title: Phosphoproteomic analysis of cells treated with longevity-related autophagy inducers.
abstract: Macroautophagy is a self-cannibalistic process that enables cells to adapt to various stresses and maintain energy homeostasis. Additionally, autophagy is an important route for turnover of misfolded proteins and damaged organelles, with important implications in cancer, neurodegenerative diseases and aging. Resveratrol and spermidine are able to induce autophagy by affecting deacetylases and acetylases, respectively, and have been found to extend the life-span of model organisms. With the aim to reveal the signaling networks involved in this drug-induced autophagic response, we quantified resveratrol and spermidine-induced changes in the phosphoproteome using SILAC and mass spectrometry. The data were subsequently analyzed using the NetworKIN algorithm to extract key features of the autophagy-responsive kinase-substrate network. We found that two distinct sequence motifs were highly responsive to resveratrol and spermidine and that key proteins modulating the acetylation, phosphorylation, methylation and ubiquitination status were affected by changes in phosphorylation during the autophagic response. Essential parts of the apoptotic signaling network were subjected to post-translational modifications during the drug-induced autophagy response, suggesting potential crosstalk and balancing between autophagy and apoptosis. Additionally, we predicted cellular signaling networks affected by resveratrol and spermidine using a computational framework. Altogether, these results point to a profound crosstalk between distinct networks of post-translational modifications and provide a resource for future analysis of autophagy and cell death.
authors.affiliation: Center for Experimental BioInformatics, Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense, Denmark.
authors.forename: Martin V
authors.initials: MV
authors.lastname: Bennetzen
authors.forename: Guillermo
authors.initials: G
authors.lastname: Mariño
authors.forename: Dennis
authors.initials: D
authors.lastname: Pultz
authors.forename: Eugenia
authors.initials: E
authors.lastname: Morselli
authors.forename: Nils J
authors.initials: NJ
authors.lastname: Færgeman
authors.forename: Guido
authors.initials: G
authors.lastname: Kroemer
authors.forename: Jens S
authors.initials: JS
authors.lastname: Andersen
mesh_terms: D000107:Acetylation; D000465:Algorithms; D020816:Amino Acid Motifs; D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis; D001343:Autophagy; D019295:Computational Biology / Q000379:methods; D051357:Cyclin-Dependent Kinase 2 / Q000378:metabolism; D004789:Enzyme Activation; D059585:G1 Phase Cell Cycle Checkpoints; D045325:HCT116 Cells; D006801:Humans; D008136:Longevity / Q000187:drug effects; D013058:Mass Spectrometry; D008745:Methylation; D010750:Phosphoproteins / Q000032:analysis* / Q000378:metabolism; D010766:Phosphorylation; D025941:Protein Interaction Mapping; D060066:Protein Interaction Maps; D040901:Proteomics; D000077185:Resveratrol; D015398:Signal Transduction; D013095:Spermidine / Q000494:pharmacology*; D013267:Stilbenes / Q000494:pharmacology*; D054875:Ubiquitination
chemical_list: D000970:Antineoplastic Agents; D010750:Phosphoproteins; D013267:Stilbenes; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; D000077185:Resveratrol; D013095:Spermidine