JournalofVisualizedExperimentswww.jove.comCopyright©2010CreativeCommonsAttribution-NonCommercialLicenseJuly2010|41|e2101|Page1of7VideoArticleGenome-wideAnalysisusingChIPtoIdentifyIsoform-specificGeneTargetsMichaelL.Beshiri1,AbulIslam2,DannielleC.DeWaal1,WilliamF.Richter1,JenniferLove3,NuriaLopez-Bigas2,ElizavetaV.Benevolenskaya11DepartmentofBiochemistryandMolecularGenetics,UniversityofIllinoisChicago-UIC2ResearchUnitonBiomedicalInformatics,UniversitatPompeuFabra3GenomeTechnologyCore,WhiteheadInstituteforBiomedicalResearchCorrespondenceto:ElizavetaV.Benevolenskayaatevb@uic.eduURL:http://www.jove.com/video/2101DOI:doi:10.3791/2101Keywords:Biochemistry,Issue41,chromatinimmunoprecipitation,ChIP-Seq,RBP2,JARID1A,KDM5A,isoform-specificrecruitmentDatePublished:7/7/2010Citation:Beshiri,M.L.,Islam,A.,DeWaal,D.C.,Richter,W.F.,Love,J.,Lopez-Bigas,N.,Benevolenskaya,E.V.Genome-wideAnalysisusingChIPtoIdentifyIsoform-specificGeneTargets.J.Vis.Exp.(41),e2101,doi:10.3791/2101(2010).AbstractRecruitmentoftranscriptionalandepigeneticfactorstotheirtargetsisakeystepintheirregulation.Prominentlyfeaturedinrecruitmentaretheproteindomainsthatbindtospecifichistonemodifications.Onesuchdomainistheplanthomeodomain(PHD),foundinseveralchromatin-bindingproteins.TheepigeneticfactorRBP2hasmultiplePHDdomains,however,theyhavedifferentfunctions(Figure4).Inparticular,theC-terminalPHDdomain,foundinaRBP2oncogenicfusioninhumanleukemia,bindstotrimethylatedlysine4inhistoneH3(H3K4me3)1.ThetranscriptcorrespondingtotheRBP2isoformcontainingtheC-terminalPHDaccumulatesduringdifferentiationofpromonocytic,lymphoma-derived,U937cellsintomonocytes2.Consistentwithbothsetsofdata,genome-wideanalysisshowedthatindifferentiatedU937cells,theRBP2proteingetslocalizedtogenomicregionshighlyenrichedforH3K4me33.LocalizationofRBP2toitstargetscorrelateswithadecreaseinH3K4me3duetoRBP2histonedemethylaseactivityandadecreaseintranscriptionalactivity.Incontrast,twootherPHDsofRBP2areunabletobindH3K4me3.Notably,theC-terminaldomainPHDofRB...
