Dusi S, Della Bianca V, Grzeskowiak M, Rossi F
Istituto di Patologia Generale, Strada Le Grazie, Verona, Italy.
Biochem J. 1993 Feb 15;290 ( Pt 1)(Pt 1):173-8. doi: 10.1042/bj2900173.
Stimulation of neutrophils with different agonists activates a latent multicomponent NADPH oxidase that reduces molecular oxygen to superoxide anion. Evidence has accumulated that phosphorylation of p47phox (the 47 kDa cytosolic phagocyte oxidase factor) and translocation of the two cytosolic components p47phox and p67phox are essential steps in the activation of NADPH oxidase in response to phorbol esters. We analysed the relationships between activation of the NADPH oxidase and phosphorylation and translocation of p47phox and p67phox in normal and Ca(2+)-depleted neutrophils stimulated by the receptor-mediated agonists formyl-methionyl-leucyl-phenylalanine and concanavalin A. The results produced the following conclusions: (1) Translocation of p47phox and p67phox is an essential mechanism for activation of the NADPH oxidase. (2) A continuous translocation of p47phox and p67phox is necessary to maintain the NADPH oxidase in an activated state. (3) Only a fraction of p47phox and p67phox translocated to the plasma membrane is functional for the activation of the oxidase. (4) Translocation is independent of protein kinase C, and is linked to transmembrane signalling involving Ca2+ transients and production of lipidic second messengers. However, under some conditions, such as in Ca(2+)-depleted neutrophils, translocation can also occur independently of signalling pathways involving production of second messengers from hydrolysis of phospholipids and Ca2+ transients. (5) Phosphorylation of p47phox and p67phox can be quantitatively dissociated from translocation, as staurosporine markedly inhibits phosphorylation but not translocation. (6) The activity of NADPH oxidase is not correlated with the amounts of the phosphorylated proteins present in the plasma membrane.
用不同激动剂刺激中性粒细胞会激活一种潜在的多组分NADPH氧化酶，该酶可将分子氧还原为超氧阴离子。越来越多的证据表明，p47phox（47 kDa胞质吞噬细胞氧化酶因子）的磷酸化以及两个胞质组分p47phox和p67phox的转位是佛波酯刺激下NADPH氧化酶激活的关键步骤。我们分析了在受体介导的激动剂甲酰甲硫氨酰亮氨酰苯丙氨酸和伴刀豆球蛋白A刺激下，正常和Ca(2+) 耗尽的中性粒细胞中NADPH氧化酶的激活与p47phox和p67phox的磷酸化及转位之间的关系。结果得出以下结论：(1)p47phox和p67phox的转位是NADPH氧化酶激活的关键机制。(2)p47phox和p67phox的持续转位对于维持NADPH氧化酶的激活状态是必要的。(3)只有转位到质膜的一部分p47phox和p67phox对氧化酶的激活具有功能作用。(4)转位独立于蛋白激酶C，并与涉及Ca2+瞬变和脂质第二信使产生的跨膜信号传导相关。然而，在某些条件下，如在Ca(2+) 耗尽的中性粒细胞中，转位也可独立于涉及磷脂水解产生第二信使和Ca2+瞬变的信号通路发生。(5)p47phox和p67phox的磷酸化在数量上可与转位分离，因为星形孢菌素显著抑制磷酸化但不抑制转位。(6)NADPH氧化酶的活性与质膜中存在的磷酸化蛋白的量无关。
