Conservation and diversity of radiation and ox.pdf

FEMS Microbiology Reviews,fuy037,43,2019,1952doi:10.1093/femsre/fuy037Advance Access Publication Date:18 October 2018Review ArticleREVIEW ARTICLEConservation and diversity of radiation and oxidativestress resistance mechanisms in Deinococcus speciesSangyong Lim1,Jong-Hyun Jung1,Laurence Blanchard2and Arjan de Groot2,1Biotechnology Research Division,Korea Atomic Energy Research Institute,Jeongeup 56212,Republic of Koreaand2Aix Marseille Univ,CEA,CNRS,BIAM,Saint Paul-Lez-Durance,FranceCorresponding author:DRF/BIAM/LBC,UMR 7265 CEA-CNRS-AMU CEA,Cadarache,13108 Saint Paul Lez Durance,France.Tel:+33 04 42 25 39 35;Fax:+33 04 42 25 47 01;E-mail:nicolaas.degrootcea.frOne sentence summary:The authors reviewed the mechanisms and factors involved in the extreme radiation and oxidative stress resistance inDeinococcus radiodurans in comparison with 10 other resistant Deinococcus species,and highlighted not only conserved pathways but also a large diversityof the repair,protection and regulation toolbox among the different deinococci.Editor:Kenn GerdesArjan de Groot,http:/orcid.org/0000-0003-2202-5279ABSTRACTDeinococcus bacteria are famous for their extreme resistance to ionising radiation and other DNA damage-and oxidativestress-generating agents.More than a hundred genes have been reported to contribute to resistance to radiation,desiccation and/or oxidative stress in Deinococcus radiodurans.These encode proteins involved in DNA repair,oxidativestress defence,regulation and proteins of yet unknown function or with an extracytoplasmic location.Here,we analysedthe conservation of radiation resistance-associated proteins in other radiation-resistant Deinococcus species.Strikingly,homologues of dozens of these proteins are absent in one or more Deinococcus species.For example,only a fewDeinococcus-specific proteins and radiation resistance-associated regulatory proteins are present in each Deinococcus,notablythe metallopeptidase/repressor pair IrrE/DdrO that controls the radiation/desiccation response regulon.Inversely,someDeinococcus species possess proteins that D.radiodurans lacks,including DNA repair proteins consisting of novel domaincombinations,translesion polymerases,additional metalloregulators,redox-sensitive regulator SoxR andmanganese-containing catalase.Moreover,the comparisons improved the characterisation of several proteins regardingimportant conserved residues,cellular location and possible proteinprotein interactions.This comprehensive analysisindicates not only conservation but also large diversity in the molecular mechanisms involved in radiation resistance evenwithin the Deinococcus genus.Keywords:DNA repair;oxidative damage protection;regulatory proteins;stress response;metal homeostatis;biodiversityINTRODUCTIONIn 1956,scientists described a bacterium that was found as acontaminant in a can of ground meat.This bacterium had sur-vived exposure to a high dose of ionising radiation(IR)that wassupposed to sterilise the canned meat(Anderson et al.1956).Now known as Deinococcus radiodurans,this bacterial species isnot only extremely tolerant to gamma radiation,but also toother DNA damage-and oxidative stress-generating conditionssuch as UV and desiccation(Battista 1997).Exposure to highdoses of IR generates massive DNA damage,including hundredsof double-strand breaks,but D.radiodurans is able to reconstituteits genome completely within hours after irradiation.Therefore,Received:28 July 2018;Accepted:17 October 2018C?FEMS 2018.This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License(http:/creativecommons.org/licenses/by-nc/4.0/),which permits non-commercial re-use,distribution,and reproduction in any medium,provided theoriginal work is properly cited.For commercial re-use,please contact 1920FEMS Microbiology Reviews,2019,Vol.43,No.1D.radiodurans is a good model organism to study DNA repair,DNA damage and oxidative stress response,and radiation resis-tance.Deinococcus radiodurans and other Deinococcus species show noloss of viability after exposure to IR doses up to 5 kGy.For com-parion,a few hundred Gy will kill most known bacterial species,including Escherichia coli and Thermus thermophilus,and 510 Gyare lethal to most vertebrates,including humans(Daly 2012).Nevertheless,IR resistance is not unique to Deinococcus,and sev-eral organisms tolerating more than 1 kGy have been described,including not only bacteria(e.g.Chroococcidiopsis of the phylumCyanobacteria)and archaea(e.g.Thermococcus gammatolerans),butalso some small eukaryotes(e.g.tardigrades and bdelloid ro-tifers)(Cox and Battista 2005;Daly 2012).Of these IR-resistantspecies,D.radiodurans has been studied most extensively,whichwas accelerated after obtaining its genome sequence(Whiteet al.1999)and by the development of techniques for its geneticmanipulation.Characterisation of the mechanisms underlyingIR resistance in Deinococcus is also useful to understand IR resis-tance,or sensitivity,in other organisms.The various in vivo and in vitro approaches used in recentyears to study D.radiodurans have indicated that its toleranceto radiation,desiccation and oxidative stress results from acombination of different physiological determinants and well-regulated molecular mechanisms(Fig.1)(Cox and Battista 2005;Confalonieri and Sommer 2011;Slade and Radman 2011;Daly2012;Agapov and Kulbachinskiy 2015;Timmins and Moe 2016).Compared to radiation-sensitive species such as E.coli,pro-teins in D.radiodurans and other radiation-resistant organismsare much better protected against oxidative damage(Daly et al.2007,2010;Krisko and Radman 2010).Radiation and desiccationlead to generation of reactive oxygen species(ROS),but D.ra-diodurans has developed efficient enzymatic and non-enzymaticantioxidant systems to remove ROS and limit protein damage.Sufficient proteome protection is crucial for survival after irradi-ation because protein activity is required for essential processesincluding transcription,translation and DNA repair.Comparedto IR-sensitive bacteria,the nucleoid of Deinococcus species ap-pears more condensed,which may contribute to radiation re-sistance by limiting diffusion of DNA fragments(Levin-Zaidmanet al.2003;Zimmerman and Battista 2005).Following exposureto IR or desiccation,the expression of many genes and proteinsis induced in D.radiodurans,including DNA repair proteins andproteins of yet unknown function(Liu et al.2003;Tanaka et al.2004;Lu et al.2009;Basu and Apte 2012),and several regulatorproteins involved in the radiation or oxidative stress responsehave been described(Agapov and Kulbachinskiy 2015).Deinococcus radiodurans was the first species of the genusDeinococcus that was isolated,and was also the first Deinococcusspecies for which the genome sequence was thoroughly anal-ysed(White et al.1999;Makarova et al.2001).Deinococus bacteriaare ubiquitous in nature and have been isolated from variousFigure 1.Extreme radiation and oxidative stress resistance in Deinococcus in-volves multiple factors and well-regulated mechanisms.environments and locations(e.g.hot and cold desert soil,air,high atmosphere,water).At present,more than 50 radiation-resistant Deinococcus species have been described,and for someof these a complete or draft genome sequence has been ob-tained.Here,we review the reported data about the mecha-nisms involved in radiation resistance,oxidative stress defence,DNA repair,and in their regulation in D.radiodurans.The con-servation of the proteins involved in these processes was in-vestigated in the 10 other radiation-resistant Deinococcus speciesfor which a complete and assembled genome sequence wasavailable.The 11 analysed Deinococcus species have been iso-lated from various locations worldwide(Table 1).This compari-son showed a remarkable diversity of the radiation resistance-associated proteins among deinococci.Furthermore,sequenceanalysis improved the characterisation of several of these pro-teins.Throughout this article we discuss our findings regardingprotein functions and resistance-associated mechanisms in thegenus Deinococcus.DEINOCOCCUS RADIODURANS MUTANTSAFFECTED IN RADIATION AND OXIDATIVESTRESS RESISTANCEMorethanahundredradiation-and/oroxidativestress-sensitive mutant strains of D.radiodurans have been describedin numerous studies(Table S1,Supporting Information).Aschematic overview of proteins required for radiation and ox-idative stress resistance is shown in Fig.2.Many of the mu-tants were obtained after deleting or disrupting a specific genethat was selected because of its expected or possible role inDNA repair,oxidative stress defence and regulation of radiation-resistance-associated genes,or because of its radiation-inducedexpression.Other mutants were obtained after chemical ortransposon mutagenesis followed by screening for increased ra-diation sensitivity.Only a few mutant strains were found to be very sensitive toIR,showing a strong decrease in survival after exposure to rel-atively low doses(5 kGy),orwere found only slightly IR sensitive with less than 10-fold de-creased survival compared to the wild type at the highest dosetested.Several gene mutant strains have been characterised by twoor more research teams,and,remarkably,the reported resultsare sometimes rather different,with mutants found sensitive toIR or other agents in one study but resistant in another study(fordetails,see legend of Table S1,Supporting Information).Suchcontrasting results may be due to differences in the bacterialstrains used and/or in the experimental procedures.Lim et al.21Table 1.Information of complete genomes of Deinococcus species.SpeciesIdentified inTotal genome size(Mb)Replicons(sizes in kb)ProteinsReferencesDeinococcus radiodurans(Drad)Canned meat,USA3.284(2649,412,177,46)3167Anderson et al.(1956);Brooks and Murray(1981);White et al.(1999)Deinococcus geothermalis(Dgeo)Hot spring,Italy3.253(2467,574,206)3003Ferreira et al.(1997);Makarova et al.(2007)Deinococcus deserti(Ddes)Sahara Desert sand,Morocco/Tunisia3.864(2820,325,314,396)3503de Groot et al.(2005,2009)Deinococcus maricopensis(Dmar)Sonoran Desert soil,USA3.51(3499)3242Rainey et al.(2005);Pukallet al.(2011)Deinococcus gobiensis(Dgob)Gobi Desert sand,China4.417(3137,433,425,232,72,55,53)4140Yuan et al.(2009,2012)Deinococcus proteolyticus(Dpro)Lama glama feces,Japan2.895(2147,315,196,132,97)2645Kobatake,Tanabe andHasegawa(1973);Brooksand Murray(1981);Copeland et al.(2012)Deinococcus peraridilitoris(Dper)Coastal desert soil,Chile4.513(3882,557,75)4223Rainey et al.(2007)Deinococcus swuensis(Dswu)Mountain soil,SouthKorea3.531(3531)3217Lee et al.(2013)Deinococcus soli(Dsol)Rice field soil,SouthKorea3.241(3237)3055Cha et al.(2014);Joo et al.(2015)Deinococcus actinosclerus(Dact)Rocky hillside soil,South Korea3.261(3264)3073Joo et al.(2016);Kim et al.(2016)Deinococcus puniceus(Dpun)Mountain soil,SouthKorea2.971(2972)2681Lee et al.(2015)The species name is followed by an abbreviation that is used in Tables 2 to 6.Different results have also been reported with respect toobtaining mutant strains:it appeared to be impossible to ob-tain a recJ(DR 1126,single-stranded-DNA-specific exonuclease)or gyrA(DR 1913,DNA gyrase subunit A)null mutant in oneor two studies(Nguyen et al.2009;Bentchikou et al.2010;Cao,Mueller and Julin 2010),suggesting that these genes are es-sential for viability,whereas others successfully obtained nullmutants for these genes(Jiao et al.2012;Kota,Charaka andMisra 2014).Besides for the naturally transformable D.radiodurans,ge-netic tools allowing construction of mutant strains have alsobeen developed for D.deserti and D.geothermalis.Like in D.ra-diodurans,a D.deserti irrE mutant is highly sensitive to gammaand UV radiation(Vujicic-Zagar et al.2009).Deletion of the chro-mosomal recA gene in D.deserti,the third and last gene of anoperon equivalent to that in D.radiodurans,did not lead to radia-tion sensitivity due to the presence of two additional recA geneslocated on large plasmids(Dulermo et al.2009).A D.geothermaliscystine ABC transporter mutant showed increased sensitivity toH2O2(Kim et al.2017).One might expect that genes important for radiation resis-tance in D.radiodurans be conserved in other radiation-resistantspecies within the genus Deinococcus.To investigate this,notonly homologues of the gene products listed in Table S1(Sup-porting Information)but also other proteins involved in radi-ation resistance-associated processes such as DNA repair andoxidative stress defence(Tables S2S6,Supporting Information)were searched in 10 other complete and assembled Deinococcusgenome sequences(Table 1).Besides showing presence or ab-sence of protein homologues,we included a comparative anal-ysis of domain composition in multidomain proteins and offunctionally important residues in proteins.The results are de-scribed in the following sections.DNA REPAIR IN DEINOCOCCUSDeinococcus radiodurans DNA repair proteins andcomparison with E.coliThe genome sequence of D.radiodurans revealed the presenceof homologues of most well-known prokaryotic DNA repair pro-teins involved in base excision repair(BER),nucleotide excisionrepair(NER),mismatch repair(MMR)and recombinational re-pair,suggesting that the DNA repair machinery of D.radiodu-rans is globally similar to that of other bacteria(Makarova et al.2001),but that it functions more efficiently than in radiation-sensitive species because of better protection of the(DNA repair)proteins against oxidative damage(Daly 2012).Indeed,at leastsome DNA repair proteins of E.coli,namely PolA(Gutman,Fuchsand Minton 1994),RadA(Zhou et al.2006)and UvrA(Agostini,Carroll and Minton 1996),can functionally substitute for theircounterparts in D.radiodurans.However,genetic,biochemical and structural studies haveshown that several other classical DNA repair proteins fromD.radiodurans have characteristics different from their E.colicounterparts.Concerning recombinational repair,E.coli recA(Schlesinger 2007)and recO(Xu et al.2008)only partially com-plement the corresponding gene deletion in D.radiodurans.IncontrasttoE.coliRecA,purifiedD.radioduransRecApreferentiallybindstodouble-strandedDNAwhenalsosingle-strandedDNAispresent in the solution,and initiates DNA-strand exchange pri-marily from the double-stranded DNA(Kim and Cox 2002;Kim22FEMS Microbiology Reviews,2019,Vol.43,No.1Figure 2.Schematic overview of ionising radiation and oxidative stress resistance-associated proteins in D.radiodurans.Many D.radiodurans gene deletion or disruptionmutants with more than 10-fold increased sensitivity compared to the wild-type strain have been described(Table S1,Supporting Information),and the correspondingproteins are indicated in the figure.Red box,ionising radiation sensitive;green box,oxidative stress sensitive;blue box,ionising radiation and oxidative stress sensitive.et al.2002).Such inverse DNA-strand exchange pathway has alsobeen observed for D.geothermalis RecA in one biochemical study(Sghaieretal.2010)butnot