Citation: Cox MM, Keck JL, Battista JR (2010) Rising from the Ashes: DNA Repair in Deinococcus radiodurans. PLoS Genet 6(1): e1000815. https://doi.org/10.1371/journal.pgen.1000815
Editor: Susan M. Rosenberg, Baylor College of Medicine, United States of America
Published: January 15, 2010
Copyright: © 2010 Cox et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Funding: The authors received no specific funding for this work.
Competing interests: The authors have declared that no competing interests exist.
The extraordinary resistance of Deinococcus radiodurans to ionizing radiation (IR) and desiccation is slowly drawing more intense scrutiny. Relative to most other organisms, Deinococcus has a survival advantage measured in orders of magnitude. Exposure to 5 kGy of IR reduces the genome of any bacterium to hundreds of fragments. Deinococcus is no exception. However, Deinococcus seems to take this catastrophe in stride. Over a period of 3–4 hours, overlapping fragments are spliced together into complete chromosomes, and the cells soon resume normal growth. There is no measurable lethality. Attempts to understand the molecular basis of this phoenix-like capability has given rise to numerous hypotheses. Notable among them are proposals that the condensed nature of the Deinococcus genome  or an unusual capacity to avoid protein oxidation  are keys to radiation resistance.
Regardless of the physiological or metabolic adaptations that Deinococcus may employ to enhance survival, it is hard to explain extreme genome reconstitution without considering DNA repair. As originally defined by Daly and Minton , and reinforced more recently by Radman and colleagues ,, genome reconstitution in Deinococcus proceeds in two phases. The first phase has been attributed to a process dubbed extended synthesis-dependent single-strand DNA annealing (ESDSA) ,. The second phase involves RecA protein-mediated double-strand break repair. Some initial studies suggested that the first phase of repair did not involve the Deinococcus RecA protein, but more recent work has documented a role for RecA in both phases ,. ESDSA involves considerable nuclease activity to generate single-stranded DNA, strand invasion mediated by the RecA and/or RadA proteins, and extensive DNA synthesis primed by the invading strands prior to the annealing steps ,.
The emerging picture (Figure 1) provides a useful framework, but one with many questions. Most of these involve enzymes and their roles. The generation of single-stranded DNA in ESDSA requires the function of at least one nuclease and helicase and perhaps several of both. Proteins are also needed to load RecA protein onto single-strand DNA binding protein (SSB)-coated single-stranded DNA. The most important pathway for double-strand break repair in Escherichia coli utilizes the RecBCD enzyme for all of these roles. However, Deinococcus encodes no homologue of RecB or RecC. Deinococcus does encode homologues of every protein involved in what is considered (in E. coli) an auxiliary pathway for recombinational DNA repair—the RecFOR pathway. In the RecFOR pathway, the RecJ and RecQ proteins take up the nuclease and helicase roles, respectively, while the RecFOR proteins function to load RecA protein onto the DNA. The absence of RecBC homologues in Deinococcus seems to imply that the RecFOR path is particularly important.
The first stage, extended synthesis-dependent single-strand annealing (ESDSA) is dominated by nuclease and DNA polymerase functions. The second stage is a more conventional RecA-mediated double-strand break repair process focused on the final splicing of large chromosomal segments.
In two important reports in this issue of PLoS Genetics, Sommer and colleagues convert much recent speculation into substance. In the first report , the Chandler and Sommer laboratories collaborate to explore the mechanism of transposition of element ISDra2. This transposon is a member of a family of elements that transpose via single-stranded DNA intermediates. Transposition is activated by irradiation of Deinococcus. The work not only documents the transposition mechanism, it reinforces the proposition that extensive lengths of single-stranded genomic DNA are generated in the early stages of genome reconstitution in this bacterium. As a bonus, the work provides hope for the development of in vivo transposition as a tool for genetic manipulation of this genome.
The second report  clearly establishes the central role of the RecFOR pathway in genome reconstitution. Where the effects of RecFOR pathway gene deficiencies are generally modest in E. coli, they are dramatic in D. radiodurans. Sommer and colleagues demonstrate that cells lacking functional recF, recO, or recR genes are essentially as dysfunctional in genome reconstitution as recA mutants. Cells with these deficiencies are viable in the absence of extreme DNA damage, but grow much slower than wild type. Thus, the RecFOR pathway is important during normal replication, as well as during genome reconstitution. If the gene encoding the RecJ nuclease is inactivated, the cells are inviable. Surprisingly, cells lacking the RecQ helicase exhibit wild type resistance to IR. Instead, the helicase that appears to be critical is UvrD. In E. coli, the function of UvrD in recombinational DNA repair is to remove RecA filaments from the DNA ,. In Deinococcus, it almost certainly does more. In summary, Deinococcus now presents a unique opportunity to demonstrate what the RecFOR pathway can really do.
This RecFOR pathway may look a bit different from its cousin in E. coli. After irradiation, about 60 Deinococcus genes are induced, and quite a number of them have roles in genome reconstitution . Many of these genes are novel. Knockouts do not have the dramatic effects of recAJFOR knockouts, but their cumulative effects can be significant. Some, like the DdrA protein (a distant homologue of the eukaryotic Rad52 protein ), and DdrB (a novel single-strand DNA binding protein ) must be worked into the schemes.
The work of Chandler and Sommer has a few more far-reaching implications. The spectacular feat of genome reconstitution after heavy irradiation does not require a completely new pathway for double-strand break repair, and no such pathway appears to be present. Instead, D. radiodurans relies heavily on a set of recombinational DNA repair functions that are recognizable in almost all species. In large measure, efficient genome reconstitution involves tweaking those repair proteins, providing a few novel augmentations, and perhaps modifying the environment in which all of these proteins function. However, the properties already noted that distinguish the RecFOR pathway in Deinococcus from the same process in E. coli bear reiteration. The roles that well-known repair proteins play in radioresistance are not perfectly predictable, based on what we understand about their function in E. coli. An orthologous relationship between proteins can inform speculation, but it must be subjected to experimental substantiation. Every DNA repair protein examined to date in D. radiodurans has provided one or more novel twists in our understanding of its function, structure, interaction with other proteins, and role in repair.
Last but not least, we may soon see a first in vitro reconstitution of a complete DNA double-strand break repair reaction. In this arena, D. radiodurans is gradually eclipsing E. coli as the most pliable bacterial model system. The proteins, or at least most of them, are in hand. Fortuitously, the enzymes from Deinococcus appear to be more amenable to structural determination than their E. coli cousins. The only structural information currently available about RecF, RecO, and RecR come from the Deinococcus enzymes –. Activities are being characterized, and more surprises are anticipated. Deinococcus simply does it better.
- 1. Levin-Zaidman S, Englander J, Shimoni E, Sharma AK, Minton KW, et al. (2003) Ringlike structure of the Deinococcus radiodurans genome: a key to radioresistance? Science 299: 254–256.S. Levin-ZaidmanJ. EnglanderE. ShimoniAK SharmaKW Minton2003Ringlike structure of the Deinococcus radiodurans genome: a key to radioresistance?Science299254256
- 2. Daly MJ, Gaidamakova EK, Matrosova VY, Vasilenko A, Zhai M, et al. (2007) Protein oxidation implicated as the primary determinant of bacterial radioresistance. PLoS Biol 5: e92.MJ DalyEK GaidamakovaVY MatrosovaA. VasilenkoM. Zhai2007Protein oxidation implicated as the primary determinant of bacterial radioresistance.PLoS Biol5e92
- 3. Daly MJ, Minton KW (1996) An alternative pathway of recombination of chromosomal fragments precedes recA-dependent recombination in the radioresistant bacterium Deinococcus radiodurans. J Bacteriol 178: 4461–4471.MJ DalyKW Minton1996An alternative pathway of recombination of chromosomal fragments precedes recA-dependent recombination in the radioresistant bacterium Deinococcus radiodurans.J Bacteriol17844614471
- 4. Slade D, Lindner AB, Paul G, Radman M (2009) Recombination and replication in DNA repair of heavily irradiated Deinococcus radiodurans. Cell 136: 1044–1055.D. SladeAB LindnerG. PaulM. Radman2009Recombination and replication in DNA repair of heavily irradiated Deinococcus radiodurans.Cell13610441055
- 5. Zahradka K, Slade D, Bailone A, Sommer S, Averbeck D, et al. (2006) Reassembly of shattered chromosomes in Deinococcus radiodurans. Nature 443: 569–573.K. ZahradkaD. SladeA. BailoneS. SommerD. Averbeck2006Reassembly of shattered chromosomes in Deinococcus radiodurans.Nature443569573
- 6. Pasternak C, Ton-Hoang B, Coste G, Bailone A, Chandler M, et al. (2009) Irradiation-induced D. radiodurans genome fragmentation triggers transposition of a single resident insertion sequence. PLoS Genet 6: e1000799.C. PasternakB. Ton-HoangG. CosteA. BailoneM. Chandler2009Irradiation-induced D. radiodurans genome fragmentation triggers transposition of a single resident insertion sequence.PLoS Genet6e1000799
- 7. Bentchikou E, Servant P, Coste G, Sommer S (2009) A major role of the RecFOR pathway in DNA double-strand-break repair through ESDSA in Deinococcus radiodurans. PLoS Genet 6: e1000774.E. BentchikouP. ServantG. CosteS. Sommer2009A major role of the RecFOR pathway in DNA double-strand-break repair through ESDSA in Deinococcus radiodurans.PLoS Genet6e1000774
- 8. Centore RC, Sandler SJ (2007) UvrD limits the number and intensities of RecA-green fluorescent protein structures in Escherichia coli K-12. J Bacteriol 189: 2915–2920.RC CentoreSJ Sandler2007UvrD limits the number and intensities of RecA-green fluorescent protein structures in Escherichia coli K-12.J Bacteriol18929152920
- 9. Veaute X, Delmas P, Selva M, Jeusset J, Le Cam E, et al. (2005) UvrD helicase, unlike Rep helicase, dismantles RecA nucleoprotein filaments in Escherichia coli. EMBO J 24: 180–189.X. VeauteP. DelmasM. SelvaJ. JeussetE. Le Cam2005UvrD helicase, unlike Rep helicase, dismantles RecA nucleoprotein filaments in Escherichia coli.EMBO J24180189
- 10. Tanaka M, Earl AM, Howell HA, Park MJ, Eisen JA, et al. (2004) Analysis of Deinococcus radiodurans's transcriptional response to ionizing radiation and desiccation reveals novel proteins that contribute to extreme radioresistance. Genetics 168: 21–33.M. TanakaAM EarlHA HowellMJ ParkJA Eisen2004Analysis of Deinococcus radiodurans's transcriptional response to ionizing radiation and desiccation reveals novel proteins that contribute to extreme radioresistance.Genetics1682133
- 11. Harris DR, Tanaka M, Saveliev SV, Jolivet E, Earl AM, et al. (2004) Preserving genome integrity: the DdrA protein of Deinococcus radiodurans R1. PLoS Biology 2: e304.DR HarrisM. TanakaSV SavelievE. JolivetAM Earl2004Preserving genome integrity: the DdrA protein of Deinococcus radiodurans R1.PLoS Biology2e304
- 12. Norais C, Chitteni-Pattu S, Wood EA, Inman RB, Cox MM (2009) DdrB protein, an alternative Deinococcus radiodurans SSB induced by ionizing radiation. J Biol Chem 284: 21402–21411.C. NoraisS. Chitteni-PattuEA WoodRB InmanMM Cox2009DdrB protein, an alternative Deinococcus radiodurans SSB induced by ionizing radiation.J Biol Chem2842140221411
- 13. Koroleva O, Makharashvili N, Courcelle CT, Courcelle J, Korolev S (2007) Structural conservation of RecF and Rad50: implications for DNA recognition and RecF function. EMBO J 26: 867–877.O. KorolevaN. MakharashviliCT CourcelleJ. CourcelleS. Korolev2007Structural conservation of RecF and Rad50: implications for DNA recognition and RecF function.EMBO J26867877
- 14. Leiros I, Timmins J, Hall DR, McSweeney S (2005) Crystal structure and DNA-binding analysis of RecO from Deinococcus radiodurans. EMBO J 24: 906–918.I. LeirosJ. TimminsDR HallS. McSweeney2005Crystal structure and DNA-binding analysis of RecO from Deinococcus radiodurans.EMBO J24906918
- 15. Makharashvili N, Koroleva O, Bera S, Grandgenett DP, Korolev S (2004) A novel structure of DNA repair protein RecO from Deinococcus radiodurans. Structure 12: 1881–1889.N. MakharashviliO. KorolevaS. BeraDP GrandgenettS. Korolev2004A novel structure of DNA repair protein RecO from Deinococcus radiodurans.Structure1218811889
- 16. Makharashvili N, Koroleva O, Korolev S (2005) Structural studies of replication/recombination mediator proteins. Biophys J 88: 14A–14A.N. MakharashviliO. KorolevaS. Korolev2005Structural studies of replication/recombination mediator proteins.Biophys J8814A14A
- 17. Timmins J, Leiros I, McSweeney S (2007) Crystal structure and mutational study of RecOR provide insight into its mode of DNA binding. EMBO J 26: 3260–3271.J. TimminsI. LeirosS. McSweeney2007Crystal structure and mutational study of RecOR provide insight into its mode of DNA binding.EMBO J2632603271