Antioxidant defense of Deinococcus radiodurans: how does it contribute to extreme radiation resistance?

PURPOSE: Deinococcus radiodurans is an extremely radioresistant bacterium characterized by D10 of 10 kGy, and able to grow luxuriantly under chronic ionizing radiation of 60 Gy/h. The aim of this article is to review the antioxidant system of D. radiodurans and its possible role in the unusual resistance of this bacterium to ionizing radiation. CONCLUSIONS: The unusual radiation resistance of D. radiodurans has apparently evolved as a side effect of the adaptation of this extremophile to other damaging environmental factors, especially desiccation. The antioxidant proteins and low-molecular antioxidants (especially low-molecular weight Mn2+ complexes and carotenoids, in particular, deinoxanthin), as well as protein and non-protein regulators, are important for the antioxidant defense of this species. Antioxidant protection of proteins from radiation inactivation enables the repair of DNA damage caused by ionizing radiation.

Deinococcus lineage and Rad52 family-related protein DR0041 is involved in DNA protection and compaction.

DR0041 ORF encodes an uncharacterized Deinococcus lineage protein. We earlier reported presence of DR0041 protein in DNA repair complexes of Ssb and RecA in Deinococcus radiodurans. Here, we systematically examined the role of DR0041 in DNA metabolism using various experimental methodologies including electrophoretic mobility assays, nuclease assays, strand exchange assays and transmission electron microscopy. Interaction between DR0041 and the C-terminal acidic tail of Ssb was assessed through co-expression and in vivo cross-linking studies. A knockout mutant was constructed to understand importance of DR0041 ORF for various physiological processes. Results highlight binding of DR0041 protein to single-stranded and double-stranded DNA, interaction with Ssb-coated single-stranded DNA without interference with RecA-mediated strand exchange, protection of DNA from exonucleases, and compaction of high molecular weight DNA molecules into tightly condensed forms. Bridging and compaction of sheared DNA by DR0041 protein might have implications in the preservation of damaged DNA templates to maintain genome integrity upon exposure to gamma irradiation. Our results suggest that DR0041 protein is dispensable for growth under standard growth conditions and following gamma irradiation but contributes to protection of DNA during transformation. We discuss the role of DR0041 protein from the perspective of protection of broken DNA templates and functional redundancy.

iTRAQ-based proteomic analysis of Deinococcus radiodurans in response to 12C6+ heavy ion irradiation.

BACKGROUND: Deinococcus radiodurans (D. radiodurans) is best known for its extreme resistance to diverse environmental stress factors, including ionizing radiation (IR), ultraviolet (UV) irradiation, oxidative stress, and high temperatures. Robust DNA repair system and antioxidant system have been demonstrated to contribute to extreme resistance in D. radiodurans. However, practically all studies on the mechanism underlying D. radiodurans's extraordinary resistance relied on the treated strain during the post-treatment recovery lag phase to identify the key elements involved. The direct gene or protein changes of D. radiodurans after stress have not yet been characterized. RESULTS: In this study, we performed a proteomics profiling on D. radiodurans right after the heavy ion irradiation treatment, to discover the altered proteins that were quickly responsive to IR in D. radiodurans. Our study found that D. radiodurans shown exceptional resistance to 12C6+ heavy ion irradiation, in contrast to Escherichia coli (E.coli) strains. By using iTRAQ (Isobaric Tags for Relative and Absolute Quantitation)-based quantitative mass spectrometry analysis, the kinetics of proteome changes induced by various dosages of 12C6+ heavy ion irradiation were mapped. The results revealed that 452 proteins were differentially expressed under heavy ion irradiation, with the majority of proteins being upregulated, indicating the upregulation of functional categories of translation, TCA cycle (Tricarboxylic Acid cycle), and antioxidation regulation under heavy ion irradiation. CONCLUSIONS: This study shows how D. radiodurans reacts to exposure to 12C6+ heavy ion irradiation in terms of its overall protein expression profile. Most importantly, comparing the proteome profiling of D. radiodurans directly after heavy ion irradiation with research on the post-irradiation recovery phase would potentially provide a better understanding of mechanisms underlying the extreme radioresistance in D. radiodurans.

Comprehensive Temporal Protein Dynamics during Postirradiation Recovery in Deinococcus radiodurans.

Deinococcus radiodurans (D. radiodurans) is an extremophile that can tolerate ionizing radiation, ultraviolet radiation, and oxidation. How D. radiodurans responds to and survives high levels of ionizing radiation is still not clear. In this study, we performed label-free proteomics to explore the proteome dynamics during postirradiation recovery (PIR). Surprisingly, proteins involved in translation were repressed during the initial hours of PIR. D. radiodurans also showed enhanced DNA repair and antioxidative response after 6 kGy of gamma irradiation. Moreover, proteins involved in sulfur metabolism and phenylalanine metabolism were enriched at 1 h and 12 h, respectively, indicating different energy and material needs during PIR. Furthermore, based on these findings, we proposed a novel model to elucidate the possible molecular mechanisms of robust radioresistance in D. radiodurans, which may serve as a reference for future radiation repair.

Proteomic profiling of Deinococcus radiodurans with response to thioredoxin reductase inhibitor and ionizing radiation treatment.

This study explains the importance of cellular redox system in preserving the proteome of the radioresistant Deinococcus radiodurans. The thioredoxin reductase (TrxR) redox system was inhibited by ebselen (10 µM), and then the bacterium was exposed to 4 kGy of ionizing radiation. The differentially expressed proteins were analyzed using label-free quantitative (LFQ) proteomics. The 4 kGy radiation treatment increases the expression of stress response proteins like osmotically inducible protein OsmC, catalase, and metallophosphoesterase compared to control. Ebselen plus radiation treatment augments oxidoreductases proteins in D. radiodurans. Further, the proteins involved in glycolysis, tricarboxylic acetic acid (TCA) and proteins like proteases, peptidase, and peptide transporters were significantly decreased in the ebselen plus radiation group compared to radiation treated group. Further, ebselen plus radiation treatment increases the ATP-binding cassette (ABC) transporters involved in the efflux of toxic chemicals and nutrient uptake and the stress response related membrane protein like S-layer homology domain-containing protein in D. radiodurans. Thus, the results show that the altered redox status via inhibition of TrxR redox system significantly affects the expression of essential cellular proteins for the survival. The cellular content of D. radiodurans may be used to handle redox imbalances in the normal cells during cancer radiotherapy. SIGNIFICANCE: Deinococcus radiodurans is a popular radioresistance organism with efficient antioxidant systems and DNA repair mechanisms. There are many antioxidant systems and small molecules that responsible for its resistance. The importance of thiol based antioxidant systems in its resistance property has not fully studied yet. Thioredoxin reductase is an important disulfide containing protein that involved in maintaining redox homeostasis. The TrxR inhibition affects the cell survival and synthesis of molecules against ionizing radiation. In this study we are reporting the effects of TrxR inhibitor on proteome of D. radiodurans upon ionizing radiation. This study reveals the significance of TrxR antioxidant system on the proteome of D. radiodurans. The inhibition of TrxR antioxidant system and the subsequent disturbances in the proteome content makes the organism vulnerable to oxidative stress.

Transposition of insertion sequences by dielectric barrier discharge plasma and gamma irradiation in the radiation-resistant bacterium Deinococcus geothermalis.

Insertion sequences (ISs) of the radiation-resistant bacterium Deinococcus geothermalis are transposed into other loci by oxidative stress through hydrogen peroxide treatment. Gamma irradiation and dielectric barrier discharge (DBD) plasma radiation are known to produce a variety of oxidative stress agents such as reactive oxygen species and reactive nitrogen species. Therefore, to determine whether the transposition of ISs was induced in D. geothermalis by both gamma irradiation and DBD plasma radiation, we selected non-pigmented mutants with disrupted target genes encoding carotenoid biosynthesis enzymes such as a phytoene synthase (dgeo_0523) and a phytoene desaturase (dgeo_0524). Different DNA-binding protein-deficient mutants exhibited novel transposition of ISs. Dps (dgeo_0257), OxyR (dgeo_1888), and the LysR (dgeo_2840) family regulator, in addition to cystine importer-disrupted and -overexpressed mutants (dgeo_1986-87 and dgeo_1985R) and wild-type D. geothermalis were tested in this study. Active IS transposition was not detected in two wild-type control species (Deinococcus radiodurans and Deinococcus radiopugnans) after phenotypic selection in gamma irradiation. Our finding demonstrated that gamma irradiation triggers the transposition of particular IS elements, especially ISDge2 and ISDge3 of the IS1 family, ISDge5 of the IS701 family, and ISDge6 of the IS5 family in wild-type strain and the Δdgeo_0257, Δdgeo_1986-87, Δdgeo_1985R, and Δdgeo_2840 mutants. Furthermore, DBD plasma radiation triggered the transposition of ISDge11 of the IS4 family in the wild-type strain; ISDge6 of the IS5 family on Δdgeo_0257, Δdgeo_1888 and Δdgeo_2840; ISDge5 of the IS701 family on Δdgeo_0257 strain.

Small-Molecule Mn Antioxidants in Caenorhabditis elegans and Deinococcus radiodurans Supplant MnSOD Enzymes during Aging and Irradiation.

Denham Harman's oxidative damage theory identifies superoxide (O2•-) radicals as central agents of aging and radiation injury, with Mn2+-dependent superoxide dismutase (MnSOD) as the principal O2•--scavenger. However, in the radiation-resistant nematode Caenorhabditis elegans, the mitochondrial antioxidant enzyme MnSOD is dispensable for longevity, and in the model bacterium Deinococcus radiodurans, it is dispensable for radiation resistance. Many radiation-resistant organisms accumulate small-molecule Mn2+-antioxidant complexes well-known for their catalytic ability to scavenge O2•-, along with MnSOD, as exemplified by D. radiodurans. Here, we report experiments that relate the MnSOD and Mn-antioxidant content to aging and oxidative stress resistances and which indicate that C. elegans, like D. radiodurans, may rely on Mn-antioxidant complexes as the primary defense against reactive oxygen species (ROS). Wild-type and ΔMnSOD D. radiodurans and C. elegans were monitored for gamma radiation sensitivities over their life spans while gauging Mn2+-antioxidant content by electron paramagnetic resonance (EPR) spectroscopy, a powerful new approach to determining the in vivo Mn-antioxidant content of cells as they age. As with D. radiodurans, MnSOD is dispensable for radiation survivability in C. elegans, which hyperaccumulates Mn-antioxidants exceptionally protective of proteins. Unexpectedly, ΔMnSOD mutants of both the nematodes and bacteria exhibited increased gamma radiation survival compared to the wild-type. In contrast, the loss of MnSOD renders radiation-resistant bacteria sensitive to atmospheric oxygen during desiccation. Our results support the concept that the disparate responses to oxidative stress are explained by the accumulation of Mn-antioxidant complexes which protect, complement, and can even supplant MnSOD. IMPORTANCE The current theory of cellular defense against oxidative damage identifies antioxidant enzymes as primary defenders against ROS, with MnSOD being the preeminent superoxide (O2•-) scavenger. However, MnSOD is shown to be dispensable both for radiation resistance and longevity in model organisms, the bacterium Deinococcus radiodurans and the nematode Caenorhabditis elegans. Measured by electron paramagnetic resonance (EPR) spectroscopy, small-molecule Mn-antioxidant content was shown to decline in unison with age-related decreases in cell proliferation and radioresistance, which again are independent of MnSOD presence. Most notably, the Mn-antioxidant content of C. elegans drops precipitously in the last third of its life span, which links with reports that the steady-state level of oxidized proteins increases exponentially during the last third of the life span in animals. This leads us to propose that global responses to oxidative stress must be understood through an extended theory that includes small-molecule Mn-antioxidants as potent O2•--scavengers that complement, and can even supplant, MnSOD.

A small RNA regulates pprM, a modulator of pleiotropic proteins promoting DNA repair, in Deinococcus radiodurans under ionizing radiation.

Networks of transcriptional and post-transcriptional regulators are critical for bacterial survival and adaptation to environmental stressors. While transcriptional regulators provide rapid activation and/or repression of a wide-network of genes, post-transcriptional regulators, such as small RNAs (sRNAs), are also important to fine-tune gene expression. However, the mechanisms of sRNAs remain poorly understood, especially in less-studied bacteria. Deinococcus radiodurans is a gram-positive bacterium resistant to extreme levels of ionizing radiation (IR). Although multiple unique regulatory systems (e.g., the Radiation and Desiccation Response (RDR)) have been identified in this organism, the role of post-transcriptional regulators has not been characterized within the IR response. In this study, we have characterized an sRNA, PprS (formerly Dsr2), as a post-transcriptional coordinator of IR recovery in D. radiodurans. PprS showed differential expression specifically under IR and knockdown of PprS resulted in reduced survival and growth under IR, suggesting its importance in regulating post-radiation recovery. We determined a number of potential RNA targets involved in several pathways including translation and DNA repair. Specifically, we confirmed that PprS binds within the coding region to stabilize the pprM (DR_0907) transcript, a RDR modulator. Overall, these results are the first to present an additional layer of sRNA-based control in DNA repair pathways associated with bacterial radioresistance.

Experimental evolution of extremophile resistance to ionizing radiation.

A growing number of known species possess a remarkable characteristic - extreme resistance to the effects of ionizing radiation (IR). This review examines our current understanding of how organisms can adapt to and survive exposure to IR, one of the most toxic stressors known. The study of natural extremophiles such as Deinococcus radiodurans has revealed much. However, the evolution of Deinococcus was not driven by IR. Another approach, pioneered by Evelyn Witkin in 1946, is to utilize experimental evolution. Contributions to the IR-resistance phenotype affect multiple aspects of cell physiology, including DNA repair, removal of reactive oxygen species, the structure and packaging of DNA and the cell itself, and repair of iron-sulfur centers. Based on progress to date, we overview the diversity of mechanisms that can contribute to biological IR resistance arising as a result of either natural or experimental evolution.

New insights into the activation of Radiation Desiccation Response regulon in Deinococcus radiodurans.

The highly radiation-resistant bacterium Deinococcus radiodurans responds to gamma radiation or desiccation through the coordinated expression of genes belonging to Radiation and Desiccation Resistance/Response (RDR) regulon. RDR regulon is operated through cis-acting sequence RDRM (Radiation Desiccation Response Motif), trans-acting repressor DdrO and protease IrrE (also called PprI). The present study evaluated whether RDR regulon controls the response of D. radiodurans to various other DNA damaging stressors, to which it is resistant, such as UV rays, mitomycin C (MMC), methyl methanesulfonate (MMS), ethidium bromide (EtBr), etc. Activation of 3 RDR regulon genes (ddrB, gyrB and DR1143) was studied by tagging their promoter sequences with a highly sensitive GFP reporter. Here we demonstrated that all the DNA damaging stressors elicited activation of RDR regulon of D. radiodurans in a dose-dependent and RDRM-/IrrE-dependent manner. However, ROS-mediated indirect effects [induced by hydrogen peroxide (H2O2), methyl viologen (MV), heavy metal/metalloid (zinc or tellurite), etc.] did not activate RDR regulon. We also showed that level of activation was inversely proportional to cellular abundance of repressor DdrO. Our data strongly suggests that direct DNA damage activates RDR regulon in D. radiodurans.

Redox signaling through zinc activates the radiation response in Deinococcus bacteria.

Deinococcus bacteria are extremely resistant to radiation and other DNA damage- and oxidative stress-generating conditions. An efficient SOS-independent response mechanism inducing expression of several DNA repair genes is essential for this resistance, and is controlled by metalloprotease IrrE that cleaves and inactivates transcriptional repressor DdrO. Here, we identify the molecular signaling mechanism that triggers DdrO cleavage. We show that reactive oxygen species (ROS) stimulate the zinc-dependent metalloprotease activity of IrrE in Deinococcus. Sudden exposure of Deinococcus to zinc excess also rapidly induces DdrO cleavage, but is not accompanied by ROS production and DNA damage. Further, oxidative treatment leads to an increase of intracellular free zinc, indicating that IrrE activity is very likely stimulated directly by elevated levels of available zinc ions. We conclude that radiation and oxidative stress induce changes in redox homeostasis that result in IrrE activation by zinc in Deinococcus. We propose that a part of the zinc pool coordinated with cysteine thiolates is released due to their oxidation. Predicted regulation systems involving IrrE- and DdrO-like proteins are present in many bacteria, including pathogens, suggesting that such a redox signaling pathway including zinc as a second messenger is widespread and participates in various stress responses.

G-quadruplex DNA structures and their relevance in radioprotection.

BACKGROUND: DNA, the genetic material of most of the organisms, is the crucial element of life. Integrity of DNA needs to be maintained for transmission of genetic material from one generation to another. All organisms are constantly challenged by the environmental conditions which can lead to the induction of DNA damage. Ionizing radiation (IR) has been known to induce DNA damage and IR sensitivity varies among different organisms. The causes for differential radiosensitivity among various organisms have not been studied in great detail. SCOPE OF REVIEW: We discuss DNA secondary structure formation, GC content of the genome, role of G-quadruplex formation, and its relationship with radiosensitivity of the genome. MAJOR CONCLUSION: In Deinococcus radiodurans, the bacterium that exhibits maximum radio resistance, multiple G-quadruplex forming motifs are reported. In human cells, G-quadruplex formation led to differential radiosensitivity. In this article, we have discussed, the role of secondary DNA structure formation like G-quadruplex in shielding the genome from radiation and its implications in understanding evolution of radio protective effect of an organism. We also discuss role of GC content and its correlation with radio resistance. GENERAL SIGNIFICANCE: This review provides an insight into the role of G-quadruplexes in providing differential radiosensitivity at different site of the genome and in different organisms. It further discusses the possibility of higher GC content contributing towards reduced radiosensitivity in different organisms, evolution of radiosensitivity, and regulation of multiple cellular processes.

Topoisomerase IB interacts with genome segregation proteins and is involved in multipartite genome maintenance in Deinococcus radiodurans.

Deinococcus radiodurans, an extremophile, resistant to many abiotic stresses including ionizing radiation, has 2 type I topoisomerases (drTopo IA and drTopo IB) and one type II topoisomerase (DNA gyrase). The role of drTopo IB in guanine quadruplex DNA (G4 DNA) metabolism was demonstrated earlier in vitro. Here, we report that D. radiodurans cells lacking drTopo IB (ΔtopoIB) show sensitivity to G4 DNA binding drug (NMM) under normal growth conditions. The activity of G4 motif containing promoters like mutL and recQ was reduced in the presence of NMM in mutant cells. In mutant, the percentage of anucleate cells was more while the copy number of genome elements were less as compared to wild type. Protein-protein interaction studies showed that drTopo IB interacts with genome segregation and DNA replication initiation (DnaA) proteins. The typical patterns of cellular localization of GFP-PprA were affected in the mutant cells. Microscopic examination of D. radiodurans cells expressing drTopo IB-RFP showed its localization on nucleoid forming a streak parallel to the old division septum and perpendicular to newly formed septum. These results together suggest the role of drTopo IB in genome maintenance in this bacterium.

Characterization of ori and parS-like functions in secondary genome replicons in Deinococcus radiodurans.

The mechanisms underlying multipartite genome maintenance and its functional significance in extraordinary radioresistance of Deinococcus radiodurans are not well understood. The sequences upstream to parAB operons in chrII (cisII) and MP (cisMP) could stabilize an otherwise, non-replicative colE1 plasmid, in D. radiodurans DnaA and cognate ParB proteins bound specifically with cisII and cisMP elements. The ΔcisII and ΔcisMP cells showed the reduced copy number of cognate replicons and radioresistance as compared with wild type. Fluorescent reporter-operator system inserted in chrI, chrII, and MP in wild type and cisII mutants showed the presence of all three replicons in wild-type cells. Although chrI was present in all the ΔcisII and ΔcisMP cells, nearly half of these cells had chrII and MP, respectively, and the other half had the reduced number of foci representing these replications. These results suggested that cisII and cisMP elements contain both origin of replication and parS-like functions and the secondary genome replicons (chrII and MP) are maintained independent of chrI and have roles in radioresistance of D. radiodurans.

Ultraviolet-B Radiation Stress-Induced Toxicity and Alterations in Proteome of Deinococcus radiodurans.

Deinococcus radiodurans is a polyextremophilic bacterium capable to survive and grow at high doses of ionizing radiation. Besides resistance to ionizing radiation, the bacterium is also resistant to toxic chemicals and desiccation. This study deals with the effects of non-ionizing radiation (ultraviolet-B) on survival, alterations in proteomic profile, and gene expression in D. radiodurans. Exposure of culture to UV-B caused decrease in the percentage survival with increasing duration, complete killing occurred after 16 h. D. radiodurans also showed enhancement in the generation of reactive oxygen species and activities of antioxidative enzymes. Separation of proteins by 2-dimensional gel electrophoresis revealed major changes in number and abundance of different proteins. Twenty-eight differentially abundant protein spots were identified by MALDI-TOF MS/MS analysis and divided into 8 groups including unknown proteins. Gene expression of a few identified proteins was also analyzed employing qRT-PCR, which showed differential expression corresponding to the respective proteins. In silico analysis of certain hypothetical proteins (HPs) suggested that these are novel and as yet not reported from D. radiodurans subjected to UV-B stress. These HPs may prove useful in future studies especially for assessing their significance in the adaptation and management of stress responses against UV-B stress.

Comparative genomics of wild-type and laboratory-evolved biofilm-overproducing Deinococcus metallilatus strains.

Deinococcus metallilatus MA1002 was exposed to ultraviolet radiation to generate mutants with enhanced biofilm production. Two strains (nos 5 and 6) were then selected based on their high biofilm formation, as well as their possession of higher concentrations of extracellular matrix components (eDNA, protein and saccharides) than the wild-type (WT). Genomic sequencing revealed the presence of large genome deletions in a secondary chromosome in the mutants. Expression analyses of the WT and mutant strains indicated the upregulation of genes associated with exopolysaccharide synthesis and stress response. The mutant strains showed high mortality in glucose-supplemented (TYG) medium; however, cell death and biofilm formation were not increased in mutant cells grown under acetate- or glyoxylate-added media, suggesting that metabolic toxicity during glucose metabolism induced a high rate of cell death but improved biofilm formation in mutant strains. In damaged cells, eDNAs contributed to the enhanced biofilm formation of D. metallilatus.

Molecular repertoire of Deinococcus radiodurans after 1 year of exposure outside the International Space Station within the Tanpopo mission.

BACKGROUND: The extraordinarily resistant bacterium Deinococcus radiodurans withstands harsh environmental conditions present in outer space. Deinococcus radiodurans was exposed for 1 year outside the International Space Station within Tanpopo orbital mission to investigate microbial survival and space travel. In addition, a ground-based simulation experiment with conditions, mirroring those from low Earth orbit, was performed. METHODS: We monitored Deinococcus radiodurans cells during early stage of recovery after low Earth orbit exposure using electron microscopy tools. Furthermore, proteomic, transcriptomic and metabolomic analyses were performed to identify molecular mechanisms responsible for the survival of Deinococcus radiodurans in low Earth orbit. RESULTS: D. radiodurans cells exposed to low Earth orbit conditions do not exhibit any morphological damage. However, an accumulation of numerous outer-membrane-associated vesicles was observed. On levels of proteins and transcripts, a multi-faceted response was detected to alleviate cell stress. The UvrABC endonuclease excision repair mechanism was triggered to cope with DNA damage. Defense against reactive oxygen species is mirrored by the increased abundance of catalases and is accompanied by the increased abundance of putrescine, which works as reactive oxygen species scavenging molecule. In addition, several proteins and mRNAs, responsible for regulatory and transporting functions showed increased abundances. The decrease in primary metabolites indicates alternations in the energy status, which is needed to repair damaged molecules. CONCLUSION: Low Earth orbit induced molecular rearrangements trigger multiple components of metabolic stress response and regulatory networks in exposed microbial cells. Presented results show that the non-sporulating bacterium Deinococcus radiodurans survived long-term low Earth orbit exposure if wavelength below 200 nm are not present, which mirrors the UV spectrum of Mars, where CO2 effectively provides a shield below 190 nm. These results should be considered in the context of planetary protection concerns and the development of new sterilization techniques for future space missions. Video Abstract.

Enhanced Lycopene Production by UV-C Irradiation in Radiation-Resistant Deinococcus radiodurans R1.

Although classical metabolic engineering strategies have succeeded in developing microbial strains capable of producing desired bioproducts, metabolic imbalance resulting from extensive genetic manipulation often leads to decreased productivity. Thus, abiotic strategies for improving microbial production performance can be an alternative to overcome drawbacks arising from intensive metabolic engineering. Herein, we report a promising abiotic method for enhancing lycopene production by UV-C irradiation using a radiation-resistant ΔcrtLm/crtB+dxs+ Deinococcus radiodurans R1 strain. First, the onset of UV irradiation was determined through analysis of the expression of 11 genes mainly involved in the carotenoid biosynthetic pathway in the ΔcrtLm/crtB+dxs+ D. radiodurans R1 strain. Second, the effects of different UV wavelengths (UV-A, UV-B, and UV-C) on lycopene production were investigated. UV-C irradiation induced the highest production, resulting in a 69.9% increase in lycopene content [64.2 ± 3.2 mg/g dry cell weight (DCW)]. Extended UV-C irradiation further enhanced lycopene content up to 73.9 ± 2.3 mg/g DCW, a 95.5% increase compared to production without UV-C irradiation (37.8 ± 0.7 mg/g DCW).

Exposure of Deinococcus radiodurans to both static magnetic fields and gamma radiation: observation of cell recuperation effects.

The extremophilic bacterium Deinococcus radiodurans displays an extraordinary ability to withstand lethal radiation effects, due to its complex mechanisms for both proteome radiation protection and DNA repair. Published results obtained recently at this laboratory show that D. radiodurans submitted to ionizing radiation results in its DNA being shattered into small fragments which, when exposed to a "static electric field' (SEF), greatly decreases cell viability. These findings motivated the performing of D. radiodurans exposed to gamma radiation, yet exposed to a different exogenous physical agent, "static magnetic fields" (SMF). Cells of D. radiodurans [strain D.r. GY 9613 (R1)] in the exponential phase were submitted to 60Co gamma radiation from a gamma cell. Samples were exposed to doses in the interval 0.5-12.5 kGy, while the control samples were kept next to the irradiation setup. Exposures to SMF were carried out with intensities of 0.08 T and 0.8 T delivered by two settings: (a) a device built up at this laboratory with niobium magnets, delivering 0.08 T, and (b) an electromagnet (Walker Scientific) generating static magnetic fields with intensities from 0.1 to 0.8 T. All samples were placed in a bacteriological incubator at 30 °C for 48 h, and after incubation, a counting of colony forming units was performed. Two sets of cell surviving data were measured, each in triplicate, obtained in independent experiments. A remarkable similarity between the two data sets is revealed, underscoring reproducibility within the 5% range. Appraisal of raw data shows that exposure of irradiated cells to SMF substantially increases their viability. Data interpretation strongly suggests that the increase of D. radiodurans cell viability is a sole magnetic physical effect, driven by a stochastic process, improving the efficiency of the rejoining of DNA fragments, thus increasing cell viability. A type of cut-off dose is identified at 10 kGy, above which the irradiated cellular system loses recovery and the cell survival mechanism collapses.

Deinococcus terrestris sp. nov., a gamma ray- and ultraviolet-resistant bacterium isolated from soil.

Strain SDU3-2T was isolated from a soil sample collected in Shandong Province, PR China. Cells of SDU3-2T were spherical, Gram-stain-positive, aerobic and non-motile. Cellular growth of the strain occurred at 25-45 °C, pH 5.5-8.5 and with 0-1.5 % (w/v) of NaCl. Phylogenetic analysis based on the 16S rRNA gene sequences showed that strain SDU3-2T was closest to the type strain Deinococcus murrayi ALT-1bT with a similarity of 95.2 %. The draft genome was 3.49 Mbp long with 69.2 mol% G+C content. Strain SDU3-2T exhibited high resistance to gamma radiation (D10 >12 kGy) and UV (D10 >900 J m-2). The strain encoded many genes for resistance to radiation and oxidative stress, which were highly conserved with other Deinococcus species, but possessed interspecific properties. The major fatty acids of SDU3-2T cells were C15 : 1 ω6c, C16 : 1 ω7c/C16 : 1 ω6c, and C17 : 1 ω8c, the major menaquinone was menaquinone-8, and the major polar lipids were an unidentified phosphoglycolipid, four unidentified glycolipids and an unidentified phospholipid. The average nucleotide identity and DNA-DNA hybridization results further indicated that strain SDU3-2T represents a new species in the genus Deinococcus, for which the name Deinococcus terrestris sp. nov. is proposed. The type strain is SDU3-2T (=CGMCC 1.17147T=KCTC 43098T).