Dynamic localization of human RAD18 during the cell cycle and a functional connection with DNA double-strand break repair |
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| Authors: | Akiko Inagaki Wiggert A. van Cappellen Roald van der Laan Adriaan B. Houtsmuller Jan H.J. Hoeijmakers J. Anton Grootegoed Willy M. Baarends |
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| Affiliation: | 1. Department of Reproduction and Development, Erasmus MC, University Medical Center, PO Box 2040, 3000 CA, Rotterdam, The Netherlands;2. Department of Cell Biology and Genetics, Erasmus MC, University Medical Center, Rotterdam, The Netherlands;3. Department of Pathology, Josephine Nefkens Institute Erasmus MC, University Medical Center, Rotterdam, The Netherlands;2. Internal Medicine Department, Texas Tech University Health Sciences Center of El Paso, El Paso, Texas;3. Department of Psychiatry, University of Missouri School of Medicine, Kansas City, Missouri;1. Department of Pharmacological Sciences, Stony Brook University, Stony Brook, NY 11794, United States;2. Department of Chemistry, Stony Brook University, Stony Brook, NY 11794, United States;3. Department of Medicine, Stony Brook University, Stony Brook, NY 11794, United States;1. College of Life Sciences, Capital Normal University, Beijing, 100048, China;2. Department of Microbiology and Immunology, University of Saskatchewan, Saskatoon, SK, S7N 5E5, Canada;1. Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, NY 10016, USA;2. Perlmutter NYU Cancer Center, New York University School of Medicine, New York, NY 10016, USA;3. Department of Biochemistry and Structural Biology, UT Health San Antonio, San Antonio, TX 78229, USA;4. Department of Pathology, New York University School of Medicine, New York, NY 10016, USA;5. Howard Hughes Medical Institute, New York University School of Medicine, New York, NY 10016, USA;1. The Wellcome Trust and Cancer Research UK Gurdon Institute, University of Cambridge, Cambridge 2 1QN, UK;2. Institut de Pharmacologie et de Biologie Structurale, CNRS, Université de Toulouse-Université Paul Sabatier, Equipe Labellisée Ligue contre le Cancer, 31077 Toulouse, France;3. Institute of Molecular Biology (IMB), 55128 Mainz, Germany |
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| Abstract: | The ubiquitin ligase RAD18 is involved in different DNA repair processes. Here, we show that in G1 phase, human RAD18 accumulates in a few relatively large spontaneous foci that contain proteins involved in double-strand break (DSB) repair. These foci persist until cells enter S phase, when numerous small foci appear. At these sites, only 20% of RAD18 colocalizes with PCNA, a known RAD18 substrate. In late G2 phase, RAD18 relocates to nucleoli. After UVC irradiation, PCNA accumulates at the damaged site, followed by RAD18, independent of the cell cycle phase. After induction of DSBs, using low-power multi-photon laser, RAD18 accumulated at the DSB sites, but no PCNA accumulation was observed. Our data show that RAD18 accumulates on DSBs independent of the cell cycle phase. DSBs marked by RAD18 and RAD51 are also positive for RPA in G1 phase, and these DSBs persist until S phase. In addition, we show that DSBs generated in G2 phase are not all repaired, and are observed again in the next G1 phase. We conclude that repair of induced and spontaneous DSBs that accumulate RAD18 and RAD51 in G1 phase cells is delayed until S phase. |
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