Inthecrowdedanddynamicthree-dimensionalspaceofthenucleus,activegenesoftenshare localneighborhoodswithoneanother.Despitesignificantrecentprogressinanalysesof3D positioningofgenes,itisnotyetfullyunderstoodhowtheco-localizationofgenesplaysarolein theirtranscriptionaloutput.Thelong-termgoalofthisresearchistounderstandhowagene?s expressionisinfluencedbyitsneighborsinspaceandtime.Thestudyofhowco-localized genessharelocalresourceshasthepotentialtoshedsignificantlightonfundamental mechanismsoftranscription,butprogresshasbeenseverelylimitedbyarelianceonmethods thatexaminetranscriptionand3Dpositionseparatelyinfixedordisruptedtissues.Theobjective ofthisproposalistouncovertheimpactofgeneco-localizationontranscriptionaldynamicsin livingcellstoprovideadynamicspatiotemporalquantificationoftranscriptionforgenesthat sharealocal3Dneighborhood.Toaddressthisobjective,theproposalfocusesonDrosophila melanogasterandtakesadvantageofaphenomenoncalledtransvection,wheresequencesat allelicpositionsonhomologouschromosomesarestablyco-localizedviaaprocesscalled somatichomologpairing.Furthermore,theproposedexperimentsemploytheMS2andPP7 systemsforliveanalysisoftranscriptionaldynamics,whichpermitsimultaneousquantificationof transcriptionalactivityandassessmentofthe3Dpositionsofgenesovertimeinlivingcells.The centralhypothesisguidingtheproposalisthatco-localizationofgenesviasomatichomolog pairingwillresultinthesharingoflocalstoresofresourcesnecessaryfortranscription,which canbeunderstoodbyanalyzinghowco-localizationinfluencestheparametersoftranscriptional dynamics,includingthefrequency,duration,andamplitudesoftranscriptionalbursts.This hypothesisistestedinthecontextoftwocontrastingscenariosresultingfromco-localizationof genesviatransvection:inonescenario,co-localizedpromotersincisandintranstoan enhancercompeteforactivation,whileinasecondscenario,co-localizedenhancerscooperate toachieveaugmentedlevelsoftranscriptionaloutput.Theproposedexperimentshavethe expectedoutcomeofidentifyingpreciselyandquantitativelyhowinteractionsbetweenco- localizedgenesinfluencetranscriptionaldynamics,providingkeyinsightsintomechanismsby which3Dgenomeorganizationcontrolsgeneexpression.
Activegenesoftenlocalizetogethertodiscreteregionsofthenucleus,buttheconsequencesof co-localizationongeneexpressionarepoorlyunderstood.Weproposetousethemodel organismDrosophilamelanogastertounderstandhowgenesinteractwithin3Dneighborhoods, eitherincompetitionorcooperationwithoneanother,toaccessthefinitepoolofresources requiredforgeneactivation.Giventheimportanceofgeneregulationforhumandevelopment andhealth,ourstudieswillbebroadlyrelevantinthemedicalarena.
