植物学及园艺学英文

EFFECTSOFETHYLENEFruitRipening

Abscission;leafflowerfruits(thinning,harvesting)HookClosureMaintenanceInitiatesGerminationinGrainsActivatesdormantbuds(potatoesinstorage)Stemelongationindeep-waterriceInducesFloweringinPineapplePromotesFemaleExpressioninFlowersFlowerandLeafSenescence:Agpreventative(vaselife)

Ethylene第1页/共20页第一页，共21页。1.Introduction●Hormonesactaschemicalmessengersinthecontrolofthemolecular,biochemical,andphysiological

eventsunderlyinggrowthanddevelopment.●Hormonesalsoserveasessentialintegratorsofdevelopmentalprogramswiththeenvironmentalsignals.Abasicchallengeinbiologyis,therefore,tounderstandthemolecularmechanismsthatunderliehormoneaction,inotherwords,howthosechemicalsignalsaresensedbyandcommunicatedwithinthecellstotriggertherelevantresponses.●Thisprocesscanbedividedintothreemainsteps:(1)signalperception,(2)signaltransductionoracascadeofbiochemicaleventsthatultimatelyleadstotheinductionofthefinalstep,(3)theresponse.●Amongtheplanthormones,ethylenedistinguishesitselfbyitssimplehydrocarbonchemicalstructure(C2H4)anditsgaseousnature.Thissimplemolecule,however,playsamajorroleinplantgrowthanddevelopmentbyinfluencingawiderangeofcomplexphysiologicalprocessesthroughouttheentireplantlifecycle,fromseedgerminationtoflowering,fruitripening,andsenescence第2页/共20页第二页，共21页。●Oneofthemostdramaticeffectsofethyleneonplantmorphogenesisistheclassical‘‘tripleresponse’’exhibitedbydark-grownseedlingsexposedtoethylene.ThetripleresponseinArabidopsisischaracterizedby(1)exaggeratedcurvatureoftheapicalhook,(2)radialswellingofthehypocotyl,and(3)inhibitionofhypocotylandrootgrowthFig.1Phenotypesofdark-grownthree-day-oldseedlingsofArabidopsisthaliana.Theplantontheleftwasgrownwithouthormonalsupplementation,whereastheplantontherightwasexposedto10mMethyleneprecursorACCandthusshowsatypicaltripleresponse.●Aseriesofelegantgenetic,molecular,andbiochemicalstudiesareuncoveringalargelylinearpathwaythattransducestheethylenesignalfromtheendoplasmicreticulummembranetothenucleusFig.2Theethylenesignalingpathwayanditsgeneticallycharacterizedcomponents.Thesignalingpathwaycomponentsareshownintheirsequentialorderofaction.Componentsdrawninwhiterepresentactiveforms,whereasgrayovalsrepresenttheirinactiveversions.Bindingofethylenetothereceptors,representedbyETR1,leadstoactivationofethyleneresponses.DottedovalrepresentsEIN3degradationbythe26SproteasomepathwayduetoactionofEBF1andEBF2.Arrowsindicateactivationsteps,whereasablockedarrowdepictsrepressionofdownstreamelementsbyCTR1.●Thisreviewaimstosummarizethecurrentstateofknowledgeintheethylenefield.Wewillstartwiththedescriptionofthemainexperimentalbreakthroughsthatresultedinthediscoveryoftheknownethylenesignalingcomponentstothendescribethegenomicapproachesemployedtocharacterizethemolecularaspectsoftheethyleneresponse.第3页/共20页第三页，共21页。2.Ethyleneperceptionismediatedbyasmallfamilyofreceptors●ThedevelopmentofmodernmoleculargeneticapproachesandtheselectionofArabidopsisasaplantmodelsystemopenedanewdoornotonlyfortheidentificationoftheethylenereceptors,butfortheelucidationoftheentiresignaltransductionpathway.●Thenewquestforthemolecularcomponentsoftheethylenesignaltransductionmachineryfirstledtotheisolationofethyleneresponse1,etr1,adominantmutationthatconfersethyleneinsensitivity.●ETR1encodesahistidinekinasewithsimilaritytotheclassicalbacterialtwo-componenthistidinekinases.Thenovelhydrophobicamino-terminaldomainofETR1heterologouslyexpressedinyeastwasshowntopossesshigh-affinitybindingpropertiestothegasethylene.ETR1wasfoundtoactasadimerthatlocalizestoacellularmembranesystem.●AseriesofelegantstudiesperformedintheninetiesrevealedthatethylenereceptorsareencodedbyasmallgenefamilythatinArabidopsisconsistsoffivemembers:ETR1,ethyleneresponse2(etr2),ethyleneinsensitive4(ein4),ethyleneresistant1(ers1),andethyleneresistant2(ers2).第4页/共20页第四页，共21页。●

Theirsequencesimilarityandstructuralorganization,thefivereceptorsarecategorizedintotwosubfamilies.SubfamilyImembers(ETR1andERS1)harborthreehydrophobictransmembranedomainsintheamino-terminusfollowedbyaconservedhistidinekinasedomain.SubfamilyIImembers(ETR2,ERS2,andEIN4)possessfourpredictedamino-terminalhydrophobictransmembraneregionsfollowedbyalessconservedkinasedomainthatlacksseveralofthecanonicalfeaturesrequiredforhistidine-kinaseactivity.Furthermore,threeofthefivereceptors,ETR1,ETR2,andEIN4,alsopossessacarboxyl-terminalreceiverdomain.●Amajorbreakthroughcamewiththeisolationofloss-offunction(LOF)allelesofthereceptors.EachofthesingleLOFalleleswasstillabletorespondtoethylene,indicatingahighdegreeoffunctionalredundancyamongthereceptors.Furthermore,tripleandquadrupleLOFmutantsdisplayedaconstitutiveethyleneresponseintheabsenceofthehormone.第5页/共20页第五页，共21页。第6页/共20页第六页，共21页。第7页/共20页第七页，共21页。●Bothreceptorsubfamiliesappeartobeabletosenseethylene,asdoubleetr1;ers1andtripleetr2;ers2;ein4LOFmutantsarestillabletorespondtoethyleneinthetripleresponseassay.However,aparticularroleforsubfamilyIreceptorsinlight-grownplantshasbeenrecentlysuggested.Doubleetr1;ers1LOFmutantsdisplayedseverephenotypes,includingminiaturerosettes,fertilitydefects,andalteredflowermorphology.Alloftheseeffectsweredependentonafunctionalethylenesignalingpathway,implyingthattheobservedgrowthdefectsarosefromamisregulationofethyleneresponses.OverexpressionofthesubfamilyIImemberswasunabletorescuetheobservedphenotypes,whereasectopicexpressionofeitherwild-typeETR1orERS1restorednormaldevelopment,furthersupportingthenotionofauniqueroleforthesubfamilyIreceptors.第8页/共20页第八页，共21页。●Ethylenewasfoundtobindtothereceptorsthroughatransitionmetalcofactor,copper.●

Moreover,thecoppercofactorwasshowntobeessentialforethylenebinding,andthus,properreceptorfunction.Thecurrentstructuralmodelfortheethylenebindingdomainsuggeststhatthecopper(I)cofactorislocatedintheelectron-richhydrophobicpocketformedbytheN-terminaltransmembranedomainsofthereceptors.Inparticular,residuesCys65andHis69arethoughttoplayafundamentalroleinthisprotein-metal-hormoneinteraction.●

Inplanta,therelevanceofthisinteractionwasfurtherconfirmedwiththeidentificationofRESPONSIVETOANTAGONIST1(RA\N1).RAN1wasisolatedusingascreeningformutantswithalteredspecificityinhormonebindingbyemployingtheethyleneantagonisttrans-cyclooctene(TCO).ran1plantsaredefectiveinacoppertransportersimilartoP-typeATPases.●Thelackofthemetalcofactorimpairsreceptorfunctioninran1mutantsbycausingalteredligandspecificityandthusrenderingtheplantsresponsivetotheantagonistTCO.Furthermore,astrongLOFran1alleleresultsinaconstitutiveethyleneresponsephenotypeintheabsenceofthegaseoushormone.●

AutophosphorylationactivityhasbeendemonstratedforETR1andallothermembersofthereceptorfamily.However,whileETR1autophosphorylatesinthepredictedconservedhistidineresidue,ERS1andallofthesubfamilyIImembersdisplayaserine-kinaseactivityinvitro.ERS1alsopossessesahistidine-kinaseactivity,whereasitsserineautophosphorylationisthoughtnottobesignificantinvivo.第9页/共20页第九页，共21页。●Anotherinterestingstudyaddressedtherolesofthekinaseactivityandofthecarboxyl-terminalreceiverdomainofETR1,ETR2,andEIN4inethylenesignalingbylookingattheeffectofexpressingtruncatedversionsofETR1inatripleLOFetr1;etr2;ein4mutantbackground.●TransformationofthetriplemutantwithatruncatedversionofETR1thatlacksboththehistidine-kinaseandthereceiverdomainfailedtorestoreethyleneresponsivenesstothetriplemutant.Conversely,asecondtruncatedETR1constructinwhichonlythereceiverdomainwasmissingdidrescuethepartialconstitutivetripleresponsephenotypeofthetripleknockoutline.Moreover,transgenicplantsharboringthisparticularconstructdisplayedhypersensitivitytoethylene.Theseobservationsimpliedthatthekinasedomainwasnecessaryforsignaltransmissionbythereceptorsandthatthereceiverdomainwasnotessentialforrestoringethyleneresponsiveness.●

Modulationoftheethyleneresponsesbythereceiverdomaincouldbeachieved,forinstance,throughitsreportedinteractionwithanothernegativeregulatorofthepathway,CONSTITUTIVETRIPLERESPONSE1(CTR1)(seebelow),assumingthatintheabsenceofthereceiverdomainthefunctionofCTR1isimpaired.Interestingly,adetailedkineticanalysisofseedlinggrowthresponseinthepresenceofexogenousethyleneandoftheconsecutiverecoveryafterethylenewithdrawalalsoindicatedtheimportanceof

boththekinaseandreceiverdomainsinspecificaspectsoftheethyleneresponse.●Theseresultssuggestthatboththekinaseandthereceiverdomainareimportantforthehypocotylrecoveryfromshortethylenetreatmentandindicatethat,despitethelargefunctionaloverlap,somespecificitycanbeattributedtothedifferentreceptors.第10页/共20页第十页，共21页。●Theaccumulateddatahaveprovidedamodelfortheroleofthereceptorsinethylenesignaling.(1)Intheabsenceofthehormone,thereceptorsactivelyrepressdownstreamcomponentsofthepathwayandinhibitethyleneresponses.

(2)Ethylenegasbindstoallfivereceptors,causingthereceptorstobecomeinactiveandreleasingthepathwayfromtheirrepression.Althoughstillcontroversial,(3)receptorkinaseactivitypossiblyplaysanimportantroleinsignaling.3.ARaf-likekinaseactsdownstreamofthereceptorsrepressingethyleneresponses●Asubsetofthosewasfurthercharacterizedasdefectiveinethylenebiosynthesis,namelyethyleneoverproducer(eto).Conversely,thectr1mutantdidnotrespondtoinhibitorsofethylenebiosynthesisindicativeofanalterationinsignaltransduction.CTR1encodesaserine/threoninekinasewhosecarboxyl-terminussharessequencesimilaritywiththeRaffamilyofproteinkinases.TheconstitutiveresponsephenotypeexhibitedbyLOFmutantsindicatesthatCTR1negativelyregulatestheethylenesignalingpathway.●Interestingly,CTR1wasshowntophysicallyinteractwiththereceptors.Subsequently,CTR1wasshowntoco-localizewiththereceptorsattheERmembranes.Moreover,co-purificationofanaffinity-taggedCTR1andendogenousETR1intransgenicArabidopsislinesstronglysupportedtheinvivointeractionoftheseproteins.ThesignificanceofthisinteractioninsignaltransmissioniscorroboratedbytheobservationthatdoubleandtripleLOFreceptormutantsresultinalossofER-localizationofCTR1.Takentogether,thesedatasuggestedthatthereceptorsandCTR1functionaspartofanER-localizedcomplexthatactivelyrepressesethyleneresponses.第11页/共20页第十一页，共21页。●Theserine/threoninekinaseactivityofCTR1wasdemonstratedinvitroandshowntobeessentialforproperfunctioningofthereceptors/CTR1signalingcomplex.IthasbeenestablishedthatethylenebindingtothereceptorsaffectsneithertheinteractionbetweenthereceptorsandCTR1northeirsub-cellularlocalization.Thecurrentproposedmodelpredictsthatuponethylenebinding,thereceptors/CTR1signalingcomplexesareturned‘‘off’’.4.Auniqueplantproteinisacentralcomponentofthesignalingpathwayandpositivelyregulatesethyleneresponses●Downstreamofthereceptors/CTR1complexesthereactsapositiveregulatorofthepathway,ETHYLENEINSENSITIVE2(EIN2).EIN2isrequiredforallethyleneresponsesstudiedandconstitutesacriticalstepinthesignaltransduction.Thehydrophobicamino-terminusispredictedtoformtwelvetransmembranedomainsandsharessequencesimilaritywiththefamilyofNRAMPmetaliontransporters.●ThelongEIN2carboxyl-terminuscontainsacoiled-coilstructure(amotiftypicallyinvolvedinprotein–proteininteractions)butotherwisedisplaysnosimilaritytoknownprotein.Conversely,theNRAMP-likeamino-terminaldomainofEIN2isbelievedto‘‘sense’’theupstreamsignalingevents.第12页/共20页第十二页，共21页。5.Atranscriptionalcascademediatesethyleneresponsesatthegeneexpressionlevel●

ETHYLENEINSENSITIVE3,EIN3,isanuclear-localizedproteinrequiredforethylenesignalingthatgeneticallyworksdownstreamofEIN2.EIN3belongstoasmallgenefamilythatinArabidopsisalsoincludesfiveEIN3-LIKE(EIL)proteins.TheseobservationssuggestedthatEIN3andatleastsomeoftheEILsmayactastranscriptionalregulatorsofethyleneresponses.第13页/共20页第十三页，共21页。●InArabidopsis,EIN3,EIL1,andEIL2weredemonstratedtobindtoashortpalindromicregion,knownastheEIN3-bindingsite,orEBS,inthepromoteroftheEREBPfamilymemberETHYLENERESPONSEFACTOR1(ERF1).ERF1,inturn,isaGCCbox-bindingtranscriptionfactorthatactsdownstreamofEIN3andEILsandisresponsibleforthemodulationofasetofsecondaryethyleneresponsivegenes.Consistentwiththeseresults,transgeniclinesoverexpressingEIN3showaconstitutivetripleresponse,whereasinERF1-overexpressingplantsonlysomeoftheethyleneresponsesareactivated.●ERF1isalsorequiredfortheactivationofdefense-relatedresponsesbytheplanthormonejasmonicacid.Moreover,bothethyleneandjasmonatepathways

mustbeintactforproperERF1expression.EthyleneandjasmonateinteractsynergisticallytoachievemaximalexpressionofERF1.Thistranscriptionfactor,therefore,representsacriticalelementintheinteractionbetweenthesetwoimportanthormones.6.Theubiquitin/26SproteasomepathwayregulatesEIN3activity●ArecentlyemergingpicturepositionstheSCF-E3ligasesinthecenterofmanyplanthormonesignalingpathways,includingthoseofauxin,gibberellins,andjasmonicacid.●TheethylenesignalingpathwayalsoemploystheSCF/26Sproteasometoregulatethelevelsofatleastoneofitscomponents,EIN3.Recently,twoF-boxproteins,EIN3BINDINGFACTOR1(EBF1)andEBF2,wereshowntoactaspartofanE3-ligaseandtobindandtargettheconstitutivelyproducedEIN3fordegradationintheabsenceofethylene.●Inthepresenceofexogenousethylene,EIN3proteinlevelswerefoundtoincrease.第14页/共20页第十四页，共21页。●InadditiontothenegativeregulationbyEBF1andEBF2,astimulatorymechanismthatactivatesthestabilizedEIN3proteinisalsoexpectedtoparticipateintheregulationoftheactivityofthiscriticaltranscriptionfactor.ThisobservationissupportedbytheethyleneresponsivenessofEIN3-overexpressionlinesandebf1;ebf2doubleLOFmutants.TheseplantsrespondtoexogenousethyleneinspiteofhavingelevatedlevelsofEIN3,suggestingthatregulationoftheproteinlevelsisnottheonlymechanismthatgovernstheEIN3activity.7.Ethylenetriggersgenome-widechangesingeneexpression●EIN3istheinitialfactorinthetranscriptionalcascaderesponsibleforinducingethyleneresponsesatthemolecularlevel.Theimportanceofthistranscriptionalregulationinethyleneresponsesisemphasizedbytheobservationthatein3;eil1doublemutantsdisplayalmostcompleteinsensitivitytotheethylenegas.Inaddition,regulationofEIN3bytheSCF/26Sproteasomepathwayservesasanefficientmechanismforthepromptandfine-tunedcontrolofthistranscriptionalcascade.第15页/共20页第十五页，共21页。●

Inordertogainabetterunderstandingoftheethyleneeffectsintranscription,severalresearchgroupshaveperformedgenome-scalestudiesusingmicroarraysandotherhigh-throughputtechniques.●EarlystudiesreliedoncDNAmicroarrayscoveringonlyafractionofthegenome,andthusSchenkandcoworkersanalyzedtheexpressionof2375ESTsenrichedinputativedefense-relatedgenes.Interestingly,theauthorsevaluatedthepatternofgeneexpressioninresponsenotonlytoethylene,butalsotoafungalpathogen,salicylicacid,andjasmonate.Ahighlevel(50%)ofcoordinatedgeneexpressionwasobservedfortheethyleneandjasmonatetreatments.●ThisinitialstudywasfollowedbyabroaderanalysisofethyleneresponsesrelyingonanESTmicroarrayrepresenting6000uniquegenes.Theauthorsalsotookadvantageoftheethylenemutantsetr1-1andctr1-1ascontrols.Thecomparativeanalysisoftheprofilesidentifiedapproximately7%ofgenesasethylene-regulated.Ethylenewasalsofoundtochangeexpressionlevelsofknownjasmonate-andauxin-regulatedgenes,indicativeoftheinteractionbetweenethyleneandthesehormones.●Ahierarchicalclusteranalysisofthedatarevealeddistinctsubsetsofgeneswithcorrelatedpatternsofexpression.Theseincluded:(1)agroupofveryearly,transientlyinduciblegenes(after10to30minofthebeginningofthetreatment),

(2)anearly-induciblesubset(30minto1h),

(3)agroupofgenesup-regulatedattheintermediate(between1–6hofethyleneexposure)time-points,and(4)agroupconsistingoflateinduciblegenes(at6hoftreatment).Alargenumberofclonesalsodisplayedhigherexpressionlevelsintheein2-1ethyleneinsensitivebackgroundandweredown-regulatedinwild-typeethylene-treatedplants.第16页/共20页第十六页，共21页。●OneofthenoveltiesinthestudybyDePaepeandcoworkers55wasthediscoveryofanethylene-mediatedregulationofthegenesinvolvedintheubiquitin/26Sproteasomepathway.TheseresultswerefurtherconfirmedbyRT-PCRanalysis,whichrevealedthreeubiquitin-specificproteasesandanubiquitin-conjugatingenzymeasbeingup-regulatedbytheethylenetreatment.Itispossiblethatthesegenesworkincoordinationwiththewell-characterizedEBF1andEBF2.●Thephysiologicalsignificanceoftheinteractionbetweenethyleneandglucose,aswellastheinplantaroleofEIN3inthecrosstalkofthesetwosignals,awaitsfurtherexperimentalconfirmation.●Mostimportantly,hormonecrosstalkhasalsoemergedasaprominentthemeintheseanalyses,particularly,theinteractionbetweenethyleneandjasmonateinplantdefenseresponsesleadingtotheidentificationofseveralcommontargetgenes.●Inadditiontoco-regulatinganumberoftargetgenes,theethylene-jasmonateandethylene-auxininteractionswerealsosuggestedtotakep