BiologyLecture1-2:MOLECULARGENETICS-Transcription,TranslationandControlofgeneexpression

• Describetheessentialfeaturesoftranscriptionandtranslationincellsofbothprokaryotes(bacteria)andeukaryotes(fungi,plantsandanimals).

• Understandhowpointandframe-shiftmutationscanaffectproductionoffunctionalgeneproducts/proteins.• Understandthebasicprocessesbywhichgeneexpressioncanbecontrolledattheleveloftranscriptioninprokaryotes

andeukaryotes.

Transcription: thesynthesisofRNAusinginformationintheDNA.Thetwonucleicacidsarewrittenindifferentformsofthesamelanguageandtheinformationisjustsimply‘rewritten’fromDNAtoRNA.

MessengerRNA(mRNA): carriesageneticmessagefromtheDNAtotheprotein-synthesisingmachineryofthecell.Translation: thesynthesisofapolypeptideusingtheinformationinthemRNA.Duringthisstage,thereis

achangeinlanguage:ThecellmusttranslatethenucleotidesequenceofanmRNAmoleculeintotheaminoacidsequenceofapolypeptide.Thesitesoftranslationareribosomes,molecularcomplexesthatfacilitatetheorderlylinkingofaminoacidsintopolypeptidechains.

Transcription/Translationinbacteriavs.eukaryotes:

Becausebacteriadonothavenuclei,theirDNAisnotseparatedbynuclearmembranesfromribosomesandotherprotein-synthesisingequipment.ThislackofcompartmentalisationallowstranslationofanmRNAtobeginwhileitstranscriptionisstillinprogress.Inaeukaryoticcell,bycontrast,thenuclearenvelopeseparatestranscriptionfromtranslationinspaceandtime.Transcriptionoccursinthenucleus,andmRNAisthentransportedtothecytoplasm,wheretranslationoccurs.

Primarytranscript: theinitialRNAtranscriptfromanygene,includingthosespecifyingRNAthatisnottranslatedintoprotein.Templatestrand:thestrandofDNAthatprovidesthepatternortemplateforthesequenceofnucleotidesinanRNAtranscript.

*ThemRNAisreadinthe5’-3’direction.

TRANSCRIPTION

MolecularcomponentsofTranscription:

• RNApolymerase:priesthetwostrandsofDNAapartandjoinstogetherRNAnucleotidescomplementarytotheDNAtemplatestrand.RNApolymerasescanassembleapolynucleotideonlyinits5’!3’direction.UnlikeDNApolymerasehowever,RNApolymerasesareabletostartsachainfromscratch;theydon’tneedaprimer.

• Promoter:theDNAsequencewhereRNApolymeraseattachesandinitiatestranscription.

• TranscriptionUnit:ThestretchofDNAdownstreamfromthepromoter.

SynthesisofanRNATranscript:

Therearethreestagesoftranscription:initiation,elongationandterminationoftheRNAchain.

1. Initiation• RNApolymerasebindstoapromoterthatdetermineswhere

transcriptionstartsandwhichofthetwostrandsoftheDNAhelixisusedasthetemplate.

• Ineukaryotes,acollectionofproteinscalledtranscription

factorsmediatethebindingofRNApolymeraseandtheinitiationoftranscription.OnlyaftertranscriptionfactorsareattachedtothepromoterdoesRNApolymeraseIIbindtoit.ThecomplexoftranscriptionfactorsandRNApolymeraseIIboundtothepromoteriscalledatranscriptioninitiationcomplex.

• OncetheappropriatetranscriptionfactorsarefirmlyattachedtothepromoterDNAandthepolymeraseisboundinthecorrectorientation,theenzymeunwindsthetwoDNAstrandsandbeginstranscribingthetemplatestrandatthestartpoint.

2. Elongation• AsRNApolymerasemovesalongtheDNA,ituntwists

thedoublehelix,exposingDNAnucleotidesforpairingwithRNAnucleotides.

• Theenzymeaddsnucleotidestothe3’endofthegrowingRNAmoleculeasitcontinuesalongthedoublehelix.

• ThenewRNAmoleculepeelsawayfromitsDNAtemplateandtheDNAdoublehelixre-forms.

3. Termination• Terminationdiffersbetweenbacteriaandeukaryotes.Inbacteria,transcriptionproceedsthroughaterminator

sequenceintheDNA.Thetranscribedterminator(anRNAsequence)functionsastheterminationsignal,causingthepolymerasetostitchfromtheDNAandreleasethetranscript,whichrequiresnofurthermodificationbeforetranslation.

• Ineukaryotes,RNApolymeraseIItranscribesasequenceontheDNAcalledthepolyadenylationsignalsequence,whichspecifiesapolyadenylationsignalinthepre-mRNA.Thepre-mRNAthenundergoesprocessing,thetopicofthenextsection.Al-thoughthatcleavagemarkstheendofthemRNA,theRNApolymeraseIIcontinuestotranscribe.Sincethenew59endisn’tprotectedbyacap,however,enzymesdegradetheRNAfromthe59end.Thepolymerasecontinuestranscribing,pursuedbytheenzymes,untiltheycatchuptothepolymeraseanditdissociatesfromtheDNA.

Q.WhatenablesRNApolymerasetostarttranscribingageneattherightplaceontheDNAinabacterialcell?Inaeukaryoticcell?

ModifyingRNAaftertranscription:

• 5’endreceivesa5’capandapoly-Atailisaddedtothe3’end(termedpre-RNA).ThesefeaturesfacilitatetheexportofmaturemRNAfromthenucleus,protectthemRNAfromdegradationbyhydrolyticenzymesandhelpribosomesattachtothe5’endofthemRNAoncethemRNAreachesthecytoplasm.

• RNAsplicing:noncodingregions(introns)arecutoutoftheRNAmolecule(bysmallRNAscalledspliceosomes)whilstexonsaremaintainedandexpressed.

AlternateRNAsplicing: Alternativesplicingisaregulatedprocessduringgeneexpressionthatresultsinasinglegenecodingformultipleproteins.Inthisprocess,particularexonsofagenemaybeincludedwithinorexcludedfromthefinal,processedmessengerRNA(mRNA)producedfromthatgene.

TRANSLATION

MolecularcomponentsofTranslation:

• tRNAtranslatesmessagesintheformofseriesofcodonsalonganmRNAmolecule.ThefunctionoftRNAistotransferaminoacidsfromthecytoplasmicpoolofaminoacidstoagrowingpolypeptideinaribosome.

• EachtRNAmoleculetranslatesagivenmRNAcodonintoacertainaminoacid.ThisispossiblebecausetRNAbearsaspecificaminoacidatoneend,whichattheotherendarenucleotidetripletsthatcanbasepairwiththecomplementarycodononmRNA.

• TheanticodonisthepartofthenucleotidetripletthatbasepairstoaspecificmRNAcodon.• LikemRNAandothertypesofcellularRNA,transferRNAmoleculesaretranscribedfromDNAtemplates.Ina

eukaryoticcell,tRNA,likemRNA,ismadeinthenucleusandthentravelsfromthenucleustothecytoplasm,whereitwillparticipateintheprocessoftranslation.Inbothbacterialandeukaryoticcells,eachtRNAmoleculeisusedrepeatedly,pickingupitsdesignatedaminoacidinthecytosol,depositingthiscargoontoapolypeptidechainattheribosome,andthenleavingtheribosome,readytopickupanotherofthesameaminoacid.

InadditiontoabindingsiteformRNA,eachribosomehasthreebindingsitesfortRNA,asdescribedinFigure17.17.ThePsite(peptidyl-tRNAbindingsite)holdsthetRNAcarryingthegrowingpolypeptidechain,whiletheAsite(aminoacyl-tRNAbindingsite)holdsthetRNAcarryingthenextaminoacidtobeaddedtothechain.DischargedtRNAsleavetheribosomefromtheEsite(exitsite).TheribosomeholdsthetRNAandmRNAincloseproximityandpositionsthenewaminoacidsothatitcanbeaddedtothecarboxylendofthegrowingpolypeptide.Itthencatalysestheformationofthepeptidebond.Asthepolypeptidebecomeslonger,itpassesthroughanexittunnelintheribosome’slargesubunit.Whenthepolypeptideiscomplete,itisreleasedthroughtheexittunnel.