Oxidative Stress Resistance in Deinococcus radiodurans

Summary: Deinococcus radiodurans is a robust bacterium best known for its capacity to repair massive DNA damage efficiently and accurately. It is extremely resistant to many DNA-damaging agents, including ionizing radiation and UV radiation (100 to 295 nm), desiccation, and mitomycin C, which induce oxidative damage not only to DNA but also to all cellular macromolecules via the production of reactive oxygen species. The extreme resilience of D. radiodurans to oxidative stress is imparted synergistically by an efficient protection of proteins against oxidative stress and an efficient DNA repair mechanism, enhanced by functional redundancies in both systems. D. radiodurans assets for the prevention of and recovery from oxidative stress are extensively reviewed here. Radiation- and desiccation-resistant bacteria such as D. radiodurans have substantially lower protein oxidation levels than do sensitive bacteria but have similar yields of DNA double-strand breaks. These findings challenge the concept of DNA as the primary target of radiation toxicity while advancing protein damage, and the protection of proteins against oxidative damage, as a new paradigm of radiation toxicity and survival. The protection of DNA repair and other proteins against oxidative damage is imparted by enzymatic and nonenzymatic antioxidant defense systems dominated by divalent manganese complexes. Given that oxidative stress caused by the accumulation of reactive oxygen species is associated with aging and cancer, a comprehensive outlook on D. radiodurans strategies of combating oxidative stress may open new avenues for antiaging and anticancer treatments. The study of the antioxidation protection in D. radiodurans is therefore of considerable potential interest for medicine and public health.     

Death by protein damage in irradiated cells

A founding concept of radiobiology that deals with X-rays, γ-rays and ultraviolet light is that radiation indiscriminately damages cellular macromolecules. Mounting experimental evidence does not fit into this theoretical framework. Whereas DNA lesion-yields in cells exposed to a given dose and type of radiation appear to be fixed, protein lesion-yields are highly variable. Extremely radiation resistant bacteria such as Deinococcus radiodurans have evolved extraordinarily efficient antioxidant chemical defenses which specifically protect proteins and the functions they catalyze. In diverse prokaryotes, the lethal effects of radiation appear to be governed by oxidative protein damage, which inactivates enzymes including those needed to repair and replicate DNA. These findings offer fresh insight into the molecular mechanisms of radiation resistance and present themselves as new opportunities to study and control oxidative stress in eukaryotes, including mammalian cells and their cancer cell counterparts.     

Endurance of the endolithic desert cyanobacterium Chroococcidiopsis under UVC radiation

Desert cyanobacteria of the genus Chroococcidiopsis are extremely resistant to desiccation and ionizing radiation. When an endolithic strain was exposed to UVC radiation cell lysis, genome damage, photosynthetic pigment bleaching and reduced photochemical performance occurred. Nevertheless, survivors were scored after UVC doses as high as 13?kJ/m² and their endurance ascribed to multicellular aggregates enveloped in thick envelopes, so that attenuated UVC radiation reached the inner cells. In addition, the accumulation of carotenoids contributed to UVC resistance by providing protection against oxidative stress. Finally, in survivors repair mechanisms were responsible for the recovery of the induced damage to genome and photosynthetic apparatus.     

Intracellular Copper Accumulation Enhances the Growth of Kineococcus radiotolerans during Chronic Irradiation

The actinobacterium Kineococcus radiotolerans is highly resistant to ionizing radiation, desiccation, and oxidative stress, though the underlying biochemical mechanisms are unknown. The purpose of this study was to explore a possible linkage between the uptake of transition metals and extreme resistance to ionizing radiation and oxidative stress. The effects of six different divalent cationic metals on growth were examined in the absence of ionizing radiation. None of the metals tested were stimulatory, though cobalt was inhibitory to growth. In contrast, copper supplementation dramatically increased colony formation during chronic irradiation. K. radiotolerans exhibited specific uptake and intracellular accumulation of copper, compared to only a weak response to both iron and manganese supplementation. Copper accumulation sensitized cells to hydrogen peroxide. Acute-irradiation-induced DNA damage levels were similar in the copper-loaded culture and the age-synchronized no-copper control culture, though low-molecular-weight DNA was more persistent during postirradiation recovery in the Cu-loaded culture. Still, the estimated times for genome restoration differed by only 2 h between treatments. While we cannot discount the possibility that copper fulfills an unexpectedly important biochemical role in a low-radioactivity environment, K. radiotolerans has a high capacity for intracellular copper sequestration and presumably efficiently coordinated oxidative stress defenses and detoxification systems, which confers cross-protection from the damaging effects of ionizing radiation.     

Genome of the extremely radiation-resistant bacterium Deinococcus radiodurans viewed from the perspective of comparative genomics.

The bacterium Deinococcus radiodurans shows remarkable resistance to a range of damage caused by ionizing radiation, desiccation, UV radiation, oxidizing agents, and electrophilic mutagens. D. radiodurans is best known for its extreme resistance to ionizing radiation; not only can it grow continuously in the presence of chronic radiation (6 kilorads/h), but also it can survive acute exposures to gamma radiation exceeding 1,500 kilorads without dying or undergoing induced mutation. These characteristics were the impetus for sequencing the genome of D. radiodurans and the ongoing development of its use for bioremediation of radioactive wastes. Although it is known that these multiple resistance phenotypes stem from efficient DNA repair processes, the mechanisms underlying these extraordinary repair capabilities remain poorly understood. In this work we present an extensive comparative sequence analysis of the Deinococcus genome. Deinococcus is the first representative with a completely sequenced genome from a distinct bacterial lineage of extremophiles, the Thermus-Deinococcus group. Phylogenetic tree analysis, combined with the identification of several synapomorphies between Thermus and Deinococcus, supports the hypothesis that it is an ancient group with no clear affinities to any of the other known bacterial lineages. Distinctive features of the Deinococcus genome as well as features shared with other free-living bacteria were revealed by comparison of its proteome to the collection of clusters of orthologous groups of proteins. Analysis of paralogs in Deinococcus has revealed several unique protein families. In addition, specific expansions of several other families including phosphatases, proteases, acyltransferases, and Nudix family pyrophosphohydrolases were detected. Genes that potentially affect DNA repair and recombination and stress responses were investigated in detail. Some proteins appear to have been horizontally transferred from eukaryotes and are not present in other bacteria. For example, three proteins homologous to plant desiccation resistance proteins were identified, and these are particularly interesting because of the correlation between desiccation and radiation resistance. Compared to other bacteria, the D. radiodurans genome is enriched in repetitive sequences, namely, IS-like transposons and small intergenic repeats. In combination, these observations suggest that several different biological mechanisms contribute to the multiple DNA repair-dependent phenotypes of this organism.     

The effect of gamma-radiation and desiccation on the viability of the soil bacteria isolated from the alienated zone around the Chernobyl Nuclear Power Plant

Methylobacterium extorquens, M. mesophilicum, and Bacillus subtilis strains were found to be resistant to gamma-radiation, irrespective of whether they were isolated from the alienated zone around the Chernobyl Nuclear Power Plant or outside this zone. The LD90 of Methylobacterium and B. subtilis strains with respect to gamma-radiation was 2.0-3.4 and 3.7-4.4 kGy, respectively, whereas their LD99.99 values were 4.5-6.9 and more than 10 kGy, respectively. The high threshold levels of gamma-radiation for Methylobacterium and B. subtilis imply the efficient functioning of DNA repair systems in these bacteria. Unlike Bacillus polymyxa cells, the cells of M. extorquens, M. mesophilicum, and B. subtilis were also resistant to desiccation. Pseudomonas sp., Nocardia sp., and nocardioform actinomycetes were sensitive to both gamma-radiation and desiccation. Similar results were obtained when the bacteria studied were exposed to hydrogen peroxide and ultraviolet radiation. The results obtained indicate that the bacteria that are resistant to gamma-radiation are also resistant to desiccation, UV radiation, and hydrogen peroxide. The possibility of using simple laboratory tests (such as the determination of bacterial resistance to UV light and desiccation) for the evaluation of bacterial resistance to gamma-radiation is discussed.     

Effect of <Superscript>60</Superscript>Co-Gamma Ionizing Radiation and Desiccation Stress on Protein Profile of <Emphasis Type="Italic">Anabaena</Emphasis> 7120

Physiological stress can bring major molecular and cellular change to a living cell which further decide its survival or tolerance to the stress exposure. Cyanobacteria like Anabaena has been shown to tolerate high levels of different stresses like oxidative, desiccation, UV, and gamma radiation. They are able to withstand and recover remarkably without any lethal mutation when exposed to high doses of gamma radiation or prolonged duration of desiccation. In the present work, the modifications in protein profiles of Anabaena 7120 cells after exposure to 6 kGy of ⁶⁰Co γ-rays and 6 days of desiccation, and the proteome dynamics during post stress recovery were investigated. Differentially expressed proteins during stress and recovery were identified by MALDI-ToF or LC-MS, which generated a partial proteome map of Anabaena 7120. Anabaena cells went through protein recycling—phase of protein degradation following by their resynthesis, which helped them to recover remarkably. The data suggests an overlap in proteome changes during recovery against radiation and desiccation stress.     

紫外线-B辐射对植物DNA及蛋白质的影响

大气平流层中的臭氧衰减,导致太阳辐射中的紫外辐射量有明显的增加,其中UV-B辐射对植物会产生不同程度的影响。分子生态学理论认为,UV-B辐射对植物造成的损伤,首先伤害植物的生物大分子,即进行光化学修饰。本文就臭氧衰减对生态环境和植物的影响途径进行了讨论,重点论述了UV-B辐射对植物蛋白质合成的抑制和DNA的损伤修复途径。并应用分子生物学技术研究植物对UV-B辐射的抗性机理和DNA修复技术的前景进行了展望。     

Unstructured Hydrophilic Sequences in Prokaryotic Proteomes Correlate with Dehydration Tolerance and Host Association

Water loss or desiccation is among the most life-threatening stresses. It leads to DNA double-strand breakage, protein aggregation, cell shrinkage, and low water activity precluding all biological functions. Yet, in all kingdoms of life, rare organisms are resistant to desiccation through prevention or reversibility of such damage. Here, we explore possible hallmarks of prokaryotic desiccation tolerance in their proteomes. The content of unstructured, low complexity (LC) regions was analyzed in a total of 460 bacterial and archaeal proteomes.It appears that species endowed with proteomes abundant in unstructured hydrophilic LC regions are desiccation-tolerant or sporulating bacteria, halophilic archaea and bacteria, or host-associated species. In the desiccation- and radiation-resistant bacterium Deinococcus radiodurans, most proteins that contain large hydrophilic LC regions have unassigned function, but those with known function are mostly involved in diverse cellular recovery processes. Such LC regions are typically absent in orthologous proteins in desiccation-sensitive species. D. radiodurans encodes also special LC proteins, akin to those associated with desiccation resistance of plant seeds and some plants and animals. Therefore, we postulate that large unstructured hydrophilic LC regions and proteins provide for cellular resistance to dehydration and we discuss mechanisms of their protective activity.     

The chicken or the egg? Adaptation to desiccation and salinity tolerance in a lineage of water beetles

Transitions from fresh to saline habitats are restricted to a handful of insect lineages, as the colonization of saline waters requires specialized mechanisms to deal with osmotic stress. Previous studies have suggested that tolerance to salinity and desiccation could be mechanistically and evolutionarily linked, but the temporal sequence of these adaptations is not well established for individual lineages. We combined molecular, physiological and ecological data to explore the evolution of desiccation resistance, hyporegulation ability (i.e., the ability to osmoregulate in hyperosmotic media) and habitat transitions in the water beetle genus Enochrus subgenus Lumetus (Hydrophilidae). We tested whether enhanced desiccation resistance evolved before increases in hyporegulation ability or vice versa, or whether the two mechanisms evolved in parallel. The most recent ancestor of Lumetus was inferred to have high desiccation resistance and moderate hyporegulation ability. There were repeated shifts between habitats with differing levels of salinity in the radiation of the group, those to the most saline habitats generally occurring more rapidly than those to less saline ones. Significant and accelerated changes in hyporegulation ability evolved in parallel with smaller and more progressive increases in desiccation resistance across the phylogeny, associated with the colonization of meso‐ and hypersaline waters during global aridification events. All species with high hyporegulation ability were also desiccation‐resistant, but not vice versa. Overall, results are consistent with the hypothesis that desiccation resistance mechanisms evolved first and provided the physiological basis for the development of hyporegulation ability, allowing these insects to colonize and diversify across meso‐ and hypersaline habitats.     

Induction of Hsp70 by desiccation, ionising radiation and heat-shock in the eutardigrade Richtersius coronifer

The physiology and biochemistry behind the extreme tolerance to desiccation shown by the so-called anhydrobiotic animals represents an exciting challenge to biology. The current knowledge suggests that both carbohydrates and proteins are often involved in protecting the dry cell from damage, or in the repair of induced damage. Tardigrades belong to the most desiccation-tolerant multicellular organisms, but very little research has been reported on the biochemistry behind desiccation tolerance in this group. We quantified the induction of the heat-shock protein Hsp70, a very wide-spread stress protein, in response to desiccation, ionising radiation, and heating, in the anhydrobiotic tardigrade Richtersius coronifer using an immuno-westernblot method. Elevated levels of Hsp70 were recorded after treatment of both heat and ionising radiation, and also in rehydrated tardigrades after a period of desiccation. In contrast, tardigrades in the desiccated (dry) state had reduced Hsp70 levels compared to the non-treated control group. Our results suggest that Hsp70 may be involved in the physiological and biochemical system underlying desiccation (and radiation) tolerance in tardigrades, and that its role may be connected to repair processes after desiccation rather than to biochemical stabilization in the dry state.     

Radioresistance of Deinococcus radiodurans: functions necessary to survive ionizing radiation are also necessary to survive prolonged desiccation.

Forty-one ionizing radiation-sensitive strains of Deinococcus radiodurans were evaluated for their ability to survive 6 weeks of desiccation. All exhibited a substantial loss of viability upon rehydration compared with wild-type D. radiodurans. Examination of chromosomal DNA from desiccated cultures revealed a time-dependent increase in DNA damage, as measured by an increase in DNA double-strand breaks. The evidence presented suggests that D. radiodurans' ionizing radiation resistance is incidental, a consequence of this organism's adaptation to a common physiological stress, dehydration.     

The influence of UV radiation on protistan evolution

Ultraviolet radiation has provided an evolutionary challenge to life on Earth. Recent increases in surficial ultraviolet B fluxes have focused attention on the role of UV radiation in protistan ecology, cancer, and DNA damage. Exploiting this new wealth of data, I examine the possibility that ultraviolet radiation may have played a significant role in the evolution of the first eukaryotes, that is, protists. Protists probably arose well before the formation of a significant ozone shield, and thus were probably subjected to substantial ultraviolet A, ultraviolet B, and ultraviolet C fluxes early in their evolution. Evolution consists of the generation of heritable variations and the subsequent selection of these variants. Ultraviolet radiation has played a role both as a mutagen and as a selective agent. In its role as a mutagen, it may have been crucial in the origin of sex and as a driver of molecular evolution. As a selective agent, its influence has been broad. Discussed in this paper are the influence of ultraviolet radiation on biogeography, photosynthesis, and desiccation resistance.     

Green algae in alpine biological soil crust communities: acclimation strategies against ultraviolet radiation and dehydration

Green algae are major components of biological soil crusts in alpine habitats. Together with cyanobacteria, fungi and lichens, green algae form a pioneer community important for the organisms that will succeed them. In their high altitudinal habitat these algae are exposed to harsh and strongly fluctuating environmental conditions, mainly intense irradiation, including ultraviolet radiation, and lack of water leading to desiccation. Therefore, green algae surviving in these environments must have evolved with either avoidance or protective strategies, as well as repair mechanisms for damage. In this review we have highlighted these mechanisms, which include photoprotection, photochemical quenching, and high osmotic values to avoid water loss, and in some groups flexibility of secondary cell walls to maintain turgor pressure even in water-limited situations. These highly specialized green algae will serve as good model organisms to study desiccation tolerance or photoprotective mechanisms, due to their natural capacity to withstand unfavorable conditions. We point out the urgent need for modern phylogenetic approaches in characterizing these organisms, and molecular methods for analyzing the metabolic changes involved in their adaptive strategies.     

抗辐射菌Deinococcus radiodurans的多样性

抗辐射菌Deinococcus radiodurans是一种对电离辐射和其他DNA损伤因子具有极强抵抗能力的细菌,是研究DNA损伤与修复的模式生物。综述了国内外在抗辐射菌研究上取得的最新研究成果,从生存环境、对DNA损伤因子的抗性、抗性机理及其损伤修复关联基因等方面报道了抗辐射菌的多样性,并探讨了该细菌高效正确的DNA损伤修复机理的相关研究成果在生命科学、农业、环境修复及医学等领域的应用前景。     

A novel cold-inducible gene from Arabidopsis,RCI3, encodes a peroxidase that constitutes a component for stress tolerance

Myrothamnus flabellifolia, a short woody shrub from southern Africa, can survive severe desiccation of its vegetative organs. We studied mechanisms protecting this plant from oxidative damage during desiccation for 2 weeks, 4 and 8 months, and also during subsequent rehydration. This plant retains high concentrations of chlorophyll during desiccation, and these chlorophyll molecules are probably a source for potentially harmful singlet oxygen production. Desiccation triggered substantial increases in zeaxanthin and redox shifts of the antioxidants glutathione and ascorbate towards their oxidised forms. Simultaneously, the concentrations of violaxanthin, beta-carotene, ascorbate, alpha-tocopherol, and glutathione reductase activity progressively decreased. Antheraxanthin, gamma-tocopherol, lutein, neoxanthin and glucose-6-phosphate dehydrogenase displayed less pronounced changes in response to desiccation. Even after 4 months of desiccation, Myrothamnus flabellifolia recovered rapidly upon rehydration. Re-watering induced formation of ascorbate and glutathione, simultaneous reduction of their oxidised forms, and rapid production of alpha-tocopherol and of various carotenoids. Only after 8 months of desiccation did the antioxidant system of M. flabellifolia break down; 3 weeks after the onset of rehydration, these plants abscised their leaves, but even then they were still able to recover and develop new ones. Ascorbate, beta-carotene and alpha-tocopherol were totally depleted after 8 months of desiccation and did not recover upon rehydration; glutathione was partly maintained, but only in the oxidised form. We present a model demonstrating which parts of antioxidant pathways break down as oxidative stress becomes detrimental and we discuss some potential implications of our results for the genetic modification of crop plants to improve their drought tolerance.     

Rate of dehydration and cumulative desiccation stress interacted to modulate desiccation tolerance of recalcitrant cocoa and ginkgo embryonic tissues

Rate of dehydration greatly affects desiccation tolerance of recalcitrant seeds. This effect is presumably related to two different stress vectors: direct mechanical or physical stress because of the loss of water and physicochemical damage of tissues as a result of metabolic alterations during drying. The present study proposed a new theoretic approach to represent these two types of stresses and investigated how seed tissues responded differently to two stress vectors, using the models of isolated cocoa (Theobroma cacao) and ginkgo (Ginkgo biloba) embryonic tissues dehydrated under various drying conditions. This approach used the differential change in axis water potential (DeltaPsi/Deltat) to quantify rate of dehydration and the intensity of direct physical stress experienced by embryonic tissues during desiccation. Physicochemical effect of drying was expressed by cumulative desiccation stress [integralf(psi,t)], a function of both the rate and time of dehydration. Rapid dehydration increased the sensitivity of embryonic tissues to desiccation as indicated by high critical water contents, below which desiccation damage occurred. Cumulative desiccation stress increased sharply under slow drying conditions, which was also detrimental to embryonic tissues. This quantitative analysis of the stress-time-response relationship helps to understand the physiological basis for the existence of an optimal dehydration rate, with which maximum desiccation tolerance could be achieved. The established numerical analysis model will prove valuable for the design of experiments that aim to elucidate biochemical and physiological mechanisms of desiccation tolerance.