A comprehensive analysis of radiosensitization targets; functional inhibition of DNA methyltransferase 3B radiosensitizes by disrupting DNA damage regulation

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A comprehensive genome-wide screen of radiosensitization targets in HeLa cells was performed using a shRNA-library/functional cluster analysis and DNMT3B was identified as a candidate target. DNMT3B RNAi increased the sensitivity of HeLa, A549 and HCT116 cells to both γ3-irradiation and carbon-ion beam irradiation. DNMT3B RNAi reduced the activation of DNA damage responses induced by γ3-irradiation, including HP1β-, γ3H2AX- and Rad51-foci formation. DNMT3B RNAi impaired damage-dependent H2AX accumulation and showed a reduced level of γ3H2AX induction after γ3-irradiation. DNMT3B interacted with HP1β in non-irradiated conditions, whereas irradiation abrogated the DNMT3B/HP1β complex but induced interaction between DNMT3B and H2AX. Consistent with radiosensitization, TP63, BAX, PUMA and NOXA expression was induced after γ3-irradiation in DNMT3B knockdown cells. Together with the observation that H2AX overexpression canceled radiosensitization by DNMT3B RNAi, these results suggest that DNMT3B RNAi induced radiosensitization through impairment of damage-dependent HP1β foci formation and efficient γ3H2AX-induction mechanisms including H2AX accumulation. Enhanced radiosensitivity by DNMT3B RNAi was also observed in a tumor xenograft model. Taken together, the current study implies that comprehensive screening accompanied by a cluster analysis enabled the identification of radiosensitization targets. Downregulation of DNMT3B, one of the targets identified using this method, radiosensitizes cancer cells by disturbing multiple DNA damage responses.

Scientific Reports, 5, 18231; 2015

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