affects intrinsic protein stability. C-terminal PEST domain by casein kinase II Phosphorylation of IkappaBalpha in the

NF-kBproteinsareafamilyofpleiotropictranscriptionfac-torsthatregulateexpressionofnumerouscytokines,andviralgenes(forreviews,seereferences27and48).Incells,NF-kBispresentinaninactivecytoplasmicform;localizationofthep50-p65heterodimericformofNF-kBtothecytoplasmismediatedbytheinhibitoryIkBa proteinthatbindstoandmasksthenuclearlocalizationsignalofthep65subunitofNF-kB(5,7).Activatingagents,suchasviruses,phorbolesters,cytokines,radicaloxygenintermediates,andbacteriallipo-polysaccharide,promotethedissociationofthecytosolicNF-kB/IkBcomplexes,inpartbyactivatingcellularkinasesthatphosphorylateIkB(5–7,22,27,48).TheDNA-bindingNF-kBfamilymembersshareaRelho-mologydomainthatisresponsibleforDNAbinding,nuclearlocalization,andproteindimerization.DNA-bindingmembersofNF-kB/Relincludep50(NFKB1)(9,23,41),p65(RelA)(52,58),c-Rel(13,24),p52(NFKB2orLyt-10)(8,51,62),RelB(I-Rel)(59,60),anddorsal(65).p50andp52aresyn-thesizedasprecursorsofp105andp100,respectively,thatareproteolyticallyprocessedtogenerateactiveDNA-bindingp50andp52(8,9,23,41,51,62).DifferentNF-kBdimersbindtovariantNF-kBsites(44)(consensus,59-GGGANNYYCC-39)presentinthepromoterregionsofmanygenes(reviewedinreferences2and48).TheintracellularlocalizationandposttranslationalactivityofNF-kB/Relproteinsareregulatedbytheankyrinrepeat-containingI kBproteins(I Ba,Ig,Bcl-3,p105,andp100)(reviewedinreferences5,25,28,and39).PhosphorylationbyactivatedcellularkinasesandsubsequentdegradationofIkBaappeartobecriticalregulatorystepsinthecontrolofNF-kBactivation.Recentstudiesdemonstratedthatmutationofei-therSer-32orSer-36blockedsignal-inducedIkBa phosphor-ylationanddegradation(10,11,69),andatruncatedIkBa thatwasmissing45aminoacidsattheNterminuscorrectedradi-ationsensitivityinataxiatelangiectasiacells(37).Further-more,IkBa transcriptionisupregulatedbyNF-kB,sincetheIkBa promotercontainsfunctionalNF-kBsites.Thus,anau-toregulatoryloopisestablished,andtheultimatelyincreasedIkBa concentrationslimitthenucleartranslocationofNF-kBproteins(12,18,45,66).Inrecentstudies,disruptionofIkBaautoregulationwithIkBa antisenseRNAresultedinmalignanttransformationandalteredNF-kB-regulatedgeneactivity,in-dicatingthatIkBa mayrepresentapotentialgrowthsuppres-soractivity(4).TofurthercharacterizethesignalingeventsthatcontributetoIkBa phosphorylationanddegradation,akinaseactivitythatspecificallyinteractedwithIkBa inanaffinitychromatog-raphystepandphosphorylatedIkBa withhighspecificityinvitrowasisolatedfromJurkatTcells.Biochemicalandimmu-nologicalcross-reactivityexperimentsidentifiedthekinaseac-tivityascaseinkinaseII(CKII).Pointmutationanalysisre-vealedthatatriplemutationofresiduesT-291,S-283,andT-299eliminatedphosphorylationofIkBa bytheCKIIinvitro