title: DNA repair kinetics in SCID mice Sertoli cells and DNA-PKcs-deficient mouse embryonic fibroblasts.
abstract: Noncycling and terminally differentiated (TD) cells display differences in radiosensitivity and DNA damage response. Unlike other TD cells, Sertoli cells express a mixture of proliferation inducers and inhibitors in vivo and can reenter the cell cycle. Being in a G1-like cell cycle stage, TD Sertoli cells are expected to repair DSBs by the error-prone nonhomologous end-joining pathway (NHEJ). Recently, we have provided evidence for the involvement of Ku-dependent NHEJ in protecting testis cells from DNA damage as indicated by persistent foci of the DNA double-strand break (DSB) repair proteins phospho-H2AX, 53BP1, and phospho-ATM in TD Sertoli cells of Ku70-deficient mice. Here, we analyzed the kinetics of 53BP1 foci induction and decay up to 12 h after 0.5 Gy gamma irradiation in DNA-PKcs-deficient (Prkdcscid) and wild-type Sertoli cells. In nonirradiated mice and PrkdcscidSertoli cells displayed persistent DSBs foci in around 12 % of cells and a fivefold increase in numbers of these DSB DNA damage-related foci relative to the wild type. In irradiated mice, PrkdcscidSertoli cells showed elevated levels of DSB-indicating foci in 82 % of cells 12 h after ionizing radiation (IR) exposure, relative to 52 % of irradiated wild-type Sertoli cells. These data indicate that Sertoli cells respond to and repair IR-induced DSBs in vivo, with repair kinetics being slow in the wild type and inefficient in Prkdcscid. Applying the same dose of IR to Prdkc-/-and Ku-/-mouse embryonic fibroblast (MEF) cells revealed a delayed induction of 53BP1 DSB-indicating foci 5 min post-IR in Prdkc-/-cells. Inefficient DSB repair was evident 7 h post-IR in DNA-PKcs-deficient cells, but not in Ku-/-MEFs. Our data show that quiescent Sertoli cells repair genotoxic DSBs by DNA-PKcs-dependent NEHJ in vivo with a slower kinetics relative to somatic DNA-PKcs-deficient cells in vitro, while DNA-PKcs deficiency caused inefficient DSB repair at later time points post-IR in both conditions. These observations suggest that DNA-PKcs contributes to the fast and slow repair of DSBs by NHEJ.
authors.affiliation: Laboratory of Immunology and Molecular Physiology, Zoology Department, Faculty of Science, Assiut University, Assiut, Egypt. emad20us@yahoo.com.|Department of Molecular Biology and Biotechnology, University of Sheffield, Sheffield, UK. emad20us@yahoo.com.
authors.forename: Emad A
authors.initials: EA
authors.lastname: Ahmed
authors.affiliation: Institut für Radiobiologie der Bundeswehr in Verb. mit der Universitat Ulm, Neuherbergstr 11, D-80937, Munich, Germany.|Department of Histology, University of Navarra, Pamplona, Spain.
authors.forename: Eukene
authors.initials: E
authors.lastname: Vélaz
authors.affiliation: Institute of Radiation Biology, Helmholtz Zentrum München, Neuherberg, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Rosemann
authors.affiliation: Institut für Radiobiologie der Bundeswehr in Verb. mit der Universitat Ulm, Neuherbergstr 11, D-80937, Munich, Germany.
authors.forename: Klaus-P
authors.initials: KP
authors.lastname: Gilbertz
authors.affiliation: Institut für Radiobiologie der Bundeswehr in Verb. mit der Universitat Ulm, Neuherbergstr 11, D-80937, Munich, Germany. scherth@web.de.
authors.forename: Harry
authors.initials: H
authors.lastname: Scherthan
mesh_terms: D000818:Animals; D002453:Cell Cycle / Q000235:genetics; D002460:Cell Line; D053903:DNA Breaks, Double-Stranded / Q000528:radiation effects; D004249:DNA Damage; D059766:DNA End-Joining Repair; D004260:DNA Repair; D051747:DNA-Activated Protein Kinase / Q000172:deficiency*; D004268:DNA-Binding Proteins / Q000172:deficiency*; D005347:Fibroblasts / Q000378:metabolism*; D055786:Gene Knockout Techniques; D007700:Kinetics; D008297:Male; D051379:Mice; D016513:Mice, SCID; D009687:Nuclear Proteins / Q000172:deficiency*; D011839:Radiation, Ionizing; D012708:Sertoli Cells / Q000378:metabolism* / Q000528:radiation effects; D016615:Telomere / Q000235:genetics / Q000378:metabolism
chemical_list: D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D051747:DNA-Activated Protein Kinase; C497264:Prkdc protein, mouse
keywords: 53BP1; DNA-PKcs; DSB repair; Ku70; NHEJ; Sertoli cells