title: ATR inhibition rewires cellular signaling networks induced by replication stress.
abstract: The slowing down or stalling of replication forks is commonly known as replication stress and arises from multiple causes such as DNA lesions, nucleotide depletion, RNA-DNA hybrids, and oncogene activation. The ataxia telangiectasia and Rad3-related kinase (ATR) plays an essential role in the cellular response to replication stress and inhibition of ATR has emerged as therapeutic strategy for the treatment of cancers that exhibit high levels of replication stress. However, the cellular signaling induced by replication stress and the substrate spectrum of ATR has not been systematically investigated. In this study, we employed quantitative MS-based proteomics to define the cellular signaling after nucleotide depletion-induced replication stress and replication fork collapse following ATR inhibition. We demonstrate that replication stress results in increased phosphorylation of a subset of proteins, many of which are involved in RNA splicing and transcription and have previously not been associated with the cellular replication stress response. Furthermore, our data reveal the ATR-dependent phosphorylation following replication stress and discover novel putative ATR target sites on MCM6, TOPBP1, RAD51AP1, and PSMD4. We establish that ATR inhibition rewires cellular signaling networks induced by replication stress and leads to the activation of the ATM-driven double-strand break repair signaling.
authors.affiliation: Department of Medicine, Hematology/Oncology, Goethe University, Frankfurt, Germany.|German Cancer Consortium (DKTK), Heidelberg, Germany.|German Cancer Research Center (DKFZ), Heidelberg, Germany.
authors.forename: Sebastian A
authors.initials: SA
authors.lastname: Wagner
authors.affiliation: Department of Medicine, Hematology/Oncology, Goethe University, Frankfurt, Germany.
authors.forename: Hannah
authors.initials: H
authors.lastname: Oehler
authors.affiliation: Institute of Molecular Biology (IMB), Mainz, Germany.
authors.forename: Andrea
authors.initials: A
authors.lastname: Voigt
authors.affiliation: Department of Medicine, Hematology/Oncology, Goethe University, Frankfurt, Germany.
authors.forename: Denis
authors.initials: D
authors.lastname: Dalic
authors.affiliation: Institute of Molecular Biology (IMB), Mainz, Germany.
authors.forename: Anja
authors.initials: A
authors.lastname: Freiwald
authors.affiliation: Department of Medicine, Hematology/Oncology, Goethe University, Frankfurt, Germany.|German Cancer Consortium (DKTK), Heidelberg, Germany.|German Cancer Research Center (DKFZ), Heidelberg, Germany.
authors.forename: Hubert
authors.initials: H
authors.lastname: Serve
authors.affiliation: Institute of Molecular Biology (IMB), Mainz, Germany.
authors.forename: Petra
authors.initials: P
authors.lastname: Beli
mesh_terms: D000595:Amino Acid Sequence; D000970:Antineoplastic Agents / Q000494:pharmacology*; D064007:Ataxia Telangiectasia Mutated Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects / Q000235:genetics; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004261:DNA Replication / Q000187:drug effects*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D053263:Gene Regulatory Networks; D006801:Humans; D006918:Hydroxyurea / Q000494:pharmacology*; D064151:Minichromosome Maintenance Complex Component 6 / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D010006:Osteoblasts / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D046988:Proteasome Endopeptidase Complex / Q000235:genetics / Q000378:metabolism; D025941:Protein Interaction Mapping; D012326:RNA Splicing; D016601:RNA-Binding Proteins; D015398:Signal Transduction / Q000187:drug effects*; D013312:Stress, Physiological; D014158:Transcription, Genetic
chemical_list: D000970:Antineoplastic Agents; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; C498506:PSMD4 protein, human; D010750:Phosphoproteins; C110410:RAD51AP1 protein, human; D016601:RNA-Binding Proteins; C110513:TOPBP1 protein, human; D004247:DNA; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D046988:Proteasome Endopeptidase Complex; C103258:MCM6 protein, human; D064151:Minichromosome Maintenance Complex Component 6; D006918:Hydroxyurea
keywords: ATR kinase; Cancer; DNA replication stress; Phosphoproteomics; Quantitative MS; Systems biology