title: Suppression of DNA-dependent protein kinase sensitize cells to radiation without affecting DSB repair.
abstract: Efficient and correct repair of DNA double-strand break (DSB) is critical for cell survival. Defects in the DNA repair may lead to cell death, genomic instability and development of cancer. The catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) is an essential component of the non-homologous end joining (NHEJ) which is the major DSB repair pathway in mammalian cells. In the present study, by using siRNA against DNA-PKcs in four human cell lines, we examined how low levels of DNA-PKcs affected cellular response to ionizing radiation. Decrease of DNA-PKcs levels by 80-95%, induced by siRNA treatment, lead to extreme radiosensitivity, similar to that seen in cells completely lacking DNA-PKcs and low levels of DNA-PKcs promoted cell accumulation in G2/M phase after irradiation and blocked progression of mitosis. Surprisingly, low levels of DNA-PKcs did not affect the repair capacity and the removal of 53BP1 or γ-H2AX foci and rejoining of DSB appeared normal. This was in strong contrast to cells completely lacking DNA-PKcs and cells treated with the DNA-PKcs inhibitor NU7441, in which DSB repair were severely compromised. This suggests that there are different mechanisms by which loss of DNA-PKcs functions can sensitize cells to ionizing radiation. Further, foci of phosphorylated DNA-PKcs (T2609 and S2056) co-localized with DSB and this was independent of the amount of DNA-PKcs but foci of DNA-PKcs was only seen in siRNA-treated cells. Our study emphasizes on the critical role of DNA-PKcs for maintaining survival after radiation exposure which is uncoupled from its essential function in DSB repair. This could have implications for the development of therapeutic strategies aiming to radiosensitize tumors by affecting the DNA-PKcs function.
authors.affiliation: Section of Biomedical Radiation Sciences, Department of Radiology, Oncology and Radiation Science, Rudbeck Laboratory, Uppsala University, Dag Hammarskjölds Väg 20, SE-751 85 Uppsala, Sweden. Electronic address: ann-sofie.gustafsson@bms.uu.se.
authors.forename: Ann-Sofie
authors.initials: AS
authors.lastname: Gustafsson
authors.affiliation: Section of Biomedical Radiation Sciences, Department of Radiology, Oncology and Radiation Science, Rudbeck Laboratory, Uppsala University, Dag Hammarskjölds Väg 20, SE-751 85 Uppsala, Sweden.
authors.forename: Andris
authors.initials: A
authors.lastname: Abramenkovs
authors.affiliation: Section of Biomedical Radiation Sciences, Department of Radiology, Oncology and Radiation Science, Rudbeck Laboratory, Uppsala University, Dag Hammarskjölds Väg 20, SE-751 85 Uppsala, Sweden.
authors.forename: Bo
authors.initials: B
authors.lastname: Stenerlöw
mesh_terms: D002470:Cell Survival / Q000187:drug effects / Q000528:radiation effects; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology*; D053903:DNA Breaks, Double-Stranded / Q000187:drug effects*; D004260:DNA Repair / Q000187:drug effects* / Q000235:genetics; D051747:DNA-Activated Protein Kinase / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D045325:HCT116 Cells; D006801:Humans; D008938:Mitosis / Q000187:drug effects / Q000235:genetics / Q000528:radiation effects; D009025:Morpholines / Q000494:pharmacology*; D034741:RNA, Small Interfering / Q000494:pharmacology; D011836:Radiation Tolerance / Q000187:drug effects* / Q000235:genetics; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology
chemical_list: C499693:8-dibenzothiophen-4-yl-2-morpholin-4-yl-chromen-4-one; D002867:Chromones; D004791:Enzyme Inhibitors; D009025:Morpholines; D034741:RNA, Small Interfering; D011838:Radiation-Sensitizing Agents; D051747:DNA-Activated Protein Kinase
keywords: DNA repair; DNA-PK deficiency; DNA-PKcs; Ionizing radiation; NU7441