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To determinewhether these proteins could bind to the histone H3 N-terminus, you have covalently attached to separatebeads various versions of the H3 N-terminal peptide-unmodified,**'il*specificityof HP1proteins(Problem4-13).Eachproteinatthe left was detectedbyimmunoblottingusingaspecificantibodyafterseparationby 5D5-polyacrytForamidegel electrophoresis.eachhistoneN-terminalpeptidethe total input protein(l),the unboundprotein(U),and the bound protein(B)areindicated.(AdaptedfromM.
Lachneret al.,Nature410:116-120,2001.Withfrom MacmillanoermissionLtd.)PublishersLys-9-dimethylated (Kg-Me), and Ser-10-phosphorylated(S10-P)-along with an unmodified tail from histone H4'This arrangement allows you to incubate the beadswith various proteins,wash away unbound proteins,and then elutebound proteins for assaybyWesternblotting' The resultsof'pull-dor.tm' assay for the HPI proteins are shor,rminyourFigure Q4-3, along with the results from severalcontrol proteins, including Pax5,a gene regulatory protein, polycombprotein Pcl, which is known to bind to histones, andSuv39h1,a histone methyltransferase.Basedon these results,which of the proteins tested bindto the unmodified tails of histones?Do any of the HPl proteins and control proteins selectivelybind to the modifiedhistone N-terminal peptides?\A/hat histone modificationwould you predict would be found in heterochromatin?4-14 Mobile piecesof DNA-transposable elements-thatinsert themselvesinto chromosomes and accumulate duringevolution make up more than 40Toof the human genome.Transposableelements of four types-long interspersedelements (LINEs), short interspersedelements (SINEs),LIRand DNA transposons-are insertedmoreretrotransposons,or less randomly throughout the human genome.These elements are conspicuously rare at the four homeobox geneclusters, HoxA, HoxB, HoxC, and HoxD, as illustrated forHoxD in Figure Q4-4, along with an equivalent region ofchromosome 22,which lacks a ,flor cluster' Each Ilor clusteris about 100kb in length and contains9 to 1l genes,whosedifferential expressionalong the anteroposterior axis of thedeveloping embryo establishesthe basic body plan forhumans (and for other animals).
Vfhy do you suppose thattransposableelements are so rare in the Ho.t clusters?chromosome22raakbHoxD clusterelementsand genesin 1 Mb regionsofFigureQ4-4 Transposablechromosomes2 and 22 (Problem4-14). Linesthat projectupwardindicateexonsof known genes.Linesthat proiectdownwardthey areso numerous(constitutingelements;indicatetransposablegenome)that they mergeinto nearlyhumanofthemorethan 40olo(Adaptedfrom E.Landeret al.,a solidblockoutsidethe Hoxclusters.Nature409'.860-921,2001.With permissionfrom MacmillanLtd.)Publishers262Chapter4: DNA,Chromosomes,and GenomesREFERENCESGeneralHartwelL,HoodL,GoldbergML et al (2006)Genetics:fromGenestoGenomes,3rded Boston:McGrawHillOlsonMV(2002)TheHumanGenomeProject:perspectivea p/ayer'sJ |'/olBiol3l9.931942.StrachanT & ReadAP(2004)HumanMolecularGeneticsNewYork:G a r l a nSdc i e n c eWolffeA ('l999)Chromatin:Structureand Function,3rded NewYork:AcademlcPressThe Structureand Function of DNAAveryOT,MacleodCM & McCartyM (1944)Srudieson the chemicalnatureofthe substanceinducingtransformationof pneumococcaltypes ) Expltled79:137-158MeselsonM & StahlFW(1958)Therepltcationof DNAin E.coli procNatlAcadScrUSA44.671-682) o e c u l asr t r u c t u roef n u c l e iac c i d sWatsonJ D& C r i c kF H C( 1 9 5 3MA structurefor deoxyribosenucleicacidsNature171737738ChromosomalDNA and lts Packagingin the Chromatin FiberJinJ,CaiY,Li B et a (2005)In andout:histonevariantexchangeinchromatinTrendsBiochemScr30:680-687KornbergRD& LorchY (1999)Twentyfiveyearsof the nucleosome,fundamentaparticleof the eukaryotechromosomeCell98:285-294Li G,[evitusM, BustamanteC & W dom J (2005)Rapidsponraneousaccessbilityof nucleosomalDNA lVarure5tructIVolBiol12.46-53LorchY,Maier-DavisB & KornbergRD(2006)Chromatinremodelingbynucleosomedisassemblyin vitraPracNatlAcadSciIJSA103:3090-3093LugerK,MaderAW,RichmondRKet al (j997)Crystalstructureofthenucleosomecoreparticleat 2B A resolutionNature389:251260LugerK & RichmondTJ(1998)Thehistonetailsof the nucleosomeCurapinGenetDevBi4a-146(2003)PhylogenomMa ik H S & HenikoffScs of rhe nucleosomeNatureStructBrol1O:BB2-B9tRiedT,SchrockE,NingY & WienbergI (1998)Chromosomepainring:a usefulart Huml\/tolGenet7:1619-1626RobinsonPJ& RhodesR (2006)Structureofthe 30 nm chromatinfibre:A keyroleforthelinkerhistoneCurrOpinStructBiol16.1BSahaA,WittmeyerJ & CairnsBR(2006)Chromatinremodeling:theindustrialrevolurionof DNAaroundhistonesNatureRevMolCellBiol7 437 446WoodcockCL(2006)Chromatn architectureCurrOpinStructBial16:213-224The Regulationof Chromatin StructureEggerG,Liang,G,AparicioA & JonesPA(2004)Epigeneticsin humandiseaseand prospectsfor epigenetictherapyNature429:457-.463HenikoffS (1990)Posltioneffectvariegationafter60 yearsTrendsGenet6:422-426HenikoffS & AhmadK (2005)Assemblyof varianthistonesintochromatiinAnnuRevCellDevBiol21:,l33-153GasznerM & FelsenfeldG (2006)Insulators:exploitingtranscriptionalandepigeneticmechanismsNatureRevGenet71A3-713HakeSB& AllisCD (2006)HistoneH3variantsandtheirpotentiaroleini n d e x i nmg a m m a l i agne n o r n etsh.
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