Poaceae plants transfer cyclobutane pyrimidine dimer photolyase to chloroplasts for ultraviolet-B resistance.

Photoreactivation enzyme that repairs cyclobutane pyrimidine dimer (CPD) induced by ultraviolet-B radiation, commonly called CPD photolyase (PHR) is essential for plants living under sunlight. Rice (Oryza sativa) PHR (OsPHR) is a unique triple-targeting protein. The signal sequences required for its translocation to the nucleus or mitochondria are located in the C-terminal region but have yet to be identified for chloroplasts. Here, we identified sequences located in the N-terminal region, including the serine-phosphorylation site at position 7 of OsPHR, and found that OsPHR is transported/localized to chloroplasts via a vesicle transport system under the control of serine-phosphorylation. However, the sequence identified in this study is only conserved in some Poaceae species, and in many other plants, PHR is not localized to the chloroplasts. Therefore, we reasoned that Poaceae species need the ability to repair CPD in the chloroplast genome to survive under sunlight and have uniquely acquired this mechanism for PHR chloroplast translocation.

High ultraviolet-B sensitivity due to lower CPD photolyase activity is needed for biotic stress response to the rice blast fungus, Magnaporthe oryzae.

Sensitivity to ultraviolet-B (UVB, 280-315 nm) radiation varies widely among rice (Oryza sativa) cultivars due to differences in the activity of cyclobutane pyrimidines dimer (CPD) photolyase. Interestingly, cultivars with high UVB sensitivity and low CPD photolyase activity have been domesticated in tropical areas with high UVB radiation. Here, we investigated how differences in CPD photolyase activity affect plant resistance to the rice blast fungus, Magnaporthe oryzae, which is one of the other major stresses. We used Asian and African rice cultivars and transgenic lines with different CPD photolyase activities to evaluate the interaction effects of CPD photolyase activity on resistance to M. oryzae. In UVB-resistant rice plants overexpressing CPD photolyase, 12 h of low-dose UVB (0.4 W m-2) pretreatment enhanced sensitivity to M. oryzae. In contrast, UVB-sensitive rice (transgenic rice with antisense CPD photolyase, A-S; and rice cultivars with low CPD photolyase activity) showed resistance to M. oryzae. Several defense-related genes were upregulated in UVB-sensitive rice compared to UVB-resistant rice. UVB-pretreated A-S plants showed decreased multicellular infection and robust accumulation of reactive oxygen species. High UVB-induced CPD accumulation promoted defense responses and cross-protection mechanisms against rice blast disease. This may indicate a trade-off between high UVB sensitivity and biotic stress tolerance in tropical rice cultivars.

222 nm far-UVC efficiently introduces nerve damage in Caenorhabditis elegans.

Far-ultraviolet radiation C light (far-UVC; 222 nm wavelength) has received attention as a safer light for killing pathogenic bacteria and viruses, as no or little DNA damage is observed after irradiation in mammalian skin models. Far-UVC does not penetrate deeply into tissues; therefore, it cannot reach the underlying critical basal cells. However, it was unclear whether far-UVC (222-UVC) irradiation could cause more biological damage at shallower depths than the 254 nm UVC irradiation (254-UVC), which penetrates more deeply. This study investigated the biological effects of 222- and 254-UVC on the small and transparent model organism Caenorhabditis elegans. At the same energy level of irradiation, 222-UVC introduced slightly less cyclobutane pyrimidine dimer damage to naked DNA in solution than 254-UVC. The survival of eggs laid during 0-4 h after irradiation showed a marked decrease with 254-UVC but not 222-UVC. In addition, defect of chromosomal condensation was observed in a full-grown oocyte by 254-UVC irradiation. In contrast, 222-UVC had a significant effect on the loss of motility of C. elegans. The sensory nervous system, which includes dopamine CEP and PVD neurons on the body surface, was severely damaged by 222-UVC, but not by the same dose of 254-UVC. Interestingly, increasing 254-UVC irradiation by about 10-fold causes similar damage to CEP neurons. These results suggest that 222-UVC is less penetrating, so energy transfer occurs more effectively in tissues near the surface, causing more severe damage than 254-UVC.

Transgenic rice Oryza glaberrima with higher CPD photolyase activity alleviates UVB-caused growth inhibition.

The ultraviolet B (UVB) sensitivity of rice cultivated in Asia and Africa varies greatly, with African rice cultivars (Oryza glaberrima Steud. and O. barthii A. Chev.) being more sensitive to UVB because of their low cyclobutane pyrimidine dimer (CPD) photolyase activity, which is a CPD repair enzyme, relative to Asian rice cultivars (O. sativa L.). Hence, the production of UVB-resistant African rice with augmented CPD photolyase activity is of great importance, although difficulty in transforming the African rice cultivars to this end has been reported. Here, we successfully produced overexpressing transgenic African rice with higher CPD photolyase activity by modifying media conditions for callus induction and regeneration using the parental line (PL), UVB-sensitive African rice TOG12380 (O. glaberrima). The overexpressing transgenic African rice carried a single copy of the CPD photolyase enzyme, with a 4.4-fold higher level of CPD photolyase transcripts and 2.6-fold higher activity than its PL counterpart. When the plants were grown for 21 days in a growth chamber under visible radiation or with supplementary various UVB radiation, the overexpressing transgenic plants have a significantly increased UVB resistance index compared to PL plants. These results strongly suggest that CPD photolyase remains an essential factor for tolerating UVB radiation stress in African rice. As a result, African rice cultivars with overexpressed CPD photolyase may survive better in tropical areas more prone to UVB radiation stress, including Africa. Collectively, our results provide strong evidence that CPD photolyase is a useful biotechnological tool for reducing UVB-induced growth inhibition in African rice crops of O. glaberrima.

222 nm ultraviolet radiation C causes more severe damage to guard cells and epidermal cells of Arabidopsis plants than does 254 nm ultraviolet radiation.

Lamps that emit 222 nm short-wavelength ultraviolet (UV) radiation can be safely used for sterilization without harming human health. However, there are few studies on the effects of 222 nm UVC (222-UVC) radiation exposure on plants compared with the effects of germicidal lamps emitting primarily 254 nm UVC (254-UVC) radiation. We investigated the growth inhibition and cell damage caused by 222-UVC exposure to Arabidopsis plants, especially mitochondrial dynamics, which is an index of damage caused by UVB radiation. Growth inhibition resulted from 254-UVC or 222-UVC exposure depending on the dose of UVC radiation. However, with respect to the phenotype of 222-UVC-irradiated plants, the leaves curled under 1 kJ m-2 and were markedly bleached under 10 kJ m-2 compared with those of plants irradiated with 254-UVC. The cellular state, especially the mitochondrial dynamics, of epidermal and mesophyll cells of Arabidopsis leaves exposed to 254-UVC or 222-UVC radiation was investigated using Arabidopsis plants expressing mitochondrial matrix-targeted yellow fluorescent protein (MT-YFP) under the control of Pro35S to visualize the mitochondria. 222-UVC (1 or 5 kJ m-2) severely damaged the guard cells within the epidermis, and YFP signals and chloroplast autofluorescence in guard cells within the epidermis exposed to 222-UVC (1 or 5 kJ m-2) were not detected compared with those in cells exposed to 254-UVC radiation. In addition, 222-UVC irradiation led to mitochondrial fragmentation in mesophyll cells, similar to the effects of 254-UVC exposure. These results suggest that 222-UVC severely damages guard cells and epidermal cells and that such damage might have resulted in growth inhibition.

The characterization of tocols in different plants parts of six Japanese rice cultivars relating to their UVB-sensitivity.

There has been significant interest in the photosensitivity, or photo-resistance, of Japanese rice cultivars, which synthesize tocols (Vitamin E), a class of phytochemicals including tocol derivatives tocopherol (T) and tocotrienol (T3). In the present study, the distribution of tocols in the leaves, seeds, stems, and roots of six Japanese rice cultivars was investigated. The relationship between the different tocols in cultivars and their ultraviolet B sensitivity index (USB-SI) was analyzed. The leaves contained the highest average total amount of tocols at 230 µg.fresh-g-1, followed by seeds, stems, and roots. In leaves and stems, the most abundant component was α-T which was more than 85%. On the other hand, the tocols in seeds were 38% δ-T3, 32% α-T, and 20% α-T3. The tocols in roots were 55% α-T, 14% γ-T, and 13% δ-T3. The total tocol content in four plant parts exhibited a negative correlation (P < 0.05) in stem and root, and a negative relationship (r < -0.70) with the UVB-SI of the cultivars, suggesting that the total tocol contents were closely related to the resistance to UVB in Japanese rice plants.

UV-B signalling in rice: Response identification, gene expression profiling and mutant isolation.

Responses of rice seedlings to UV-B radiation (UV-B) were investigated, aiming to establish rice as a model plant for UV-B signalling studies. The growth of japonica rice coleoptiles, grown under red light, was inhibited by brief irradiation with UV-B, but not with blue light. The effective UV-B fluences (10-1 -103 µmol m-2 ) were much lower than those reported in Arabidopsis. The response was much less in indica rice cultivars and its extent varied among Oryza species. We next identified UV-B-specific anthocyanin accumulation in the first leaf of purple rice and used this visible phenotype to isolate mutants. Some isolated mutants were further characterized, and one was found to have a defect in the growth response. Using microarrays, we identified a number of genes that are regulated by low-fluence-rate UV-B in japonica coleoptiles. Some up-regulated genes were analysed by real-time PCR for UV-B specificity and the difference between japonica and indica. More than 70% of UV-B-regulated rice genes had no homologs in UV-B-regulated Arabidopsis genes. Many UV-B-regulated rice genes are related to plant hormones and especially to jasmonate biosynthetic and responsive genes in apparent agreement with the growth response. Possible involvement of two rice homologs of UVR8, a UV-B photoreceptor, is discussed.

Autophagy Contributes to the Quality Control of Leaf Mitochondria.

In autophagy, cytoplasmic components of eukaryotic cells are transported to lysosomes or the vacuole for degradation. Autophagy is involved in plant tolerance to the photooxidative stress caused by ultraviolet B (UVB) radiation, but its roles in plant adaptation to UVB damage have not been fully elucidated. Here, we characterized organellar behavior in UVB-damaged Arabidopsis (Arabidopsis thaliana) leaves and observed the occurrence of autophagic elimination of dysfunctional mitochondria, a process termed mitophagy. Notably, Arabidopsis plants blocked in autophagy displayed increased leaf chlorosis after a 1-h UVB exposure compared to wild-type plants. We visualized autophagosomes by labeling with a fluorescent protein-tagged autophagosome marker, AUTOPHAGY8 (ATG8), and found that a 1-h UVB treatment led to increased formation of autophagosomes and the active transport of mitochondria into the central vacuole. In atg mutant plants, the mitochondrial population increased in UVB-damaged leaves due to the cytoplasmic accumulation of fragmented, depolarized mitochondria. Furthermore, we observed that autophagy was involved in the removal of depolarized mitochondria when mitochondrial function was disrupted by mutation of the FRIENDLY gene, which is required for proper mitochondrial distribution. Therefore, autophagy of mitochondria functions in response to mitochondrion-specific dysfunction as well as UVB damage. Together, these results indicate that autophagy is centrally involved in mitochondrial quality control in Arabidopsis leaves.

Autophagy-deficient Arabidopsis mutant atg5, which shows ultraviolet-B sensitivity, cannot remove ultraviolet-B-induced fragmented mitochondria.

Mitochondria damaged by ultraviolet-B radiation (UV-B, 280-315 nm) are removed by mitophagy, a selective autophagic process. Recently, we demonstrated that autophagy-deficient Arabidopsis thaliana mutants exhibit a UV-B-sensitive phenotype like that of cyclobutane pyrimidine dimer (CPD)-specific photolyase (PHR1)-deficient mutants. To explore the relationship between UV-B sensitivity and autophagy in UV-B-damaged plants, we monitored mitochondrial dynamics and autophagy in wild-type Arabidopsis (ecotype Columbia); an autophagy-deficient mutant, atg5; a PHR1-deficient mutant, phr1; an atg5 phr1 double mutant; and AtPHR1-overexpressing (AtPHR1ox) plants following high-dose UV-B exposure (1.5 W m-2 for 1 h). At 10 h after exposure, the number of mitochondria per mesophyll leaf cell was increased and the volumes of individual mitochondria were decreased independently of UV-B-induced CPD accumulation in all genotypes. At 24 h after exposure, the mitochondrial number had recovered or almost recovered to pre-exposure levels in plants with functional autophagy (WT, phr1, and AtPHR1ox), but had increased even further in atg5. This suggested that the high dose of UV-B led to the inactivation and fragmentation of mitochondria, which were removed by mitophagy activated by UV-B. The UV-B-sensitive phenotype of the atg5 phr1 double mutant was more severe than that of atg5 or phr1. In wild-type, phr1, and AtPHR1ox plants, autophagy-related genes were strongly expressed following UV-B exposure independently of UV-B-induced CPD accumulation. Therefore, mitophagy might be one of the important repair mechanisms for UV-B-induced damage. The severe UV-B-sensitive phenotype of atg5 phr1 is likely an additive effect of deficiencies in independent machineries for UV-B protection, autophagy, and CPD photorepair.

Combined Environment Simulator for Low-Dose-Rate Radiation and Partial Gravity of Moon and Mars.

Deep space exploration by humans has become more realistic, with planned returns to the Moon, travel to Mars, and beyond. Space radiation with a low dose rate would be a constant risk for space travelers. The combined effects of space radiation and partial gravity such as on the Moon and Mars are unknown. The difficulty for such research is that there are no good simulating systems on the ground to investigate these combined effects. To address this knowledge gap, we developed the Simulator of the environments on the Moon and Mars with Neutron irradiation and Gravity change (SwiNG) for in vitro experiments using disposable closed cell culture chambers. The device simulates partial gravity using a centrifuge in a three-dimensional clinostat. Six samples are exposed at once to neutrons at a low dose rate (1 mGy/day) using Californium-252 in the center of the centrifuge. The system is compact including two SwiNG devices in the incubator, one with and one without radiation source, with a cooling function. This simulator is highly convenient for ground-based biological experiments because of limited access to spaceflight experiments. SwiNG can contribute significantly to research on the combined effects of space radiation and partial gravity.

Chloroplast Autophagy and Ubiquitination Combine to Manage Oxidative Damage and Starvation Responses.

Autophagy and the ubiquitin-proteasome system are the major degradation processes for intracellular components in eukaryotes. Although ubiquitination acts as a signal inducing organelle-targeting autophagy, the interaction between ubiquitination and autophagy in chloroplast turnover has not been addressed. In this study, we found that two chloroplast-associated E3 enzymes, SUPPRESSOR OF PPI1 LOCUS1 and PLANT U-BOX4 (PUB4), are not necessary for the induction of either piecemeal autophagy of chloroplast stroma or chlorophagy of whole damaged chloroplasts in Arabidopsis (Arabidopsis thaliana). Double mutations of an autophagy gene and PUB4 caused synergistic phenotypes relative to single mutations. The double mutants developed accelerated leaf chlorosis linked to the overaccumulation of reactive oxygen species during senescence and had reduced seed production. Biochemical detection of ubiquitinated proteins indicated that both autophagy and PUB4-associated ubiquitination contributed to protein degradation in the senescing leaves. Furthermore, the double mutants had enhanced susceptibility to carbon or nitrogen starvation relative to single mutants. Together, these results indicate that autophagy and chloroplast-associated E3s cooperate for protein turnover, management of reactive oxygen species accumulation, and adaptation to starvation.

SUPPRESSOR OF GAMMA RESPONSE 1 acts as a regulator coordinating crosstalk between DNA damage response and immune response in Arabidopsis thaliana.

Plants live in constantly changing and often unfavorable or stressful environments. Environmental changes induce biotic and abiotic stress, which, in turn, may cause genomic DNA damage. Hence, plants simultaneously suffer abiotic/biotic stress and DNA damage. However, little information is available on the signaling crosstalk that occurs between DNA damage and abiotic/biotic stresses. Arabidopsis thaliana SUPPRESSOR OF GAMMA RESPONSE1 (SOG1) is a pivotal transcription factor that regulates thousands of genes in response to DNA double-strand break (DSB), and we recently reported that SOG1 has a role in immune responses. In the present study, the effects of SOG1 overexpression on the DNA damage and immune responses were examined. Results found that SOG1 overexpression enhances the regulation of numerous downstream genes. Relative to the wild type plants, then, DNA damage responses were observed to be strongly induced. SOG1 overexpression also upregulates chitin (a major components of fungal cell walls) responsive genes in the presence of DSBs, implying that pathogen defense response is activated by DNA damage via SOG1. Further, SOG1 overexpression enhances fungal resistance. These results suggest that SOG1 regulates crosstalk between DNA damage response and the immune response and that plants have evolved a sophisticated defense network to contend with environmental stress.

Space Radiation Biology for "Living in Space".

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.

Very high sensitivity of African rice to artificial ultraviolet-B radiation caused by genotype and quantity of cyclobutane pyrimidine dimer photolyase.

Ultraviolet-B (UVB) radiation damages plants and decreases their growth and productivity. We previously demonstrated that UVB sensitivity varies widely among Asian rice (Oryza sativa L.) cultivars and that the activity of cyclobutane pyrimidine dimer (CPD) photolyase, which repairs UVB-induced CPDs, determines UVB sensitivity. Unlike Asian rice, African rice (Oryza glaberrima Steud. and Oryza barthii A. Chev.) has mechanisms to adapt to African climates and to protect itself against biotic and abiotic stresses. However, information about the UVB sensitivity of African rice species is largely absent. We showed that most of the African rice cultivars examined in this study were UVB-hypersensitive or even UVB-super-hypersensitive in comparison with the UVB sensitivity of Asian O. sativa cultivars. The difference in UVB resistance correlated with the total CPD photolyase activity, which was determined by its activity and its cellular content. The UVB-super-hypersensitive cultivars had low enzyme activity caused by newly identified polymorphisms and low cellular CPD photolyase contents. The new polymorphisms were only found in cultivars from West Africa, particularly in those from countries believed to be centres of O. glaberrima domestication. This study provides new tools for improving both Asian and African rice productivity.

Expression Profile of Cell Cycle-Related Genes in Human Fibroblasts Exposed Simultaneously to Radiation and Simulated Microgravity.

Multiple unique environmental factors such as space radiation and microgravity (µG) pose a serious threat to human gene stability during space travel. Recently, we reported that simultaneous exposure of human fibroblasts to simulated µG and radiation results in more chromosomal aberrations than in cells exposed to radiation alone. However, the mechanisms behind this remain unknown. The purpose of this study was thus to obtain comprehensive data on gene expression using a three-dimensional clinostat synchronized to a carbon (C)-ion or X-ray irradiation system. Human fibroblasts (1BR-hTERT) were maintained under standing or rotating conditions for 3 or 24 h after synchronized C-ion or X-ray irradiation at 1 Gy as part of a total culture time of 2 days. Among 57,773 genes analyzed with RNA sequencing, we focused particularly on the expression of 82 cell cycle-related genes after exposure to the radiation and simulated µG. The expression of cell cycle-suppressing genes (ABL1 and CDKN1A) decreased and that of cell cycle-promoting genes (CCNB1, CCND1, KPNA2, MCM4, MKI67, and STMN1) increased after C-ion irradiation under µG. The cell may pass through the G1/S and G2 checkpoints with DNA damage due to the combined effects of C-ions and µG, suggesting that increased genomic instability might occur in space.

Divergent evolution of rice blast resistance Pi54 locus in the genus Oryza.

BACKGROUND: The rice blast resistance gene Pi54 was cloned from Oryza sativa ssp. indica cv. Tetep, which conferred broad-spectrum resistance against Magnaporthe oryzae. Pi54 allelic variants have been identified in not only domesticates but also wild rice species, but the majority of japonica and some indica cultivars lost the function. RESULTS: We here found that Pi54 (Os11g0639100) and its homolog Os11g0640600 (named as #11) were closely located on a 25 kbp region in japonica cv. Sasanishiki compared to a 99 kbp region in japonica cv. Nipponbare. Sasanishiki lost at least six genes containing one other R-gene cluster (Os11g0639600, Os11g0640000, and Os11g0640300). Eight AA-genome species including five wild rice species were classified into either Nipponbare or Sasanishiki type. The BB-genome wild rice species O. punctata was Sasanishiki type. The FF-genome wild rice species O. brachyantha (the basal lineage of Oryza) was neither, because Pi54 was absent and the orientation of the R-gene cluster was reversed in comparison with Nipponbare-type species. The phylogenetic analysis showed that #11gene of O. brachyantha was on the root of both Pi54 and #11 alleles. All Nipponbare-type Pi54 alleles were specifically disrupted by 143 and 37/44 bp insertions compared to Tetep and Sasanishiki type. In addition, Pi54 of japonica cv. Sasanishiki lost nucleotide-binding site and leucine-rich repeat (NBS-LRR) domains owing to additional mutations. CONCLUSIONS: These results suggest that Pi54 might be derived from a tandem duplication of the ancestor #11 gene in progenitor FF-genome species. Two divergent structures of Pi54 locus caused by a mobile unit containing the nearby R-gene cluster could be developed before domestication. This study provides a potential genetic resource of rice breeding for blast resistance in modern cultivars sustainability.

Selective Elimination of Membrane-Damaged Chloroplasts via Microautophagy.

Plant chloroplasts constantly accumulate damage caused by visible wavelengths of light during photosynthesis. Our previous study revealed that entire photodamaged chloroplasts are subjected to vacuolar digestion through an autophagy process termed chlorophagy; however, how this process is induced and executed remained poorly understood. In this study, we monitored intracellular induction of chlorophagy in Arabidopsis (Arabidopsis thaliana) leaves and found that mesophyll cells damaged by high visible light displayed abnormal chloroplasts with a swollen shape and 2.5 times the volume of normal chloroplasts. In wild-type plants, the activation of chlorophagy decreased the number of swollen chloroplasts. In the autophagy-deficient autophagy mutants, the swollen chloroplasts persisted, and dysfunctional chloroplasts that had lost chlorophyll fluorescence accumulated in the cytoplasm. Chloroplast swelling and subsequent induction of chlorophagy were suppressed by the application of exogenous mannitol to increase the osmotic pressure outside chloroplasts or by overexpression of VESICLE INDUCING PROTEIN IN PLASTID1, which maintains chloroplast envelope integrity. Microscopic observations of autophagy-related membranes showed that swollen chloroplasts were partly surrounded by autophagosomal structures and were engulfed directly by the tonoplast, as in microautophagy. Our results indicate that an elevation in osmotic potential inside the chloroplast due to high visible light-derived envelope damage results in chloroplast swelling and serves as an induction factor for chlorophagy, and this process mobilizes entire chloroplasts via tonoplast-mediated sequestering to avoid the cytosolic accumulation of dysfunctional chloroplasts.

Development and performance evaluation of a three-dimensional clinostat synchronized heavy-ion irradiation system.

Outer space is an environment characterized by microgravity and space radiation, including high-energy charged particles. Astronauts are constantly exposed to both microgravity and radiation during long-term stays in space. However, many aspects of the biological effects of combined microgravity and space radiation remain unclear. We developed a new three-dimensional (3D) clinostat synchronized heavy-ion irradiation system for use in ground-based studies of the combined exposures. Our new system uses a particle accelerator and a respiratory gating system from heavy-ion radiotherapy to irradiate samples being rotated in the 3D clinostat with carbon-ion beams only when the samples are in the horizontal position. A Peltier module and special sample holder were loaded on a static stage (standing condition) and the 3D clinostat (rotation condition) to maintain a suitable temperature under atmospheric conditions. The performance of the new device was investigated with normal human fibroblasts 1BR-hTERT in a disposable closed cell culture chamber. Live imaging revealed that cellular adhesion and growth were almost the same for the standing control sample and rotation sample over 48h. Dose flatness and symmetry were judged according to the relative density of Gafchromic films along the X-axis and Y-axis of the positions of the irradiated sample to confirm irradiation accuracy. Doses calculated using the carbon-ion calibration curve were almost the same for standing and rotation conditions, with the difference being less than 5% at 1Gy carbon-ion irradiation. Our new device can accurately synchronize carbon-ion irradiation and simulated microgravity while maintaining the temperature under atmospheric conditions at ground level.

Entire Photodamaged Chloroplasts Are Transported to the Central Vacuole by Autophagy.

Turnover of dysfunctional organelles is vital to maintain homeostasis in eukaryotic cells. As photosynthetic organelles, plant chloroplasts can suffer sunlight-induced damage. However, the process for turnover of entire damaged chloroplasts remains unclear. Here, we demonstrate that autophagy is responsible for the elimination of sunlight-damaged, collapsed chloroplasts in Arabidopsis thaliana We found that vacuolar transport of entire chloroplasts, termed chlorophagy, was induced by UV-B damage to the chloroplast apparatus. This transport did not occur in autophagy-defective atg mutants, which exhibited UV-B-sensitive phenotypes and accumulated collapsed chloroplasts. Use of a fluorescent protein marker of the autophagosomal membrane allowed us to image autophagosome-mediated transport of entire chloroplasts to the central vacuole. In contrast to sugar starvation, which preferentially induced distinct type of chloroplast-targeted autophagy that transports a part of stroma via the Rubisco-containing body (RCB) pathway, photooxidative damage induced chlorophagy without prior activation of RCB production. We further showed that chlorophagy is induced by chloroplast damage caused by either artificial visible light or natural sunlight. Thus, this report establishes that an autophagic process eliminates entire chloroplasts in response to light-induced damage.

Ultraviolet-B-induced DNA damage and ultraviolet-B tolerance mechanisms in species with different functional groups coexisting in subalpine moorlands.

High doses of ultraviolet-B (UV-B; 280-315 nm) radiation can have detrimental effects on plants, and especially damage their DNA. Plants have DNA repair and protection mechanisms to prevent UV-B damage. However, it remains unclear how DNA damage and tolerance mechanisms vary among field species. We studied DNA damage and tolerance mechanisms in 26 species with different functional groups coexisting in two moorlands at two elevations. We collected current-year leaves in July and August, and determined accumulation of cyclobutane pyrimidine dimer (CPD) as UV-B damage and photorepair activity (PRA) and concentrations of UV-absorbing compounds (UACs) and carotenoids (CARs) as UV-B tolerance mechanisms. DNA damage was greater in dicot than in monocot species, and higher in herbaceous than in woody species. Evergreen species accumulated more CPDs than deciduous species. PRA was higher in Poaceae than in species of other families. UACs were significantly higher in woody than in herbaceous species. The CPD level was not explained by the mechanisms across species, but was significantly related to PRA and UACs when we ignored species with low CPD, PRA and UACs, implying the presence of another effective tolerance mechanism. UACs were correlated negatively with PRA and positively with CARs. Our results revealed that UV-induced DNA damage significantly varies among native species, and this variation is related to functional groups. DNA repair, rather than UV-B protection, dominates in UV-B tolerance in the field. Our findings also suggest that UV-B tolerance mechanisms vary among species under evolutionary trade-off and synergism.