Department of Foods and Nutrition, Purdue University, Stone Hall, 700 W. State Street, West Lafayette, Indiana 47907-2059, USA.
J Cell Biochem. 2010 Aug 15;110(6):1504-11. doi: 10.1002/jcb.22724.
ENOX2 (tNOX), a tumor-associated cell surface ubiquinol (NADH) oxidase, functions as an alternative terminal oxidase for plasma membrane electron transport. Ubiquitous in all cancer cell lines studied thus far, ENOX2 expression correlates with the abnormal growth and division associated with the malignant phenotype. ENOX2 has been proposed as the cellular target for various quinone site inhibitors that demonstrate anticancer activity such as the green tea constituent epigallocatechin-3-gallate (EGCg) and the isoflavone phenoxodiol (PXD). Here we present a possible mechanism that explains how these substances result in apoptosis in cancer cells by ENOX2-mediated alterations of cytosolic amounts of NAD(+) and NADH. When ENOX2 is inhibited, plasma membrane electron transport is diminished, and cytosolic NADH accumulates. We show in HeLa cells that NADH levels modulate the activities of two pivotal enzymes of sphingolipid metabolism: sphingosine kinase 1 (SK1) and neutral sphingomyelinase (nSMase). Their respective products sphingosine 1-phosphate (S1P) and ceramide (Cer) are key determinants of cell fate. S1P promotes cell survival and Cer promotes apoptosis. Using plasma membranes isolated from cervical adenocarcinoma (HeLa) cells as well as purified proteins of both bacterial and human origin, we demonstrate that NADH inhibits SK1 and stimulates nSMase, while NAD(+) inhibits nSMase and has no effect on SK1. Additionally, intact HeLa cells treated with ENOX2 inhibitors exhibit an increase in Cer and a decrease in S1P. Treatments that stimulate cytosolic NADH production potentiate the antiproliferative effects of ENOX2 inhibitors while those that attenuate NADH production or stimulate plasma membrane electron transport confer a survival advantage.
ENOX2（tNOX），一种肿瘤相关的细胞表面泛醇（NADH）氧化酶，作为质膜电子传递的替代末端氧化酶发挥作用。迄今为止，在所有研究的癌细胞系中普遍存在，ENOX2 的表达与与恶性表型相关的异常生长和分裂相关。ENOX2 已被提议作为各种醌部位抑制剂的细胞靶标，这些抑制剂如绿茶成分表没食子儿茶素-3-没食子酸酯（EGCg）和异黄酮 Phenoxodiol（PXD），具有抗癌活性。在这里，我们提出了一种可能的机制，解释了这些物质如何通过 ENOX2 介导的细胞质 NAD（+）和 NADH 量的改变导致癌细胞凋亡。当 ENOX2 被抑制时，质膜电子传递减少，细胞质 NADH 积累。我们在 HeLa 细胞中表明，NADH 水平调节鞘脂代谢的两个关键酶的活性：鞘氨醇激酶 1（SK1）和中性鞘磷脂酶（nSMase）。它们各自的产物 1-磷酸鞘氨醇（S1P）和神经酰胺（Cer）是细胞命运的关键决定因素。S1P 促进细胞存活，Cer 促进细胞凋亡。使用来自宫颈腺癌（HeLa）细胞的质膜分离物以及来自细菌和人类来源的纯化蛋白，我们证明 NADH 抑制 SK1 并刺激 nSMase，而 NAD（+）抑制 nSMase 并且对 SK1 没有影响。此外，用 ENOX2 抑制剂处理的完整 HeLa 细胞表现出 Cer 的增加和 S1P 的减少。刺激细胞质 NADH 产生的处理增强了 ENOX2 抑制剂的抗增殖作用，而那些减弱 NADH 产生或刺激质膜电子传递的处理赋予了生存优势。
