Poly(ADP-ribose) polymerase 1 (PARP-1) is a nuclear enzyme that catalyze the transfer of ADP-ribose units from NAD+ to several target proteins involved in cellular stress responses. Using WRL68 (HeLa derivate) cells, we previously showed that PARP-1 activation induced by oxidative stress after H2O2 treatment lead to depletion of cellular NAD+ and ATP, which promoted cell death. In this work, LC–MS/MS-based phosphoproteomics in WRL68 cells showed that the oxidative damage induced by H2O2 increased the phosphorylation of YAP1, a transcriptional co-activator involved in cell survival, and modified the phosphorylation of other proteins involved in transcription. Genetic or pharmacological inhibition of PARP-1 in H2O2-treated cells reduced YAP1 phosphorylation and degradation and increased cell viability. YAP1 silencing abrogated the protective effect of PARP-1 inhibition, indicating that YAP1 is important for the survival of WRL68 cells exposed to oxidative damage. Supplementation of NAD+ also reduced YAP1 phosphorylation, suggesting that the loss of cellular NAD+ caused by PARP-1 activation after oxidative treatment is responsible for the phosphorylation of YAP1. Finally, PARP-1 silencing after oxidative treatment diminished the activation of the metabolic sensor AMPK. Since NAD+ supplementation reduced the phosphorylation of some AMPK substrates, we hypothesized that the loss of cellular NAD+ after PARP-1 activation may induce an energy stress that activates AMPK. In summary, we showed a new crucial role of PARP-1 in the response to oxidative stress in which PARP-1 activation reduced cell viability by promoting the phosphorylation and degradation of YAP1 through a mechanism that involves the depletion of NAD+.

中文翻译：

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氧化损伤后PARP-1激活促进YAP1的能量应激依赖性磷酸化并降低细胞活力

聚（ADP-核糖）聚合酶1（PARP-1）是一种核酶，可催化ADP-核糖单元从NAD +向涉及细胞应激反应的几种靶蛋白的转移。使用WRL68（HeLa衍生物）细胞，我们先前显示过氧化氢处理后氧化应激诱导的PARP-1活化导致细胞NAD +和ATP耗竭，从而促进细胞死亡。在这项工作中，WRL68细胞中基于LC–MS / MS的磷酸蛋白质组学表明，H2O2诱导的氧化损伤增加了YAP1的磷酸化，YAP1是参与细胞存活的转录共激活因子，并修饰了其他参与转录的蛋白质的磷酸化。在H2O2处理的细胞中，PARP-1的遗传或药理抑制作用降低了YAP1的磷酸化和降解，并提高了细胞活力。YAP1沉默取消了PARP-1抑制的保护作用，表明YAP1对于暴露于氧化损伤的WRL68细胞的存活很重要。补充NAD +还可以减少YAP1的磷酸化，这表明氧化处理后由PARP-1激活引起的细胞NAD +的丢失是YAP1磷酸化的原因。最终，氧化处理后的PARP-1沉默降低了代谢传感器AMPK的激活。由于NAD +的添加减少了某些AMPK底物的磷酸化，因此我们假设PARP-1激活后细胞NAD +的丢失可能会诱导激活AMPK的能量应激。综上所述，