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

作者：Fujimori Hiroaki; Sato Akira; Kikuhara Sota; Wang Junhui; Hirai Takahisa; Sasaki Yuka; Murakami Yasufumi; Okayasu Ryuichi; Masutani Mitsuko^*

来源：Scientific Reports, 2015, 5(1): 18231.

DOI：10.1038/srep18231

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 gamma-irradiation and carbon-ion beam irradiation. DNMT3B RNAi reduced the activation of DNA damage responses induced by gamma-irradiation, including HP1 beta-,gamma H2AX-and Rad51-foci formation. DNMT3B RNAi impaired damage-dependent H2AX accumulation and showed a reduced level of gamma H2AX induction after gamma-irradiation. DNMT3B interacted with HP1 beta in non-irradiated conditions, whereas irradiation abrogated the DNMT3B/HP1 beta complex but induced interaction between DNMT3B and H2AX. Consistent with radiosensitization, TP63, BAX, PUMA and NOXA expression was induced after gamma-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 HP1 beta foci formation and efficient gamma H2AX-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.