Studies on the ability of multicellular organisms to tolerate specific environmental extremes are relatively rare compared to those of unicellular microorganisms in extreme environments. Tardigrades are extremotolerant animals that can enter an ametabolic dry state called anhydrobiosis and have high tolerance to a variety of extreme environmental conditions, particularly while in anhydrobiosis. Although tardigrades have been expected to be a potential model animal for astrobiological studies due to their excellent anhydrobiotic and extremotolerant abilities, few studies of tolerance with cultured tardigrades have been reported, possibly due to the absence of a model species that can be easily maintained under rearing conditions. We report the successful rearing of the herbivorous tardigrade, Ramazzottius varieornatus, by supplying the green alga Chlorella vulgaris as food. The life span was 35 +/- 16.4 d, deposited eggs required 5.7 +/- 1.1 d to hatch, and animals began to deposit eggs 9 d after hatching. The reared individuals of this species had an anhydrobiotic capacity throughout their life cycle in egg, juvenile, and adult stages. Furthermore, the reared adults in an anhydrobiotic state were tolerant of temperatures of 90 degrees C and -196 degrees C, and exposure to 99.8% acetonitrile or irradiation with 4000 Gy (4)He ions. Based on their life history traits and tolerance to extreme stresses, R. varieornatus may be a suitable model for astrobiological studies of multicellular organisms.

SummaryThe extraordinary radiation resistance of Deinococcus radiodurans results from the efficient capacity of the bacterium to repair DNA double-strand breaks. By analysing the DNA damage repair-deficient mutant, KH311, a unique radiation-inducible gene (designated pprA ) responsible for loss of radiation resistance was identified. Investigations in vitro showed that the gene product of pprA (PprA) preferentially bound to double-stranded DNA carrying strand breaks, inhibited Escherichia coli exonuclease III activity, and stimulated the DNA end-joining reaction catalysed by ATPdependent and NAD-dependent DNA ligases. These results suggest that D. radiodurans has a radiationinduced non-homologous end-joining repair mechanism in which PprA plays a critical role.

M. tardigradum survives high doses of ionizing radiation in both hydrated and anhydrobiotic states, but irradiation with >1000 Gy makes them sterile.

The involvement of LexA in induction of RecA was investigated in Deinococcus radiodurans. As in the wild-type strain, an increase in RecA protein synthesis following ␥ irradiation was detected in a lexA disruptant, indicating that LexA is not involved in the induction of RecA in D. radiodurans.Deinococcus radiodurans is characterized by its extraordinary radiation resistance phenotype, which is considered to be due to a highly proficient DNA repair capacity (3,25,34). The most striking feature of D. radiodurans is that it can mend over 100 double-strand breaks (DSBs) of genomic DNA during postirradiation incubation (2, 18). As the rejoining of DSBs can be prevented by adding chloramphenicol to the incubation mixture, proteins induced by irradiation are necessary for the rejoining of DNA breakages (18). Several DNA damage-inducible proteins that may be required for DNA repair have been detected in the cell extract of D. radiodurans by twodimensional polyacrylamide gel electrophoresis (PAGE) (12,35). These observations suggest that D. radiodurans possesses a DNA damage response mechanism. However, little is known about the molecular basis for the control of the inducible proteins.In Escherichia coli, the inducible DNA repair system (the SOS system) is regulated by two key proteins; RecA and LexA (8,38). E. coli RecA is activated by DNA damage to mediate the proteolytic cleavage of the E. coli LexA repressor, resulting in derepression of the SOS regulon. The SOS response in Bacillus subtilis progresses in a similar manner, with B. subtilis RecA having an identical role in controlling the SOS regulon together with a cellular repressor protein that is functionally homologous to the E. coli LexA repressor (42). The B. subtilis repressor (termed DinR) binds the promoter regions of several din genes and B. subtilis recA (20,23,40,41) and undergoes autodigestion under alkaline conditions and RecA-mediated cleavage under more physiological conditions (23, 41). It has also been shown that the intracellular level of intact DinR is significantly reduced following DNA damage (23). Thus, the basic mechanism of the SOS response seems to be conserved between E. coli and B. subtilis. Deinococcus species form a coherent phylogenetic cluster related to the Thermus-Meiothermus lineage (30), indicating that the Deinococcus lineage is distinct from the lineages of both proteobacteria and grampositive bacteria. Although SOS-like processes have been documented in a wide variety of eubacterial species (24, 32), the involvement of RecA and LexA in the SOS response is poorly understood in Deinococcus and closely related bacterial species.As expression of the deinococcal recA gene is enhanced after ␥ irradiation (4), the recA gene seems to be a member of a DNA damage response regulon in D. radiodurans. In the present study, D. radiodurans LexA was purified from E. coli cells and its ability to cleave itself was examined. The changes in intracellular levels of the LexA and RecA proteins following ␥ irradiation were also investigated by usin...

Space travel has advanced significantly over the last six decades with astronauts spending up to 6 months at the International Space Station. Nonetheless, the living environment while in outer space is extremely challenging to astronauts. In particular, exposure to space radiation represents a serious potential long-term threat to the health of astronauts because the amount of radiation exposure accumulates during their time in space. Therefore, health risks associated with exposure to space radiation are an important topic in space travel, and characterizing space radiation in detail is essential for improving the safety of space missions. In the first part of this review, we provide an overview of the space radiation environment and briefly present current and future endeavors that monitor different space radiation environments. We then present research evaluating adverse biological effects caused by exposure to various space radiation environments and how these can be reduced. We especially consider the deleterious effects on cellular DNA and how cells activate DNA repair mechanisms. The latest technologies being developed, e.g., a fluorescent ubiquitination-based cell cycle indicator, to measure real-time cell cycle progression and DNA damage caused by exposure to ultraviolet radiation are presented. Progress in examining the combined effects of microgravity and radiation to animals and plants are summarized, and our current understanding of the relationship between psychological stress and radiation is presented. Finally, we provide details about protective agents and the study of organisms that are highly resistant to radiation and how their biological mechanisms may aid developing novel technologies that alleviate biological damage caused by radiation. Future research that furthers our understanding of the effects of space radiation on human health will facilitate risk-mitigating strategies to enable long-term space and planetary exploration.

The possible mechanism of a radiation-induced bystander response was investigated by using a high-LET heavy particle microbeam, which allows selected cells to be individually hit with precise numbered particles. Even when only a single cell within the confluent culture was hit by one particle of 40Ar (approximately 1260 keV/microm) or 20Ne (approximately 380 keV/microm), a 1.4-fold increase of micronuclei (MN) was detected demonstrating a bystander response. When the number of targeted cells increased, the number of MN biphasically increased; however, the efficiency of MN induction per targeted cell markedly decreased. When 49 cells in the culture were individually hit by 1 to 4 particles, the production of MN in the irradiated cultures were approximately 2-fold higher than control levels but independent of the number and LET of the particles. MN induction in the irradiated-culture was partly reduced by treatment with DMSO, a scavenger of reactive oxygen species (ROS), and was almost fully suppressed by the mixture of DMSO and PMA, an inhibitor of gap junctional intercellular communication (GJIC). Accordingly, both ROS and GJIC contribute to the above-mentioned bystander response and GJIC may play an essential role by mediating the release of soluble biochemical factors from targeted cells.

Evidence has accumulated showing that ionizing radiations persistently perturb genomic stability and induce delayed reproductive death in the progeny of survivors; however, the linear energy transfer (LET) dependence of these inductions has not been fully characterized. We have investigated the cell killing effectiveness of gamma rays (0.2 keV/microm) and six different beams of heavy-ion particles with LETs ranging from 16.2 to 1610 keV/microm in normal human fibroblasts. First, irradiated confluent density-inhibited cultures were plated for primary colony formation, revealing that the relative biological effectiveness (RBE) based on the primary 10% survival dose peaked at 108 keV/microm and that the inactivation cross section increased proportionally up to 437 keV/microm. Second, cells harvested from primary colonies were plated for secondary colony formation, showing that delayed reproductive death occurred in a dose-dependent fashion. While the RBE based on the secondary 80% survival dose peaked at 108 keV/microm, very little difference in LET was observed in the RBE based on secondary survival at the primary 10% survival dose. Our present results indicate that delayed reproductive death arising only during secondary colony formation is independent of LET and is more likely to be dependent on initial damages having been fixed during primary colony formation.

DNA double-strand breaks (DSBs) induced by ionizing radiation pose a major threat to cell survival. The cell can respond to the presence of DSBs through two major repair pathways: homologous recombination (HR) and nonhomologous end joining (NHEJ). Higher levels of cell death are induced by high-linear energy transfer (LET) radiation when compared to low-LET radiation, even at the same physical doses, due to less effective and efficient DNA repair. To clarify whether high-LET radiation inhibits all repair pathways or specifically one repair pathway, studies were designed to examine the effects of radiation with different LET values on DNA DSB repair and radiosensitivity. Embryonic fibroblasts bearing repair gene (NHEJ-related Lig4 and/or HR-related Rad54) knockouts (KO) were used and their responses were compared to wild-type cells. The cells were exposed to X rays, spread-out Bragg peak (SOBP) carbon ion beams as well as with carbon, iron, neon and argon ions. Cell survival was measured with colony-forming assays. The sensitization enhancement ratio (SER) values were calculated using the 10% survival dose of wild-type cells and repair-deficient cells. Cellular radiosensitivity was listed in descending order: double-KO cells > Lig4-KO cells > Rad54-KO cells > wild-type cells. Although Rad54-KO cells had an almost constant SER value, Lig4-KO cells showed a high-SER value when compared to Rad54-KO cells, even with increasing LET values. These results suggest that with carbon-ion therapy, targeting NHEJ repair yields higher radiosensitivity than targeting homologous recombination repair.