Depletion of the Central Metabolite NAD Leads to Oncosis-mediated Cell Death
Depletion of the Central Metabolite NAD Leads to Oncosis-mediated Cell Death
复制标题
DOI：
10.1074/jbc.m114.580159
复制
发表时间：
2014-12-19
影响因子：
4.8
通讯作者：
O'Brien, Thomas
中科院分区：
文献类型：
--
作者：
Del Nagro, Christopher;Xiao, Yang;O'Brien, Thomas
细胞中中心代谢物 NAD 的消耗会导致广泛的代谢缺陷，从而导致细胞死亡，这是肿瘤学中提出的一种新型治疗策略。然而，人们对这种中心代谢物破坏与细胞死亡之间的潜在机制了解有限。在这里，我们利用 GNE-617（一种 NAMPT（NAD 生成所需的限速酶）的小分子抑制剂）来探索 NAD 耗尽后导致细胞死亡的途径。在所有检查的细胞系中，NAMPT 抑制后，NAD 迅速耗尽（平均 t(1/2) 为 8.1 小时）。在 NAD 耗尽的同时，细胞增殖和运动能力均下降，我们将其归因于 NAD 依赖性脱乙酰酶活性降低，因为细胞无法对微管蛋白-K40 和组蛋白 H3-K9 进行脱乙酰化。 NAD 消耗>95% 后，细胞失去再生 ATP 的能力。 ATP 消耗速度较慢的细胞系（平均 t(1/2) 为 45 小时）会激活 caspase-3，并显示细胞凋亡和自噬的证据，而 ATP 快速消耗的细胞系（平均 t(1/2) 为 32 小时）不会激活 caspase-3 或显示细胞凋亡或自噬的迹象。然而，所有细胞系中细胞死亡的主要形式是肿瘤细胞死亡，这是由于一旦 ATP 水平消耗超过 20 倍，质膜稳态就会丧失而导致的。因此，我们的工作阐明了关键代谢物耗尽后细胞中发生的事件顺序，并揭示了 NAD 损失引起的细胞死亡主要是由细胞无法再生 ATP 驱动的。
Depletion of the central metabolite NAD in cells results in broad metabolic defects leading to cell death and is a proposed novel therapeutic strategy in oncology. There is, however, a limited understanding of the underlying mechanisms that connect disruption of this central metabolite with cell death. Here we utilize GNE-617, a small molecule inhibitor of NAMPT, a rate-limiting enzyme required for NAD generation, to probe the pathways leading to cell death following NAD depletion. In all cell lines examined, NAD was rapidly depleted (average t(1/2) of 8.1 h) following NAMPT inhibition. Concurrent with NAD depletion, there was a decrease in both cell proliferation and motility, which we attribute to reduced activity of NAD-dependent deacetylases because cells fail to deacetylate -tubulin-K40 and histone H3-K9. Following depletion of NAD by >95%, cells lose the ability to regenerate ATP. Cell lines with a slower rate of ATP depletion (average t(1/2) of 45 h) activate caspase-3 and show evidence of apoptosis and autophagy, whereas cell lines with rapid depletion ATP (average t(1/2) of 32 h) do not activate caspase-3 or show signs of apoptosis or autophagy. However, the predominant form of cell death in all lines is oncosis, which is driven by the loss of plasma membrane homeostasis once ATP levels are depleted by >20-fold. Thus, our work illustrates the sequence of events that occurs in cells following depletion of a key metabolite and reveals that cell death caused by a loss of NAD is primarily driven by the inability of cells to regenerate ATP.