title: PARP-1 modulation of mTOR signaling in response to a DNA alkylating agent.
abstract: Poly(ADP-ribose) polymerase-1 (PARP-1) is widely involved in cell death responses. Depending on the degree of injury and on cell type, PARP activation may lead to autophagy, apoptosis or necrosis. In HEK293 cells exposed to the alkylating agent N-methyl-N'-nitro-N'-nitrosoguanine (MNNG), we show that PARP-1 activation triggers a necrotic cell death response. The massive poly(ADP-ribose) (PAR) synthesis following PARP-1 activation leads to the modulation of mTORC1 pathway. Shortly after MNNG exposure, NAD⁺ and ATP levels decrease, while AMP levels drastically increase. We characterized at the molecular level the consequences of these altered nucleotide levels. First, AMP-activated protein kinase (AMPK) is activated and the mTORC1 pathway is inhibited by the phosphorylation of Raptor, in an attempt to preserve cellular energy. Phosphorylation of the mTORC1 target S6 is decreased as well as the phosphorylation of the mTORC2 component Rictor on Thr1135. Finally, Akt phosphorylation on Ser473 is lost and then, cell death by necrosis occurs. Inhibition of PARP-1 with the potent PARP inhibitor AG14361 prevents all of these events. Moreover, the antioxidant N-acetyl-L-cysteine (NAC) can also abrogate all the signaling events caused by MNNG exposure suggesting that reactive oxygen species (ROS) production is involved in PARP-1 activation and modulation of mTOR signaling. In this study, we show that PARP-1 activation and PAR synthesis affect the energetic status of cells, inhibit the mTORC1 signaling pathway and possibly modulate the mTORC2 complex affecting cell fate. These results provide new evidence that cell death by necrosis is orchestrated by the balance between several signaling pathways, and that PARP-1 and PAR take part in these events.
authors.affiliation: Cancer Axis, CHUQ Research Center and Faculty of Medicine, Laval University, Quebec City, Quebec, Canada.
authors.forename: Chantal
authors.initials: C
authors.lastname: Ethier
authors.forename: Maxime
authors.initials: M
authors.lastname: Tardif
authors.forename: Laura
authors.initials: L
authors.lastname: Arul
authors.forename: Guy G
authors.initials: GG
authors.lastname: Poirier
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism; D000249:Adenosine Monophosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000477:Alkylating Agents / Q000494:pharmacology*; D001343:Autophagy / Q000187:drug effects; D015153:Blotting, Western; D004789:Enzyme Activation / Q000187:drug effects; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D008769:Methylnitronitrosoguanidine / Q000494:pharmacology; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D008954:Models, Biological; D009243:NAD / Q000378:metabolism; D009336:Necrosis; D000071137:Poly (ADP-Ribose) Polymerase-1; D011064:Poly Adenosine Diphosphate Ribose / Q000378:metabolism; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism*; D034622:RNA Interference; D015398:Signal Transduction / Q000187:drug effects*; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*
chemical_list: D000477:Alkylating Agents; C477012:MAP1LC3A protein, human; D008869:Microtubule-Associated Proteins; D009243:NAD; D008769:Methylnitronitrosoguanidine; D011064:Poly Adenosine Diphosphate Ribose; D000249:Adenosine Monophosphate; D000255:Adenosine Triphosphate; C499320:PARP1 protein, human; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; C546842:MTOR protein, human; C526181:PRKAA2 protein, human; D058570:TOR Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases; C526180:PRKAA1 protein, human