Investigating the link between tRNA and mRNA abundance

175 Pages·2016·10.96 MB·English

ThisdissertationissubmittedforthedegreeofDoctorofPhilosophy
Investigating the link between
tRNA and mRNA abundance
in mammals
Konrad Ludwig Moritz Rudolph
May 2015
St Edmund’s College,
University of Cambridge
EMBL-EBI
Thesourcecodeofthethesisisavailableathttps://github.com/klmr/thesis.
LicensedunderCreativeCommonsAttribution(CCBY)3.0
Thisdissertationistheresultofmyownworkandincludesnothingwhichis
theoutcomeofworkdoneincollaborationexceptasdeclaredinthePreface
andspecifiedinthetext.
It is not substantially the same as any that I have submitted, or, is being
concurrently submitted for a degree or diploma or other qualification at
the University of Cambridge or any other University or similar institution
except as declared in the Preface and specified in the text. I further state
that no substantial part of my dissertation has already been submitted, or,
isbeingconcurrentlysubmittedforanysuchdegree,diplomaorotherqual-
ification at the University of Cambridge or any other University of similar
institutionexceptasdeclaredinthePrefaceandspecifiedinthetext.
ItdoesnotexceedtheprescribedwordlimitfortherelevantDegreeCom-
mittee.
3

Contents
Listofabbreviations 9
ListofFigures 11
ListofTables 15
Summary 17
Acknowledgements 19
1. Introduction 21
1.1. Thecentraldogma 21
1.1.1. dna 22
1.1.2. rna 24
1.1.3. Proteins 25
1.2. Transcription&translation 26
1.2.1. Transcription 26
1.2.2. Thegeneticcode 28
1.2.3. Transferrna 28
1.2.4. Translation 34
1.3. rnasequencingquantifiesprotein-codinggeneexpression 36
1.3.1. Thetranscriptomereflectsthestateofthecell 36
1.3.2. Microarrays 36
1.3.3. rna-seq 37
1.3.4. Expressionnormalisation 40
1.3.5. Differentialexpression 42
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contents
1.4. PoliiiChip-sequencingquantifiestrnageneexpression 43
1.4.1. Chip-seqisadnabindingassay 43
1.4.2. Quantifyingexpressionoftrnagenes 44
1.5. Mouseliverdevelopment 47
1.6. Quantifyingcodonusageandanticodonabundance 49
1.7. Structureofthisthesis 54
2. Developmentalstabilityofthemrna–trnainterface 55
2.1. Protein-codinggeneexpressionchangesdynamically
duringmousedevelopment 57
2.2. Dynamicchangesoftrnageneexpressionduringmouse
development 61
2.3. Everymousemrnatranscriptomeencodesthesame
distributionoftripletcodonsandaminoacids 64
2.4. Stableisoacceptoranticodonabundancethrough
developmentindicatestightregulationoftrnagene
expression 67
2.5. mrnatripletcodonusageishighlycorrelatedwithtrna
anticodonisoacceptorabundanceduringdevelopment 67
2.6. Variablechromatinaccessibilitymayinfluencetrnagene
transcription 70
2.7. trnaanticodonisoacceptorfamiliesaretranscriptionally
compensatedacrossdevelopment 75
3. Implicationsofcodon–anticodoninteractionontheregulation
oftranslation 81
3.1. “Adualprogramfortranslationregulationincellular
proliferationanddifferentiation” 81
3.1.1. trnaanticodonschangeinabundanceintumour
tissues 82
3.1.2. Codonusagediffersbetweengenesinvolvedincell
proliferationandgenesinvolvedindifferentiation 82
6
contents
3.1.3. Differentialcodonusageincell-conditionspecific
genesetsmatchestrnaanticodonabundancein
correspondingcells 84
3.2. Aretrnaanticodonabundanceandcodonusagehighly
adaptedtodifferentcellularconditions? 87
3.2.1. Theeffectofgenesetsizeoncodonusagebias 88
3.2.2. Theextentofanticodonadaptationtodistinct
cellularconditions 90
3.2.3. Othergenomicfeaturesmaydrivetheperceived
codonusagebias 95
3.3. Discussion 97
3.3.1. Codon–anticodonadaptationtodistinctcellular
conditions 98
3.3.2. Evolutionaryconservationofcodon–anticodon
adaptation 99
4. Thepoliiitranscriptomeconsistsofmorethanjusttrna 101
4.1. Aprofileofpoliiibindingacrossdifferentfeatures 101
4.1.1. sinesaretranscribedbypoliii 104
5. Conclusion 109
5.1. Discussion 110
5.1.1. Regulationoftrnageneexpression 110
5.1.2. Absenceofevidenceforcodonbias-dependent
translationefficiencyinmammals 112
5.1.3. Theextendedpoliiitranscriptome 113
5.2. Futuredirections 114
5.2.1. Regulationoftrnatranscription 114
5.2.2. Codonusageadaptation 114
5.2.3. Poliiitranscriptionofsines 116
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list of abbreviations
APPENDIX
A. Supplementarymaterialforchapter2 121
A.1. Code 121
A.2. Supplementaryfiguresandtables 121
B. Supplementarymaterialforchapter3 147
B.1. Code 147
B.2. Supplementarytables 147
C. Supplementarymaterialforchapter4 155
C.1. Code 155
C.2. Supplementaryfigures 155
Bibliography 159
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List of abbreviations
AAB anticodonabundancebias
cDNA complementarydna
ChIP chromatinimmunoprecipitation
ChIP-seq chromatinimmunoprecipitationfollowedbysequencing
CTCF CCCTC-bindingfactor
CU codonusage
CUB codonusagebias
DE differentiallyexpressed
DNA deoxyribonucleicacid
FDR falsediscoveryrate
FPKM fragmentsperkilobaseoftranscriptpermillionmappedreadpairs
GEO GeneExpressionOmnibus
GO GeneOntology
GRCm38 GenomeReferenceConsortiumMouseBuild38
HIV-1 type-1humanimmunodeficiencyvirus
HTS high-throughputsequencing
LINE longinterspersednuclearelement
lncRNA longnon-codingrna
mRNA messengerrna
NCBIM37 ncbimousegenomebuild37
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list of abbreviations
ncRNA non-codingrna
PCA principalcomponentsanalysis
PCR polymerasechainreaction
Pol I dna-dependentrnapolymerasei
Pol II dna-dependentrnapolymeraseii
Pol III dna-dependentrnapolymeraseiii
RAA relativeanticodonabundance
RCU relativecodonusage
RISC rna-inducedsilencingcomplex
RNA ribonucleicacid
RNA-seq rnasequencing
RPKM readcountperkilobaseoftranscriptpermillionmappedreads
rRNA ribosomalrna
SINE shortinterspersednuclearelement
tAI trnaadaptationindex
TBP TATAbindingprotein
TF transcriptionfactor
TFIIIB transcriptionfactoriiiB
TFIIIC transcriptionfactoriiiC
TPM transcriptspermillion
tRNA transferrna
TSS transcriptionstartsite
UTR untranslatedregion
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