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The role of DNA repair by nonhomologous end joining (NHEJ), homologous recombination, spore photoproduct lyase, and DNA polymerase I and genome protection via ␣/␤-type small, acid-soluble spore proteins (SASP) in Bacillus subtilis spore resistance to accelerated heavy ions (high-energy charged [HZE] particles) and X rays has been studied. Spores deficient in NHEJ and ␣/␤-type SASP were significantly more sensitive to HZE particle bombardment and X-ray irradiation than were the recA, polA, and splB mutant and wild-type spores, indicating that NHEJ provides an efficient DNA double-strand break repair pathway during spore germination and that the loss of the ␣/␤-type SASP leads to a significant radiosensitivity to ionizing radiation, suggesting the essential function of these spore proteins as protectants of spore DNA against ionizing radiation.Endospores of the gram-positive bacterium Bacillus subtilis are highly resistant to inactivation by environmental stresses, such as biocidal agents and toxic chemicals, desiccation, pressure and temperature extremes, and high fluences of UV radiation (reviewed in references 44, 45, and 61) and are a powerful biodosimetric system for terrestrial environmental monitoring and astrobiological studies (44). On Earth, understanding extreme spore resistance to ionizing radiation is important in the areas of food preservation, medical sterilization, and decontamination from bioterror attack (4, 17, 47; reviewed in references 42 and 43). Off Earth, spore radiation resistance is important both in space flight and in ground-based simulations, in order to obtain information on the biological damage produced by exposure to space conditions (23,25,26,44). Onboard several spacecraft (Apollo 16, Spacelab 1, LDEF, D2, and FOTON), spores of Bacillus subtilis were exposed to selected parameters of space, such as space vacuum and different spectral ranges of solar UV radiation and cosmic rays, applied separately or in combination (5, 9, 19-21, 23, 24, 26). Especially, the radiation environment on Earth, on Mars, in low-Earth orbit, and in deep space is typified by a wide variety of primary particles covering an extended range of energies. Galactic cosmic rays (GCR) are charged particles that originate from sources beyond our solar system. The distribution of GCR is believed to be isotropic throughout interstellar space. The spectrum of the GCR consists of 98% protons and heavier ions (baryon component) and 2% electrons and positrons (lepton component). The baryon component is composed of 87% protons, 12% helium ions (alpha particles), and the remaining 1% heavy ions of charge 3 from lithium to 92 from uranium. Due to their high abundance, iron ions are highly penetrating, giving them a large potential for radiobiological damage (3,21).While the UV photochemistry of spore DNA and repair of UV damage to DNA during germination are well characterized (34,44,(59)(60)(61), there has been relatively little work on the nature of DNA damage in spores caused by ionizing radiation, the protective role of...

Human and rodent cells proficient and deficient in non-homologous end joining (NHEJ) were irradiated with X rays, 70 keV/microm carbon ions, and 200 keV/microm iron ions, and the biological effects on these cells were compared. For wild-type CHO and normal human fibroblast (HFL III) cells, exposure to iron ions yielded the lowest cell survival, followed by carbon ions and then X rays. NHEJ-deficient xrs6 (a Ku80 mutant of CHO) and 180BR human fibroblast (DNA ligase IV mutant) cells showed similar cell survival for X and carbon-ion irradiation (RBE = approximately 1.0). This phenotype is likely to result from a defective NHEJ protein because xrs6-hamKu80 cells (xrs6 cells corrected with the wild-type KU80 gene) exhibited the wild-type response. At doses higher than 1 Gy, NHEJ-defective cells showed a lower level of survival with iron ions than with carbon ions or X rays, possibly due to inactivation of a radioresistant subpopulation. The G(1) premature chromosome condensation (PCC) assay with HFL III cells revealed LET-dependent impairment of repair of chromosome breaks. Additionally, iron-ion radiation induced non-repairable chromosome breaks not observed with carbon ions or X rays. PCC studies with 180BR cells indicated that the repair kinetics after exposure to carbon and iron ions behaved similarly for the first 6 h, but after 24 h the curve for carbon ions approached that for X rays, while the curve for iron ions remained high. These chromosome data reflect the existence of a slow NHEJ repair phase and severe biological damage induced by iron ions. The auto-phosphorylation of DNA-dependent protein kinase catalytic subunits (DNA-PKcs), an essential NHEJ step, was delayed significantly by high-LET carbon- and iron-ion radiation compared to X rays. This delay was further emphasized in NHEJ-defective 180BR cells. Our results indicate that high-LET radiation induces complex DNA damage that is not easily repaired or is not repaired by NHEJ even at low radiation doses such as 2 Gy.

The biological effects of radiation originate principally in damages to DNA. DNA damages by X rays as well as heavy ions are induced by a combination of direct and indirect actions. The contribution of indirect action in cell killing can be estimated from the maximum degree of protection by dimethylsulfoxide (DMSO), which suppresses indirect action without affecting direct action. Exponentially growing Chinese hamster V79 cells were exposed to high-LET radiations of 20 to 2106 keV/mum in the presence or absence of DMSO and their survival was determined using a colony formation assay. The contribution of indirect action to cell killing decreased with increasing LET. However, the contribution did not reach zero even at very high LETs and was estimated to be 32% at an LET of 2106 keV/mum. Therefore, even though the radiochemically estimated G value of OH radicals was nearly zero at an LET of 1000 keV/mum, indirect action by OH radicals contributed to a substantial fraction of the biological effects of high-LET radiations. The RBE determined at a survival level of 10% increased with LET, reaching a maximum value of 2.88 at 200 keV/mum, and decreased thereafter. When the RBE was estimated separately for direct action (RBE(D)) and indirect action (RBE(I)); both exhibited an LET dependence similar to that of the RBE, peaking at 200 keV/mum. However, the peak value was much higher for RBE(D) (5.99) than RBE(I) (1.89). Thus direct action contributes more to the high RBE of high-LET radiations than indirect action does.

Although carbon ion therapy facilities are expensive, the biological effects of carbon ion beam treatment may be better against cancer (and cancer