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TheFASEBJournal ResearchCommunication

EssentialroleofRac1andRac3GTPasesinneuronaldevelopment

SaraCorbetta,*,1SaraGualdoni,*,1GabrieleCiceri,*MartaMonari,*EmanuelaZuccaro,*VictorL.J.Tybulewicz, andIvandeCurtis*,2

*CellAdhesionUnit,Dibit,SanRaffaeleScienti cInstituteandSanRaffaeleUniversity,Milan,Italy;and DivisionofImmuneCellBiology,NationalInstituteforMedicalResearch,TheRidgeway,MillHill,London,UK

ABSTRACT

RacGTPasesaremembersoftheRho

familyregulatingtheactincytoskeletonandimplicatedinneuronaldevelopment.UbiquitousRac1andneu-ron-speci cRac3GTPasesarecoexpressedinthedevelopingmammalianbrain.WeusedCre-mediatedconditionaldeletionofRac1inneuronscombinedwithknockoutofneuron-speci cRac3tostudytheroleoftheseGTPasesinneuraldevelopment.Wefoundthatlackofbothgenescausesmotorbehavioraldefects,epilepsy,andprematuredeathofmice.DeletionofeitherGTPasedoesnotproduceevidentphenotypes.Double-knockoutmiceshowspeci cdefectsinthedevelopmentofthehippocampus.Selectiveimpair-mentofthedorsalhilusofdouble-knockoutanimalsisassociatedwithalterationintheformationofthehippocampalcircuitry.Axonalpathwaystoandfromthedorsalhilusareaffectedbecauseofthede citofhilarmossycells.Moreover,analysisofRacfunctioninhippocampalculturesshowsthatspineformationisstronglyhamperedonlyinneuronslackingbothRacproteins.These ndingsshowforthe rsttimethatbothRac1andRac3areimportantforthedevelopmentofthenervoussystem,whereintheyplaycomplemen-taryrolesduringlatestagesofneuronalandbraindevelopment.—Corbetta,S.,Gualdoni,S.,Ciceri,G.,Monari,M.,Zuccaro,E.,Tybulewicz,V.L.J.,deCurtis,I.EssentialroleofRac1andRac3GTPasesinneuronaldevelopment.FASEBJ.23,1347–1357(2009)Keyaxons

Words:hippocampus hilarmossycells dendriticspines RacGTPasesaremembersoftheRhofamilyregulat-ingactindynamics(1).Evidenceindicatingtheinvolve-mentofRacproteinsindifferentaspectsofneuronaldevelopmentincludesformationofaxonandden-drites,axonalnavigation,andsynaptogenesis(2,3).TherearethreeRacgenesinvertebratesencodingtheRac1,Rac2,andRac3proteins,sharing88-92%se-quenceidentity.Rac1andRac3arecoexpressedinneurons(4),andRac2isexpressedinhematopoieticcells(5).Moststudiesonvertebrateneuronaldevelop-menthavefocusedonRac1,whereasveryfewstudiesexistonRac3,despiteitsneuronalcoexpressionwithRac1(4).

0892-6638/09/0023-1347©FASEBRac1isubiquitous,andRac1-knockoutmiceareearlyembryoniclethal,becauseoftherequirementforthisGTPaseinearlydevelopment(6).Todate,theanalysisoftransgenicmiceexpressingmutatedformsofRac1hasshowneffectsonaxonterminalsanddendriticspinesinPurkinjecells(7).Althoughstudieswithmutantshavebeeninformativetostartassemblingapictureofhowtheseproteinsworktobuildtheverte-bratenervoussystem,itisclearthatconclusionsfromthesestudiesmustbetakenwithcaution,becauseRac1mutantsmayinterferewiththefunctionofrelatedGTPases.Recently,earlyconditionaldeletion(8)andknockdowninvitro(9)havecon rmedaroleofRac1inaxonguidanceinvivoanddendritogenesisinvitro,respectively.

Rac3was rstidenti edinhumans(10)andinbirdsasRac1B(11).TheRac3proteinisspeci callyex-pressedinthenervoussystem(11)andisdevelopmen-tallyregulatedinavianandmousebrains,withapeakattimesofintenseneuritebranchingandsynaptogenesis(12,13).Rac3-knockoutmiceareviable(4),andrecent ndingsindicatearoleofRac3incognitivedevelop-ment(14).

TherelativecontributionsofRac1andRac3tothedevelopmentofthevertebratenervoussystemareun-known.Rac1 oxmiceallowedtheanalysisofRac1functionafterconditionaldeletionofthegeneintheimmunesystem(15,16)andinearlybraindevelop-ment(8).Ontheotherhand,theappearanceofRac3 rstinvertebrates(3)suggeststhatthisgenemayplayaroleinthelatedevelopmentofbrainsofhighercomplexity.ThepossibilityoffunctionalinteractionsbetweenRac1andRac3duringthedevelopmentofthevertebratenervoussystemhasneverbeenaddressed.BycombiningconditionalmutagenesisoftheRac1genewiththedeletionoftheRac3gene,weshowherethatbothproteinsareimportantforthelatedevelopmentofthebrainandplaykeyfunctionsinlaterstagesofneuronalmaturation.

1

2Theseauthorscontributedequallytothiswork.

ti cCorrespondence:CellAdhesionOlgettinaInstituteandUniversityVita-SaluteUnit,SanSanRaffaeleRaffaele,Scien-doi:10.1096/fj.08-121574

58-20132Milano,Italy.E-mail:decurtis.ivan@hsr.itVia1347