Department of Biological Sciences, University of Calgary, Alberta T2N 1N4, Canada.
J Biol Chem. 2010 Aug 13;285(33):25731-42. doi: 10.1074/jbc.M110.122200. Epub 2010 Jun 10.
We have previously established that the anti-cancer lysophospholipid edelfosine (1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine, Et-18-OCH(3)) induces cell death in yeast by selective modification of lipid raft composition at the plasma membrane. In this study we determined that alpha-tocopherol protects cells from the edelfosine cytotoxic effect, preventing the internalization of sterols and the plasma membrane proton pump ATPase, Pma1p. Two non-mutually exclusive hypotheses were considered to explain the protective effect of alpha-tocopherol: (i) its classical antioxidant activity is necessary to break progression of lipid peroxidation, despite the fact Saccharomyces cerevisiae does not possess polyunsaturated fatty acids and (ii) due to its complementary cone shape, insertion of alpha-tocopherol could correct membrane curvature stress imposed by edelfosine (inverted cone shape). We then developed tools to distinguish between these two hypotheses and dissect the structural requirements that confer alpha-tocopherol its protective effect. Our results indicated its lipophilic nature and the H donating hydroxyl group from the chromanol ring are both required to counteract the cytotoxic effect of edelfosine, suggesting edelfosine induces oxidation of membrane components. To further support this finding and learn more about the early cellular response to edelfosine we investigated the role that known oxidative stress signaling pathways play in modulating sensitivity to the lipid drug. Our results indicate the transcription factors Yap1 and Skn7 as well as the major peroxiredoxin, Tsa1, mediate a response to edelfosine. Interestingly, the pathway differed from the one triggered by hydrogen peroxide and its activation (measured as Yap1 translocation to the nucleus) was abolished by co-treatment of the cells with alpha-tocopherol.
我们之前已经证实，抗癌溶血磷脂酰乙醇胺（1-O-十八烷基-2-O-甲基-rac-甘油-3-磷酸胆碱，Et-18-OCH(3)）通过选择性修饰质膜上的脂筏组成诱导酵母细胞死亡。在这项研究中，我们确定α-生育酚可保护细胞免受埃德尔福辛的细胞毒性作用，防止固醇和质膜质子泵 ATP 酶 Pma1p 的内化。提出了两个非相互排斥的假说来解释 α-生育酚的保护作用：（i）尽管酿酒酵母不具有多不饱和脂肪酸，但它的经典抗氧化活性对于打破脂质过氧化的进展是必要的；（ii）由于其互补的锥形形状，α-生育酚的插入可以纠正埃德尔福辛（倒锥形）施加的膜曲率应力。然后，我们开发了工具来区分这两个假说，并剖析赋予 α-生育酚其保护作用的结构要求。我们的结果表明，其亲脂性和来自色满醇环的 H 供体羟基都需要对抗埃德尔福辛的细胞毒性作用，这表明埃德尔福辛诱导膜成分的氧化。为了进一步支持这一发现并更多地了解细胞对埃德尔福辛的早期反应，我们研究了已知的氧化应激信号通路在调节对脂质药物的敏感性方面所起的作用。我们的结果表明，转录因子 Yap1 和 Skn7 以及主要的过氧化物酶 Tsa1 介导了对埃德尔福辛的反应。有趣的是，该途径与由过氧化氢触发的途径不同，并且细胞与 α-生育酚共同处理时，其激活（如 Yap1 向核内易位的测量）被废除。
