DNA double-strand breaks activate ATM independent of mitochondrial dysfunction in A549 cells |
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| Affiliation: | 1. Department of Environmental Medicine, The University of Rochester, Rochester, NY 14642, USA;2. Department of Anesthesiology, The University of Rochester, Rochester, NY 14642, USA;3. Department of Pediatrics, The University of Rochester, Rochester, NY 14642, USA;4. Department of Biology, The University of Rochester, Rochester, NY 14642, USA;1. Departments of Pharmacology, University of North Carolina School of Medicine, Chapel Hill, NC 27599, USA;2. Biochemistry and Biophysics, University of North Carolina School of Medicine, Chapel Hill, NC 27599, USA;3. Lineberger Comprehensive Cancer Center, University of North Carolina School of Medicine, Chapel Hill, NC 27599, USA;1. Department of Thoracic and Cardiovascular Surgery, University of Texas MD Anderson Cancer Center, 1515 Holcombe Blvd, Houston, TX, United States;2. Department of Bioinformatics and Computational Biology, University of Texas MD Anderson Cancer Center, Houston, TX, United States;3. Department of Thoracic/Head and Neck Medical Oncology, University of Texas MD Anderson Cancer Center, Houston, TX, United States;4. Department of Melanoma Medical Oncology, University of Texas MD Anderson Cancer Center, Houston, TX, United States;5. Department of Molecular and Cellular Oncology, University of Texas MD Anderson Cancer Center, Houston, TX, United States;6. Department of Translational Molecular Pathology, University of Texas MD Anderson Cancer Center, Houston, TX, United States;1. Department of Spine Surgery, Xiangya Hospital, Central South University, Hunan, China;2. Clinical Laboratory, Secondary Affiliated Hospital, Fujian Medical University, Fujian, China;1. Department of Radiology, University of Michigan Health System, Ann Arbor, MI, USA;2. Applied Physics Program, University of Michigan, Ann Arbor, MI, USA;3. Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, USA;4. School of Dentistry, University of Michigan, Ann Arbor, MI, USA;5. Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor, MI, USA |
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| Abstract: | Excessive nuclear or mitochondrial DNA damage can lead to mitochondrial dysfunction, decreased energy production, and increased generation of reactive oxygen species (ROS). Although numerous cell signaling pathways are activated when cells are injured, the ataxia telangiectasia mutant (ATM) protein has emerged as a major regulator of the response to both mitochondrial dysfunction and nuclear DNA double-strand breaks (DSBs). Because mitochondrial dysfunction is often a response to excessive DNA damage, it has been difficult to determine whether nuclear and/or mitochondrial DNA DSBs activate ATM independent of mitochondrial dysfunction. In this study, mitochondrial and nuclear DNA DSBs were generated in the A549 human lung adenocarcinoma cell line by infecting with retroviruses expressing the restriction endonuclease PstI fused to a mitochondrial targeting sequence (MTS) or nuclear localization sequence (NLS) and a hemagglutinin antigen epitope tag (HA). Expression of MTS-PstI-HA or NLS-PstI-HA activated the DNA damage response defined by phosphorylation of ATM, the tumor suppressor protein p53 (TP53), KRAB-associated protein (KAP)-1, and structural maintenance of chromosomes (SMC)-1. Phosphorylated ATM and SMC1 were detected in nuclear fractions, whereas phosphorylated TP53 and KAP1 were detected in both mitochondrial and nuclear fractions. PstI also enhanced expression of the cyclin-dependent kinase inhibitor p21 and inhibited cell growth. This response to DNA damage occurred in the absence of detectable mitochondrial dysfunction and excess production of ROS. These findings reveal that DNA DSBs are sufficient to activate ATM independent of mitochondrial dysfunction and suggest that the activated form of ATM and some of its substrates are restricted to the nuclear compartment, regardless of the site of DNA damage. |
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| Keywords: | ATM DNA damage Free radicals Mitochondrial dysfunction Oxidative stress |
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