miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響

2022/12/101miR396affectsmycorrhizationandrootmeristemactivityin

thelegumeMedicagotruncatulaThePlantJournal(2013)doi:10.1111/tpj.12178miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第1頁!2022/12/102主要內(nèi)容INTRODUCTIONRESULTS

1、Mtr-miR396aandmtr-miR396bgenesshowdifferenttissue-specificexpressionprofilesinMedicagotruncatularoots2、InM.truncatularoots,miR396regulatessixGRFandtwobHLH79transcriptionfactors3、CharacterizationofthemiR396regulatorynetworkinM.truncatulamiR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第2頁!4、miR396negativelyaffectsrootgrowth,RAMsizeandcellproliferationintherootapex5、SilencingofMtGRFgenesmimicsthemiR396-OErootgrowthphenotype6、HormonalcontrolofmiR396expression7、miR396limitsmycorrhizalcolonizationbutnotnodulation2022/12/103miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第3頁!Inrecentyears,microRNAs(miRNAs)haveemergedasimportantregulatorsofthearchitectureoftherootsysteminArabidopsisthaliana,miR165/166governstheradialpatterningofvascularandpericycletissuesviaquantitativerepressionofHD-ZIPIIItranscriptionfactorsAsetofauxin-relatedmiRNAs,includingmiR160,alsoregulatelateralrootinitiationand/orrootgrowth2022/12/104miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第4頁!InA.thaliana,miR396isencodedbytwoloci,post-transcriptionallyrepressesseveralmembersofthegrowth-regulatingfactor(GRF)family.Theseplant-specificTFscontrolthegrowthanddevelopmentofleavesandstems.miR396mayhaveevolvedtoplayrolesinvariousregulatorynetworksinplants.Inthispaper,wedescribetherolesofthemiR396regulatorynetworkinM.truncatularoots.2022/12/105miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第5頁!2022/12/106miR396expressioninorgansandroottips兩種miRNAs在整株植物中都有表達，其中miR396b在根、瘤、葉和豆莢中的表達量較高，miR396a在根尖中較多miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第6頁!2022/12/1072、InM.truncatularoots,miR396regulatessixGRFandtwobHLH79transcriptionfactorsWefoundreadscorrespondingtothemiR396-relatedcleavageproductsofsixGRFmRNAs(MtGRF1,MtGi10-TC183867;MtGRF2,MtGi10-TC183494;MtGRF3,MtGi10BG454006;MtGRF4,Medtr5g027250;MtGRF5,Medtr8g020560;MtGRF6,Medtr7g126820).ThesetranscriptspossessaputativemiR396bindingsite,andtheencodedproteinsallcontaintheconserveddomainsofGRFsmiR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第7頁!2022/12/108InM.truncatula,sixMtGRFandtwobHLH79geneshavemiR396bindingsitesmiR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第8頁!2022/12/109LevelsoftranscriptsofmiR396targetsweremeasuredusingquantitativeRT-PCRinmiR396-OErootsMtGRF-mRNA水平和MtbHLH79轉(zhuǎn)錄水平都降低，說明miR396可能抑制這些基因的表達水平miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第9頁!2022/12/1010LevelsoftranscriptsofmiR396targetsweremeasuredusingquantitativeRT-PCRinrootsinactivatedformiR396activity(MIM396)除了GRF6，其他基因表達水平都上升，這里還不能排除有其他因子對GRF6進行調(diào)節(jié)的可能，本文中沒有深入研究miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第10頁!2022/12/1011miR396-OE中主根長度和干重明顯減少，而MIM396中根長稍微增加，而干重卻大量增加。說明miR396過表達能抑制根的生長miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第11頁!2022/12/1012兩者之間沒有大的變化，說明miR396對細胞從細胞周期進入分裂周期的開關(guān)沒有大的影響miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第12頁!Together,thesedatastronglysuggestthatmiR396-dependentmodulationofrootgrowthmayessentiallybeduetoarestrictionofcelldivisionactivity.2022/12/1013miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第13頁!2022/12/1014SilencingofMtGRFgenesmimicsmiR396over-expressioninroots.iGRF表達減少而ibHLH79表達不變，說明很可能是miR396對GRF基因的負調(diào)控使得miR396-OE和MIM396植株中根的生長表型不同miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第14頁!2022/12/1015Gibberellin,cytokininandauxintreatmentsdidnotaffectexpressionofanyofthesegenesmiR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第15頁!2022/12/1016LevelsofGUStranscriptinrootsexpressingtheprom-miR396b:GUSconstruct(greybars)ortheprom-GRF5:GUSconstruct(blackbars)treatedornottreatedwith1nMBRfor3hweremeasuredusingquantitativeRT-PCR.BR可以活化miR396b的啟動子活性，但prom-GRF5:GUS并沒有明顯減少，說明BR可能是在轉(zhuǎn)錄后水平抑制GRF5基因的表達，而且BR和miR396b存在潛在聯(lián)系，但本文中沒有對其研究miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第16頁!2022/12/1017miR396a啟動子首先在根瘤導管組織中發(fā)現(xiàn)，miR396b啟動子在維管組織，感染區(qū)（Ⅱ）和氮固定區(qū)（Ⅲ）含量很高miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第17頁!WealsoanalysedtheexpressionofmiR396inresponsetoMyc-LCOs,thelipochitooligosaccharidesignalmoleculesproducedbyAMfungi.TheinvolvementofmiR396duringmycorrhizationwasthenanalysedat5weeksaftercolonizationwithR.irregularis(formerlyGlomusintraradices)inmiR396-OEandMIM396roots.2022/12/1018miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第18頁!Here,weshowedthat,inthemodellegumeM.truncatula,miR396repressesnotonlyMtGRFbutalsotwoMtbHLH79TF-encodinggenes,andthatitsde-regulationaffectsrootgrowthandtheabilitytobecolonizedbymycorrhizalfungi.2022/12/1019miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第19頁!INTRODUCTIONInplants,rootgrowthiscontrolledviacoordinatedcelldivisionandexpansionattherootapex,wherethreeregionsareobserved:(i)themeristematiczone,(ii)theelongationzone,and(iii)thedifferentiationzoneRootdevelopmentisregulatedbyplexinteractionsofhormonesignallingpathways2022/12/1020miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第20頁!However,fewmiRNAshavebeenreportedtoplayrolesinRAMfunctionAlthoughlargesetsofmycorrhizationornodulation-responsivemiRNAshavebeenidentifiedinlegumes,fewhavebeenfunctionallylinkedtotheseprocesses.2022/12/1021miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第21頁!RESULTS1、Mtr-miR396aandmtr-miR396bgenesshowdifferenttissue-specificexpressionprofilesinMedicagotruncatularootsMtr-miR396a和mtr-miR396b在根中的表達具有空間特異性2022/12/1022miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第22頁!2022/12/1023兩者在根中的表達在根尖橫切面中的表達在根尖中的表達在側(cè)根中的表達miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第23頁!Wealsoobservedabundantdegradomereadscorrespondingtospecificcleavageoftwonon-GRFtranscriptsatmiR396bindingsites,BasedontBLASTXanalysis,the2predictedproteinsshowedhighsimilarity(43and46%respectively)toAtbHLH79/BIGPETAL(At1g59640),aTFinvolvedinthecontrolofpetalgrowthinA.thaliana.2022/12/1024miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第24頁!3、CharacterizationofthemiR396regulatorynetworkinM.truncatulaToinvestigatemiR396-mediatedgeneregulationinplanta,weover-expressedmtr-miR396aandmtr-miR396bprecursorsinM.truncatularoots(miR396-OE).2022/12/1025左圖說明OE突變體能成功過量表達成熟的miRNAsmiR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第25頁!Tofurthervalidatetheirtargets,wegeneratedrootsexpressingatargetmimicryconstructthatwassuccessfullydesignedtohindermiR396activityinA.thaliana.Mimicry(MIM)transcriptsspecificallytrapmembersofanmiRNAfamily,andthuspreventtheiractivityonendogenoustargets2022/12/1026miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第26頁!4、miR396negativelyaffectsrootgrowth,RAMsizeandcellproliferationintherootapexToinvestigatethefunctionofmiR396inroots,wemeasuredthelengthanddryweightofrootsthateitheroverexpressedthemtr-miR396bprecursor(miR396-OE)orwereinactivatedformiR396activity(MIM396).2022/12/1027miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第27頁!2022/12/1028RootmeristemsizeisreducedinmiR396-OEpositeplantsmiR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第28頁!2022/12/1029Theexpressionofmitoticcellcycle(CYCB1;1,CYCB1;3,CYCB2;1,CYCD3;1,histoneH4,CDC16)orendocycle(CCS52B)markerswasmeasuredusingquantitativeRT-PCRinmiR396-OEandcontrolrootsThepercentageofcellsreplicating(i.e.EdU-positivenuclei)obtainedfrom20roottipsperconstructinfourindependentexperimentsmiR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第29頁!5、SilencingofMtGRFgenesmimicsthemiR396-OErootgrowthphenotypeTostudytheroleofthemiR396targetsonrootgrowth,weinactivatedthemusinganRNAistrategy.First,wegeneratedpositeplantsexpressingthreeRNAiconstructstargetingaconservedregionofvariousGRFgenes(MtGRF2,MtGRF4andMtGRF6).Inaddition,wepreparedanRNAivectortargetingbothMtbHLH79genesandconfirmedtheirefficientsilencing2022/12/1030miR396在豆科植物苜蓿根中對其菌根化和分生組織活性的影響共36頁，您現(xiàn)在瀏覽的是第30頁!6、HormonalcontrolofmiR396expressionWetreatedwild-typeplantsfor1,3or5hwithauxin,cytokinin,gibberellin,abscisicacidorbrassinosteroids(BR),andmeasuredpre-miR396andtargetmRNAlev