RNA Detection: Methods and Protocols

492 Pages·2018·12.655 MB·English

by  Imre Gaspar (eds.)

Methods in
Molecular Biology 1649
Imre Gaspar Editor
RNA
Detection
Methods and Protocols
M M B
ETHODS IN OLECULAR IO LO GY
SeriesEditor
JohnM.Walker
School of Lifeand MedicalSciences,
University of Hertfordshire,Hatfield,
Hertfordshire AL109AB,UK
Forfurther volumes:
http://www.springer.com/series/7651
RNA Detection
Methods and Protocols
Edited by
Imre Gaspar
Developmental Biology Unit, EMBL Heidelberg, Heidelberg, Germany
Editor
ImreGaspar
DevelopmentalBiologyUnit
EMBLHeidelberg
Heidelberg,Germany
ISSN1064-3745 ISSN1940-6029 (electronic)
MethodsinMolecularBiology
ISBN978-1-4939-7212-8 ISBN978-1-4939-7213-5 (eBook)
DOI10.1007/978-1-4939-7213-5
LibraryofCongressControlNumber:2017957614
©SpringerScience+BusinessMediaLLC2018
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CoverCaption: Localizationofoskar mRNA(incyan,labelled witha set ofFIT probes)to theposteriorpole of a
developingDrosophilaoocyte(ontheright).oskarmRNAistranscribedinthenursecells(ontheleft)andtransported
throughcytoplasmicbridgesintotheoocytebycytoplasmicdynein.Intheoocyte,localizationofoskartotheposterior
poleismediatedbykinesinheavychain(inred,labelledwithmKate2fluorescentprotein).Note,thatoskarmRNAis
exclusivelyexpressedinthefemalegerm-line(nursecellsandtheoocyte)anditisabsentfromthesomaticfollicular
epithelium(ontheright,markedbykinesin)surroundingtheoocyte.
Printedonacid-freepaper
ThisHumanaPressimprintispublishedbySpringerNature
TheregisteredcompanyisSpringerScience+BusinessMediaLLCNewYork
Theregisteredcompanyaddressis:233SpringStreet,NewYork,NY10013,U.S.A.
Preface
Inthelastcoupleofyears,researchbasedonhigh-throughputassaysrevealedthattheRNA
world is much more complex than initially anticipated entangling virtually all areas of cell
anddevelopmentalbiology.Semiautomatedvisualscreensdemonstratedthatlargefraction
ofthetranscriptomeisdistributednonuniformlywithinthecell,suggestingthepresenceof
underlying active localization mechanisms. On the other hand, the nonbiased capture of
RNA interactome showed that 8–10% of the total proteome could directly bind (m)RNA,
including hundreds of novel RNA binding proteins, such as enzymes of fundamental
biosynthesis pathways, components of the cytoskeleton, the endocytosis, and secretory
pathways. Some of these novel RNA-binding proteins harbor low-complexity domains,
making them capable of spontaneously self-assembling into higher-order structures both
invitroandinvivo,dynamicallyformingRNA-containingmembranelessorganelles,suchas
Cajal bodies, nuclear speckles, and paraspeckles, RNA and stress granules, nuage or germ
granules.Inspecimenpreviouslyconsideredhomogeneousincludingtumorousmalforma-
tions,quantitativeRNAimagingandcorrelativehigh-contentimagingcoupledwithsingle
cell transcriptome analysis demonstrated a high degree of heterogeneity which directly
impacts prognosis and possible therapy. As revealed by in vivo functional assays of single-
molecule sensitivity, thisheterogeneityis mainlydue to the stochastic nature ofthe under-
lyingbiologicalprocesses,suchastranscriptionandtranslation.
The advances in RNA biology render advanced RNA detection and visualization tools
invaluable to cell and developmental biologists as well as to medical researchers and
practicing clinicians. Although the amount of technology development in the last couple
ofyearsrendersitimpossibletocovereverypossibleaspectsofRNAdetection,thisvolume
aims to introduce the various concepts and the methods of detecting RNA in biological
materialinavarietyofmodelsystems.Thedetailedprotocolsandthetipsandtricksofthe
presentedassayswillallowtheoptimizationandtheadaptationofthesemethodstoaddress
different biological questions of RNA, and hopefully, this volume of the MiMB series
becomes a useful everyday companion of every novel or experienced scientists of the
expandingRNAworld.
Heidelberg,Germany ImreGaspar
v
Contents
Preface..................................................................... v
Contributors................................................................ xi
1 TheSecretLifeofRNA:LessonsfromEmergingMethodologies............. 1
CarolineMedioniandFlorenceBesse
0
2 Quantificationof2-O-MeResiduesinRNAUsingNext-Generation
Sequencing(IlluminaRiboMethSeqProtocol).............................. 29
LiliaAyadi,YuriMotorin,andVirginieMarchand
3 Identifyingthem6AMethylomebyAffinityPurificationandSequencing ...... 49
PhillipJ.HsuandChuanHe
4 PARIS:PsoralenAnalysisofRNAInteractionsandStructures
withHighThroughputandResolution.................................... 59
ZhipengLu,JingGong,andQiangfengCliffZhang
5 Axon-TRAP-RiboTag:AffinityPurificationofTranslatedmRNAs
fromNeuronalAxonsinMouseInVivo................................... 85
ToshiakiShigeoka,JaneJung,ChristineE.Holt,andHosungJung
6 LCM-Seq:AMethodforSpatialTranscriptomicProfilingUsing
LaserCaptureMicrodissectionCoupledwithPolyA-Based
RNASequencing ....................................................... 95
SusanneNichterwitz,JulioAguilaBenitez,ReinHoogstraaten,
QiaolinDeng,andEvaHedlund
7 SpatialTranscriptomics:ConstructingaSingle-CellResolution
Transcriptome-WideExpressionAtlas ..................................... 111
KaiaAchim,HernandoMartı´nezVergara,andJean-BaptistePettit
8 SinglemRNAMoleculeDetectioninDrosophila............................ 127
ShawnC.LittleandThomasGregor
9 DetectionandAutomatedAnalysisofSingleTranscriptsatSubcellular
ResolutioninZebrafishEmbryos ......................................... 143
L.CarineStapel,ColemanBroaddus,andNadineL.Vastenhouw
10 Super-ResolutionSingleMoleculeFISHattheDrosophila
NeuromuscularJunction................................................. 163
JoshuaS.Titlow,LuYang,RichardM.Parton,AnaPalanca,
andIlanDavis
11 DetectionofmRNAandAssociatedMoleculesbyISH-IEM
onFrozenSections...................................................... 177
CatherineRabouille
12 HybridizationChainReactionforDirectmRNADetection
WithoutNucleicAcidPurification ........................................ 187
YaoXuandZhiZheng
vii
viii Contents
13 InSituDetectionofMicroRNAExpressionwithRNAscopeProbes .......... 197
ViravuthP.Yin
14 PadlockProbestoDetectSingleNucleotidePolymorphisms................. 209
TomaszKrzywkowskiandMatsNilsson
15 QuantifyingGeneExpressioninLivingCellswithRatiometric
BimolecularBeacons .................................................... 231
YantaoYang,MingmingChen,ChristopherJ.Krueger,AndrewTsourkas,
andAntonyK.Chen
16 OptimizingMolecularBeaconsforIntracellularAnalysisofRNA............. 243
MingmingChen,YantaoYang,ChristopherJ.Krueger,
andAntonyK.Chen
17 LiveImagingofNuclearRNPsinMammalianComplexTissue
withECHO-liveFISH................................................... 259
DanOhtanWang
18 InVivoVisualizationandFunctionProbingofTransportmRNPs
UsingInjectedFITProbes............................................... 273
JasmineChamiolo,ImreGaspar,AnneEphrussi,andOliverSeitz
19 VisualizingRNAinLiveBacterialCellsUsingFluorophore-
andQuencher-BindingAptamers ......................................... 289
MuratSunbul,AnkitaArora,andAndresJa€schke
20 MethodforImagingLive-CellRNAUsinganRNAAptamer
andaFluorescentProbe................................................. 305
Shin-ichiSato,KenjiYatsuzuka,YousukeKatsuda,andMotonariUesugi
21 RNALiveImagingintheModelMicroorganismUstilagomaydis............. 319
SabrinaZander,KiraMu€ntjes,andMichaelFeldbru€gge
22 Real-TimeFluorescenceImagingofSingle-MoleculeEndogenous
NoncodingRNAinLivingCells.......................................... 337
HideakiYoshimuraandTakeakiOzawa
23 LiveImagingofmRNASynthesisinDrosophila............................. 349
HernanG.GarciaandThomasGregor
24 ImagingNewlyTranscribedRNAinCellsbyUsingaClickable
Azide-ModifiedUTPAnalog............................................. 359
AnupamA.Sawant,SanjeevGalande,andSeergazhiG.Srivatsan
25 DetectionoftheFirstRoundofTranslation:TheTRICKAssay .............. 373
FrankaVoigt,JanEglinger,andJeffreyA.Chao
26 ImagingTranslationDynamicsofSinglemRNA
MoleculesinLiveCells.................................................. 385
SuzanRuijtenberg,TimA.Hoek,XiaoweiYan,
andMarvinE.Tanenbaum
27 SystematicDetectionofPoly(A)+RNA-InteractingProteins
andTheirDifferentialBinding............................................ 405
MihaMilekandMarkusLandthaler
Contents ix
28 IsolationandCharacterizationofEndogenousRNPs
fromBrainTissues...................................................... 419
RicoSchieweck,FoongyeeAng,RenateFritzsche,andMichaelA.Kiebler
29 IndividualNucleotideResolutionUVCross-Linking
andImmunoprecipitation(iCLIP)toDetermineProtein–RNAInteractions ... 427
ChristopherR.Sibley
30 RNATagging:PreparationofHigh-ThroughputSequencingLibraries........ 455
ChristopherP.LapointeandMarvinWickens
31 RAP-MS:AMethodtoIdentifyProteinsthatInteractDirectly
withaSpecificRNAMoleculeinCells..................................... 473
ColleenA.McHughandMitchellGuttman
Index ...................................................................... 489
Contributors
KAIAACHIM (cid:2) DevelopmentalBiologyUnit,EuropeanMolecularBiologyLaboratory,
Heidelberg,Germany
FOONGYEEANG (cid:2) CellBiology,BiomedicalCenter,MedicalFaculty,LMUMunich,Planegg-
Martinsried,Germany
ANKITAARORA (cid:2) InstituteofPharmacyandMolecularBiotechnology,HeidelbergUniversity,
Heidelberg,Germany
LILIAAYADI (cid:2) IMoPAUMR7365CNRS-LorraineUniversity,BioPoleLorraineUniversity,
Vandoeuvre-les-Nancy,France
JULIO AGUILABENITEZ (cid:2) DepartmentofNeuroscience,KarolinskaInstitutet,Stockholm,
Sweden;DepartmentofCellandMolecularBiology,KarolinskaInstitutet,Stockholm,
Sweden
FLORENCEBESSE (cid:2) Universite´ Coˆted’Azur,CNRS,Inserm,iBV,Nice,France
COLEMANBROADDUS (cid:2) Center forSystemsBiologyDresden,MaxPlanckInstituteof
MolecularCellBiologyandGenetics,Dresden,Germany
JASMINECHAMIOLO (cid:2) DepartmentofChemistry,HumboldtUniversityBerlin,Berlin,
Germany
JEFFREYA.CHAO (cid:2) FriedrichMiescherInstituteforBiomedicalResearch,Basel,Switzerland
ANTONY K.CHEN (cid:2) DepartmentofBiomedicalEngineering,CollegeofEngineering,Peking
University,HaidianDistrict,Beijing,China
MINGMINGCHEN (cid:2) DepartmentofBiomedicalEngineering,CollegeofEngineering,Peking
University,HaidianDistrict,Beijing,China;Peking-TsinghuaCenter forLifeSciences,
PekingUniversity,Beijing,China;AcademyforAdvancedInterdisciplinaryStudies,
PekingUniversity,Beijing,China
ILANDAVIS (cid:2) DepartmentofBiochemistry,UniversityofOxford,Oxford,UK
QIAOLIN DENG (cid:2) DepartmentofCellandMolecularBiology,KarolinskaInstitutet,
Stockholm,Sweden
JANEGLINGER (cid:2) FriedrichMiescherInstituteforBiomedicalResearch,Basel,Switzerland
ANNE EPHRUSSI (cid:2) EuropeanMolecularBiologyLaboratory(EMBL)Heidelberg,Heidelberg,
Germany
MICHAELFELDBRu€GGE (cid:2) InstituteforMicrobiology,Heinrich-HeineUniversityDu€sseldorf,
ClusterofExcellenceonPlantSciences(CEPLAS),Du€sseldorf,Germany
RENATEFRITZSCHE (cid:2) CellBiology,BiomedicalCenter,MedicalFaculty,LMUMunich,
Planegg-Martinsried,Germany
SANJEEVGALANDE (cid:2) CenterofExcellenceinEpigenetics,IndianInstituteofScienceEducation
andResearch,Pune,Pune,India
HERNANG.GARCIA (cid:2) DepartmentofMolecularandCellBiology,UniversityofCalifornia,
Berkeley,CA,USA;DepartmentofPhysicsandBiophysicsGraduateGroup,Universityof
California,Berkeley,CA,USA
IMREGASPAR (cid:2) EuropeanMolecularBiologyLaboratory(EMBL)Heidelberg,Heidelberg,
Germany
JING GONG (cid:2) MOEKeyLaboratoryofBioinformatics,BeijingAdvancedInnovationCenter
forStructuralBiology,Center forSyntheticandSystemsBiology,Tsinghua-PekingJoint
xi
xii Contributors
Center forLifeSciences,SchoolofLifeSciences,TsinghuaUniversity,Haidian,Beijing,
China
THOMASGREGOR (cid:2) DepartmentofPhysics,Lewis-SiglerInstituteforIntegrativeGenomics,
PrincetonUniversity,Princeton,NJ,USA
MITCHELLGUTTMAN (cid:2) DivisionofBiologyandBiologicalEngineering,CaliforniaInstitute
ofTechnology,Pasadena,CA,USA
CHUANHE (cid:2) DepartmentofChemistry,InstituteforBiophysicalDynamics,TheUniversityof
Chicago,Chicago,IL,USA;HowardHughesMedicalInstitute,TheUniversityofChicago,
Chicago,IL,USA;DepartmentofBiochemistryandMolecularBiology,TheUniversityof
Chicago,Chicago,IL,USA
EVAHEDLUND (cid:2) DepartmentofNeuroscience,KarolinskaInstitutet,Stockholm,Sweden
TIMA.HOEK (cid:2) HubrechtInstitute,KNAWandUniversityMedicalCenterUtrecht,Utrecht,
TheNetherlands
CHRISTINEE.HOLT (cid:2) DepartmentofPhysiology,DevelopmentandNeuroscience,University
ofCambridge,Cambridge,UK
REINHOOGSTRAATEN (cid:2) DepartmentofNeuroscience,KarolinskaInstitutet,Stockholm,
Sweden;DepartmentofTranslationalNeuroscience,BrainCenterRudolfMagnus,
Utrecht,Netherlands
PHILLIPJ.HSU (cid:2) DepartmentofChemistry,InstituteforBiophysicalDynamics,The
UniversityofChicago,Chicago,IL,USA;HowardHughesMedicalInstitute,The
UniversityofChicago,Chicago,IL,USA;CommitteeonImmunology,TheUniversityof
Chicago,Chicago,IL,USA
ANDRESJ€aSCHKE (cid:2) InstituteofPharmacyandMolecularBiotechnology,Heidelberg
University,Heidelberg,Germany
HOSUNGJUNG (cid:2) DepartmentofAnatomy,BrainResearchInstitute,andBrainKorea21
PLUSProjectforMedicalScience,YonseiUniversityCollegeofMedicine,Seoul,Republicof
Korea
JANEJUNG (cid:2) DepartmentofAnatomy,BrainResearchInstitute,andBrainKorea21PLUS
ProjectforMedicalScience,YonseiUniversityCollegeofMedicine,Seoul,RepublicofKorea
YOUSUKE KATSUDA (cid:2) InstituteforIntegratedCell-MaterialSciences(WPI-iCeMS),Kyoto
University,Kyoto,Japan
MICHAELA.KIEBLER (cid:2) CellBiology,BiomedicalCenter,MedicalFaculty,LMUMunich,
Planegg-Martinsried,Germany
CHRISTOPHERJ.KRUEGER (cid:2) DepartmentofBiomedicalEngineering,CollegeofEngineering,
PekingUniversity,HaidianDistrict,Beijing,China;WallaceHCoulterDepartmentof
BiomedicalEngineering,GeorgiaInstituteofTechnology,Atlanta,GA,USA
TOMASZKRZYWKOWSKI (cid:2) ScienceforLifeLaboratory,DepartmentofBiochemistryand
Biophysics,StockholmUniversity,Solna,Sweden
MARKUSLANDTHALER (cid:2) RNABiologyandPosttranscriptionalRegulation,MaxDelbru€ck
Center forMolecularMedicineBerlin,BerlinInstituteforMolecularSystemsBiology,
Berlin,Germany
CHRISTOPHERP.LAPOINTE (cid:2) DepartmentofBiochemistry,UniversityofWisconsin-Madison,
Madison,WI,USA
SHAWNC.LITTLE (cid:2) DepartmentofCellandDevelopmentalBiology,Universityof
PennsylvaniaPerelmanSchoolofMedicine,Philadelphia,PA,USA
ZHIPENG LU (cid:2) DepartmentofDermatology,Center forPersonalDynamicRegulomes,
StanfordUniversity,Stanford,CA,USA