modulates Nox5 Localization via an N-Terminal Polybasic Region
Phosphatidylinositol (4,5)-bisphosphate Modulates Nox5 Localization via an N-Terminal Polybasic Region
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DOI：
10.1091/mbc.e07-12-1223
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发表时间：
2008-10-01
影响因子：
3.3
通讯作者：
Lambeth, J. David
中科院分区：
文献类型：
--
作者：
Kawahara, Tsukasa;Lambeth, J. David
Nox 5是一种能产生活性氧（ROS）的NADPH氧化酶，含有两个保守的多碱基区：一个位于N端（PBR-N），位于第四个EF-手和第一个跨膜区之间;一个位于C端（PBR-C），位于第一和第二个NADPH结合亚区之间。在这里，我们表明，磷脂酰肌醇（4，5）-二磷酸[PtdIns（4，5）P-2]，一个主要的磷脂酰肌醇在质膜上，结合到人类Nox 5导致Nox 5本地化从内膜到质膜。在完整细胞中PtdIns（4，5）P-2水平的酶促调节改变了Nox 5的细胞表面定位，与细胞外ROS产生平行。PBR-N的突变阻止了PtdIns（4，5）P-2依赖性的Nox 5定位于质膜，并降低了细胞外ROS的产生。对应于PBR-N的合成肽与PtdIns（4，5）P-2结合，但不与PtdIns结合，而PBR-N肽中的突变废除了与PtdIns（4，5）P-2的结合。PBR-N中的精氨酸-197是调节Nox 5亚细胞定位及其与PtdIns（4，5）P-2相互作用的关键残基。相反，PBR-C中的突变不影响定位。因此，通过将Nox 5定位于质膜，PtdIns（4，5）P-2调节Nox 5的细胞外ROS产生。
Nox5, an EF-hand-containing reactive oxygen species (ROS)-generating NADPH oxidase, contains two conserved polybasic regions: one N-terminal (PBR-N), located between the fourth EF-hand and the first transmembrane region, and one C-terminal (PBR-C), between the first and second NADPH-binding subregions. Here, we show that phosphatidylinositol (4,5)-bisphosphate [PtdIns(4,5)P-2], a major phosphoinositide in plasma membrane, binds to human Nox5 causing Nox5 to localize from internal membranes to the plasma membrane. Enzymatic modulation of PtdIns(4,5)P-2 levels in intact cells altered cell surface localization of Nox5 in parallel with extracellular ROS generation. Mutations in PBR-N prevented PtdIns(4,5)P-2- dependent localization of Nox5 to the plasma membrane and decreased extracellular ROS production. A synthetic peptide corresponding to PBR-N bound to PtdIns(4,5)P-2, but not to PtdIns, whereas mutations in the PBR-N peptide abrogated the binding to PtdIns(4,5)P-2. Arginine-197 in PBR-N was a key residue to regulate subcellular localization of Nox5 and its interaction with PtdIns(4,5)P-2. In contrast, mutation in PBR-C did not affect localization. Thus, extracellular ROS production by Nox5 is modulated by PtdIns(4,5)P-2 by localizing Nox5 to the plasma membrane.