Stimulation of Nuclear Factor (Erythroid-Derived 2)-like 2 Signaling by Nucleus Targeted Irradiation with Proton Microbeam

SIMPLE SUMMARY: With the advancement in microbeam cell irradiation technologies, researchers can now investigate subcellular region irradiation-induced biological responses as well as whether and how they contribute to the overall radiation induced biological effects of individual cells and the cell population. It will be of great value not only in expanding our understanding of the mechanisms of ionizing interaction with cells, but also in the studies of radiation protection and radiotherapy. Radiation-induced biological effects are commonly associated with an increase in reactive free radicals. Nuclear factor erythroid 2-related factor 2 (NRF2) is a master regulator of cellular oxidative stress. We investigated the response of NRF2 signaling to nucleus targeted irradiation using the Single Particle Irradiation System to Cell (SPICE-QST microbeam) facility and evaluated the similarities and differences between irradiating the cytoplasm or nucleus in the stimulation of NRF2 signaling. ABSTRACT: Nuclear factor (erythroid-derived 2)-like 2 (NRF2), well-known as a master antioxidative response regulator in mammalian cells, is considered as a potential target for radiation protection and cancer therapy sensitization. We examined the response of NRF2 signaling in normal human lung fibroblast WI-38 cells to nucleus targeted irradiation by 3.4 MeV proton microbeam. Nucleus targeted irradiation stimulated the nucleus accumulation of NRF2 and the expression of its target gene, heme oxygenase 1 (HO-1). The nucleus accumulation of NRF2 increased from 3 h to 12 h post 500 proton irradiation. In the 500 protons range, higher number of protons resulted in increased NRF2 nucleus accumulation. Activating NRF2 with tert-butylhydroquinone reduced DNA double-strand break (DSB) formation in nucleus targeted irradiation by 15%. Moreover, ATM phosphorylation was found in nucleus targeted irradiation. Inhibiting ATM with ku55933 prevented NRF2 nucleus accumulation. Furthermore, nucleus targeted irradiation activated ERK 1/2, and ROS-ERK 1/2 signaling regulated NRF2 nucleus accumulation. Taken together, NRF2 signaling was activated by nucleus targeted irradiation and mitigated DNA DSB. The discovery of ATM and ERK 1/2 as upstream regulators of NRF2 signaling in nucleus targeted cells revealed new information regarding radiation protection.