کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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1928938 | 1536780 | 2012 | 6 صفحه PDF | دانلود رایگان |
Nonhomologous end joining (NHEJ) and homologous recombination (HR) are known as DNA double-strand break (DSB) repair pathways. It has been reported that DNA-PK, a member of PI3 kinase family, promotes NHEJ and aberrant DNA-PK causes NHEJ deficiency. However, in this study, we demonstrate that a wild-type cell line treated with DNA-PK inhibitor and a mutant cell line with dysfunctional DNA-PK showed decreased HR efficiency in fish cells (Medaka, Oryzias latipes). Previously, we reported that the radiation-sensitive mutant RIC1 strain has a defect in the Histone H2AX phosphorylation after γ-irradiation. Here, we showed that a DNA-PK inhibitor, NU7026, treatment resulted in significant reduction in the number of γH2AX foci after γ-irradiation in wild-type cells, but had no significant effect in RIC1 cells. In addition, RIC1 cells showed significantly lower levels of DNA-PK kinase activity compared with wild-type cells. We investigated NHEJ and HR efficiency after induction of DSBs. Wild-type cells treated with NU7026 and RIC1 cells showed decreased HR efficiency. These results indicated that aberrant DNA-PK causes the reduction in the number of γH2AX foci and HR efficiency in RIC1 cells. We performed phosphorylated DNA-PKcs (Thr2609) and 53BP1 focus assay after γ-irradiation. RIC1 cells showed significant reduction in the number of phosphorylated DNA-PKcs foci and no deference in the number of 53BP1 foci compared with wild-type cells. These results suggest that low level of DNA-PK activity causes aberrant DNA-PKcs autophosphorylation in RIC1 cells. It is known that 53BP1 is involved in both DNA-PK dependent and independent NHEJ. Therefore we suggest that DNA-PK independent NHEJ repair DSBs under the condition of decreased DNA-PK activity, which causes reduction of HR efficiency.
► We investigated the effect of DNA-PK inhibition on DSB repair using fish cells.
► A radiation sensitive mutant RIC1 strain showed a low level of DNA-PK activity.
► DNA-PK dysfunction leads defects in HR repair and DNA-PKcs autophosphorylation.
► DNA-PK dysfunction leads a slight increase in the number of 53BP1 foci after DSBs.
► DNA-PK dysfunction leads an alternative NHEJ that depends on 53BP1.
Journal: Biochemical and Biophysical Research Communications - Volume 429, Issues 3–4, 14 December 2012, Pages 131–136