Poor transcript‐protein correlation in the brain negatively correlating gene products reveal neuronal polarity as a potential cause Christian P. Moritz.pdf

该【Poor transcript‐protein correlation in the brain negatively correlating gene products reveal neuronal polarity as a potential cause Christian P. Moritz 】是由【小舍儿】上传分享，文档一共【38】页，该文档可以免费在线阅读，需要了解更多关于【Poor transcript‐protein correlation in the brain negatively correlating gene products reveal neuronal polarity as a potential cause Christian P. 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】的内容，可以使用淘豆网的站内搜索功能，选择自己适合的文档，以下文字是截取该文章内的部分文字，如需要获得完整电子版，请下载此文档到您的设备，方便您编辑和打印。:..(OrcidID:0000-0002-9294-2323)DR.(OrcidID:0000-0003-1928-9441)Articletype:OriginalArticleRegularresearchpaper(JNC-2018-0496)ResubmittedtoJNeurochemPoortranscript-proteincorrelationinthebrain:*,TimoMühlhausc,StefanTenzerd,ae,EckhardFriauf(/0000-0002-1833-1698)aaAnimalPhysiologyGroup,DepartmentofBiology,UniversityofKaiserslautern,Erwin-Schr?dinger-Stra?e13,67663Kaiserslautern,GermanybSynaptopathiesandAutoantibodies,RSUMR5310,FacultyofMedicine,,10ruedeMarandière,42270Saint-étienne,FrancecComputationalSystemsBiology,DepartmentofBiology,UniversityofKaiserslautern,Erwin-Schr?dinger-Stra?e56,67663Kaiserslautern,GermanydInstituteofImmunology,UniversityMedicalCenteroftheJohannesGutenbergUniversityMainz,,55131Mainz,GermanyeDivisionofAllergology,Paul-Ehrlich-Institut,Paul-Ehrlich-Stra?e51-59,63225Langen,GermanyeptedforpublicationandundergonefullpeerreviewbuthasnotAcceptedbeenthroughthecopyediting,typesetting,paginationandproofreadingprocess,:.Allrightsreserved.:..*Correspondenceto:,RSUMR5310,FacultyofMedicine,,10ruedeMarandière,42270Saint-étienne,Francechristian.******@univ-st-(upto45characters):Poortranscript-proteincorrelationinthebrainKeywords(upto6):negativetranscript|proteincorrelation;integrativeomics;neuronalpolarity;proteintransport;transcriptomics;proteomicsAbbreviationsused:CN:plex,CNS:centralnervoussystem,plementaryRNA,CV%:relativecoefficientofvariationArticleDNA:deoxyribonucleicacid,GO:geneontology,IC:inferiorcolliculus,MA(plot):logratioversusmeanaverage(plot),mRNA:messengerRNA,PC:ponent,QI–QIV:quadrantI–quadrantIV,RNA:ribonucleicacid,RoB:restofbrain,ROI:regionofinterest,RRID:ResearchResourceIdentifiersSILAC:StableisotopelabelingbyaminoacidsincellcultureSOC:plexAbstractTranscription,translation,,,,Sregions,.,,weperformedanintegrativeinter-omicsstudyandanalyzedthreeinterconnectedratauditorybrainstemregions(;plex,SOC;inferiorcolliculus,IC)(ROIs)byDNAmicroarraysandlabel-.:..spectrometry,|proteinpairsanddetectedpoortomoderatetranscript|proteincorrelationinallROIs,-(Slc6a11,Syngr1,Tppp),-relatedproteins,-relatedproteinswithpoortranscript|,ouranalysessupportthatproteintransportinpolarcellsisathirdfactorthatinfluencestheproteinleveland,thereby,thetranscript|,translation,andturnoveroftranscripts(mRNA)(,).Mappingexperimentshavedemonstratedapoorcorrelationbetweenthetranscriptomeandthecognateproteomelevel(,,;).Toquantifycorrelationinastatisticalway,thecoefficientofdetermination(R2),rangingfromnocorrelationatall(R2=0)togoodcorrelation(R2?)(R2=,;R2=,Schwanh?,;R2=-,).Therefore,transcriptlevelsareratherproxiesforproteinabundance().Thattranscriptandproteinlevelsofthesamegenedonotalwaysstronglycorrelatemaybeexplainedbyregulationoftranslation(Schwanh?)andproteinstability().AccordingtoSchwanh?usserandcolleagues,regulationoftranslationplaysthemajorrolefordeterminingtheproteinlevel,,notmutuallyexclusiveexplanation,?usserstudy,,evidencedbythefactthatthedistancebetweensomaandaxonterminalsoftenamountstoseveralmillimeters,,manyproteinsencountertransportintotheaxonterminals,which,incaseofprojectionneurons,maybelocatedindistantbrainorspinalcordAcceptedregionsoreventheperipheralmusculature()..:..molecularpolarityof(projection)neuronsresultsinaconsiderablenumberofnon-correlatingandevennegativelycorrelatinggeneproducts(thosewithopposinglevelsoftranscriptandprotein;quadrantsII(QII)).Whendisregardingdifferenthalf-livesofmolecules,translationalregulation,andtransporteffectsinpolarneurons,transcriptandproteinlevelslikelycorrelatestrongly().Transportprocesses,however,thatseparatetheproteinspatiallyfromthetranscript,causepoorcorrelation().Notably,proteintransportaffectsthetranscript|proteincorrelationeneproductsthatarehigherabundantinspecificregions,inthefollowingcalledregion-),becausetheproteinistransportedintoaregionwithlowtranscriptlevelofthecognateprotein,,thetranscript|proteincorrelationofwidelyexpressedgeneproductsshouldbelessaffectedbyproteintransport(),|proteincorrelation,,becauseitplexes(MalmiercaandHackett2010,Schofield2010),),plex(SOC),andtheinferiorcolliculi(ICs).Recently,wehaveidentifiedregion-plexesinaproteomicshotgunapproach().Becauseoftheneuronalpolarity,wehypothesizethatnegativelycorrelatingtranscript||proteinpairs,weanalyzedthetranscriptome,SOC,(RoB)-freequantitativeproteomics().Ithasbeensuggestedtocategorizetranscriptomic|proteomicstudieswiththefollowingkeyconcepts(VogelandMarcotte2012):1)Absoluteconcentrations(amountperunit)(ratiosoftwoabsoluteconcentrations))Rates(ofaprocesssuchastranslation)(relativeorabsoluteamounts).3)Steady-state(changeofgeneproducts)-steady-statesystems(perturbedbyanexternalstimulus,suchasadrug).4),weaddressedrelativetranscriptandproteinconcentrations,steady-stateAccepted(unperturbed)conditions,.:..transcript|proteincorrelationhavefocusedoncellculturessuchasyeast,,wherecellpolarityisvirtuallyabsent(,,).Incontrast,|(TierSchG§4Absatz3)(LandesuntersuchungsamtKoblenz).Weutilized60(+/-3)-day-oldSprague-Dawleyrats(RRID:RGD_1566457)ofbothsexespurchasedfromCharlesRiver(Sulzfeld,Germany).AstheROIswerepreparedfromthesamecohortofrats,,().Inbrief,,theskullwasopened,thebrainwasreleasedandtransferredtoice-coldextracellularsolution(inmM:[,CarlRothGmbH,Karlsruhe,Germany],1MgCl2[A4425,AppliChemGmbH,Darmstadt,Germany],[,CarlRothGmbH],2sodiumpyruvate[A4859,AppliChemGmbH],3myo-inositol[1045070250,MerckKGaA,Darmstadt,Germany],[K3375,Sigma-Aldrich,Darmstadt,Germany],26sodiumbicarbonate[,CarlRothGmbH,260D(+)glucose[A3666,AppliChem],2CaCl2[A4689,AppliChem]).(ponsandmedullaoblongata),300-μm-thickcoronalbrainstemsliceswerecutonavibratome(VT1200S,Leica,Wetzlar,Germany).,,ofwhichrelevantAcceptedmaterialsandmethodsweredescribedindetail().Inbrief,.:..,SOC,IC,(WatersCorporation)andanalyzedpermassspectrometryusingaQ-TOFPremiermassspectrometer(WatersCorporation).ForeachROI,threebiologicalreplicateswereprepared,prisingtissuepooledoversixanimals,andeachbiologicalreplicatewasmeasuredinfourtechnicalreplicates,resultingin12replicates(n=12).TranscriptanalysisTissuepreparationformicroarrayexperimentsBraintissuewasobtainedasdescribed(,Kaltwa?,).TissuesampleswerestoredinRNAlater?(AM7021,ThermoFisherScientific,Dreieich,Germany)-freeinstrumentsortheequipmentwascleanedusingRNaseAWAYTM(10328011,ThermoFisherScientific).ArticleTotalRNAisolation,SOC,IC)orMidi(RoB)(74104&75144,QiagenGmbH,Hilden,Germany)followingmanufacturer’,binedandhomogenizedin1mL(MiniKit)or4mL(MidiKit)RAD-8(RAGmbH,Müllheim,Germany)andmixedafterwardswith200μLor1mLchloroform,(Biofugefresco,Heraeus,Hanau,Germany,13,700×g,15min,4°CorMultifuge1S-R,ThermoFisherScientific,4,618×g,19min,4°C),thetopphasewasdilutedwithanequalvolumeof70%ethanol,appliedtospincolumns,andconcentrated(13,700×g,15s,or4,618×g,15s).DNAwasdigestedon-columnpriortoelutionoftheRNAwithRNase-(G2939BA&5067-1513,AgilentTechnologies,SantaClara,USA).Microarrayexperiments1μgofRNAwasamplifiedandlabeledusingtheQuickAmpLabelingKit,two-color(5190-04