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Acetylation dynamics of human nuclear proteins during the ionizing radiation-induced DNA damage response.

ABSTRACT: Genotoxic insults, such as ionizing radiation (IR), cause DNA damage that evokes a multifaceted cellular DNA damage response (DDR). DNA damage signaling events that control protein activity, subcellular localization, DNA binding, protein-protein interactions, etc. rely heavily on time-dependent posttranslational modifications (PTMs). To complement our previous analysis of IR-induced temporal dynamics of nuclear phosphoproteome, we now identify a range of human nuclear proteins that are dynamically regulated by acetylation, and predominantly deacetylation, during IR-induced DDR by using mass spectrometry-based proteomic approaches. Apart from cataloging acetylation sites through SILAC proteomic analyses before IR and at 5 and 60 min after IR exposure of U2OS cells, we report that: (1) key components of the transcriptional machinery, such as EP300 and CREBBP, are dynamically acetylated; (2) that nuclear acetyltransferases themselves are regulated, not on the protein abundance level, but by (de)acetylation; and (3) that the recently reported p53 co-activator and methyltransferase MLL3 is acetylated on five lysines during the DDR. For selected examples, protein immunoprecipitation and immunoblotting were used to assess lysine acetylation status and thereby validate the mass spectrometry data. We thus present evidence that nuclear proteins, including those known to regulate cellular functions via epigenetic modifications of histones, are regulated by (de)acetylation in a timely manner upon cell's exposure to genotoxic insults. Overall, these results present a resource of temporal profiles of a spectrum of protein acetylation sites during DDR and provide further insights into the highly dynamic nature of regulatory PTMs that help orchestrate the maintenance of genome integrity.

SUBMITTER: Bennetzen MV

PROVIDER: S-EPMC3713127 | biostudies-literature | 2013 Jun

REPOSITORIES: biostudies-literature

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Acetylation dynamics of human nuclear proteins during the ionizing radiation-induced DNA damage response.

Bennetzen Martin V MV Larsen Dorthe Helena DH Dinant Christoffel C Watanabe Sugiko S Bartek Jiri J Lukas Jiri J Andersen Jens S JS

Cell cycle (Georgetown, Tex.) 20130501 11

Genotoxic insults, such as ionizing radiation (IR), cause DNA damage that evokes a multifaceted cellular DNA damage response (DDR). DNA damage signaling events that control protein activity, subcellular localization, DNA binding, protein-protein interactions, etc. rely heavily on time-dependent posttranslational modifications (PTMs). To complement our previous analysis of IR-induced temporal dynamics of nuclear phosphoproteome, we now identify a range of human nuclear proteins that are dynamical ...[more]

PMID: 23656789

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Project description:Common cancer therapies employ chemicals or radiation that damage DNA. Cancer and normal cells respond to DNA damage by activating complex networks of DNA damage sensor, signal transducer, and effector proteins that arrest cell cycle progression, and repair damaged DNA. If damage is severe enough, the DNA damage response (DDR) triggers programed cell death by apoptosis or other pathways. Caspase 3 is a protease that is activated upon damage and triggers apoptosis, and production of prostaglandin E2 (PGE2), a potent growth factor that can enhance growth of surviving cancer cells leading to accelerated tumor repopulation. Thus, dying tumor cells can promote growth of surviving tumor cells, a pathway aptly named Phoenix Rising. In the present study, we surveyed Phoenix Rising responses in a variety of normal and established cancer cell lines, and in cancer cell lines freshly derived from patients. We demonstrate that IR induces a Phoenix Rising response in many, but not all cell lines, and that PGE2 production generally correlates with enhanced growth of cells that survive irradiation, and of unirradiated cells co-cultured with irradiated cells. We show that PGE2 production is stimulated by low and high LET ionizing radiation, and can be enhanced or suppressed by inhibitors of key DDR proteins. PGE2 is produced downstream of caspase 3 and the cyclooxygenases COX1 and COX2, and we show that the pan COX1-2 inhibitor indomethacin blocks IR-induced PGE2 production in the presence or absence of DDR inhibitors. COX1-2 require oxygen for catalytic activity, and we further show that PGE2 production is markedly suppressed in cells cultured under low (1%) oxygen concentration. Thus, Phoenix Rising is most likely to cause repopulation of tumors with relatively high oxygen, but not in hypoxic tumors. This survey lays a foundation for future studies to further define tumor responses to radiation and inhibitors of the DDR and Phoenix Rising to enhance the efficacy of radiotherapy with the ultimate goal of precision medicine informed by deep understanding of specific tumor responses to radiation and adjunct chemotherapy targeting key factors in the DDR and Phoenix Rising pathways.

Dataset Information

Acetylation dynamics of human nuclear proteins during the ionizing radiation-induced DNA damage response.

Publications

Acetylation dynamics of human nuclear proteins during the ionizing radiation-induced DNA damage response.

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