title: Suppression of DNA-PKcs and Ku80 individually and in combination: Different effects of radiobiology in HeLa cells.
abstract: DNA-dependent protein kinase (DNA-PK), including Ku80, Ku70 and DNA-PK catalytic subunit (DNA-PKcs), is the key protein in non-homologous end-joining (NHEJ) after DNA double-strand breaks (DSBs) appear. In this study, small hairpin interfering RNAs (siRNAs) targeting Ku80 and DNA- PKcs were used both individually and in combination, to explore the effects of these DSB proteins on HeLa cell functional changes after X-ray irradiation. HeLa cells co-transfected with Ku80-siRNA and DNA-PKcs-siRNA were more radiosensitive than the ones transfected individually. HeLa in the absence of Ku80 and pretreated with LY294002, a chemically specific PI 3-kinase inhibitor, resulted in cells that were even more sensitive to X-rays than HeLa/Ku80-siRNA transfected with DNA- PKcs-siRNA. The cells inhibited by Ku80 either individually or in combination with DNA-PKcs showed cell accumulation in the G2/M phase 48 h post-irradiation, similarly to control cells. However, cells transfected with DNA-PKcs-siRNA or pretreated with LY294002 had a prolonged G2/M delay, suggesting the accumulation of significant un-repaired DNA damage following inhibition of DSB repair proteins. In conclusion, these data indicate that the role of Ku80 in DSB repair could be compensated by other DSB repair proteins; co-inhibition would be a suitable strategy to enhance the radiosensitivity of cancer cells.
authors.affiliation: Cancer Center, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan Hubei 430030, PR China.
authors.forename: Liang
authors.initials: L
authors.lastname: Zhuang
authors.forename: Yang
authors.initials: Y
authors.lastname: Cao
authors.forename: Huihua
authors.initials: H
authors.lastname: Xiong
authors.forename: Qinglei
authors.initials: Q
authors.lastname: Gao
authors.forename: Zhe
authors.initials: Z
authors.lastname: Cao
authors.forename: Fei
authors.initials: F
authors.lastname: Liu
authors.forename: Hong
authors.initials: H
authors.lastname: Qiu
authors.forename: Shiying
authors.initials: S
authors.lastname: Yu
authors.forename: Xiaoyuan
authors.initials: X
authors.lastname: Huang
mesh_terms: D034961:Antigens, Nuclear / Q000235:genetics / Q000378:metabolism*; D002453:Cell Cycle / Q000235:genetics; D045744:Cell Line, Tumor; D002867:Chromones / Q000494:pharmacology; D051747:DNA-Activated Protein Kinase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects / Q000528:radiation effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D020868:Gene Silencing; D006367:HeLa Cells; D006801:Humans; D000072200:Ku Autoantigen; D009025:Morpholines / Q000494:pharmacology; D009369:Neoplasms / Q000201:enzymology / Q000503:physiopathology; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D011836:Radiation Tolerance / Q000187:drug effects / Q000235:genetics; D014965:X-Rays
chemical_list: D034961:Antigens, Nuclear; D002867:Chromones; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D009025:Morpholines; D034741:RNA, Small Interfering; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D051747:DNA-Activated Protein Kinase; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen