Martín-Guerrero, Sandra MCasado, PedroHijazi, MaruanRajeeve, VinothiniPlaza-Díaz, JulioAbadía-Molina, FranciscoNavascués, JulioCuadros, Miguel ACutillas, Pedro RMartín-Oliva, David2023-02-092023-02-092020http://hdl.handle.net/10668/16540Poly(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+.enAttribution 4.0 Internationalhttp://creativecommons.org/licenses/by/4.0/LC–MS/MSPARP-1YAP1cell survivalenergy stressphosphorylationAMP-Activated Protein KinasesAdaptor Proteins, Signal TransducingCell SurvivalHeLa CellsHumansHydrogen PeroxideNADOxidative StressPhosphorylationPoly (ADP-Ribose) Polymerase-1Transcription FactorsYAP-Signaling Proteinsresearch article33146386open access10.1042/BCJ202005251470-8728https://doi.org/10.1042/bcj20200525