Autochthonous Mouse Melanoma and Mammary Tumors do not Express the Pluripotency Genes Oct4 and Nanog.docVIP

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Autochthonous Mouse Melanoma and Mammary Tumors do not Express the Pluripotency Genes Oct4 and Nanog.doc

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Autochthonous Mouse Melanoma and Mammary Tumors do not Express the Pluripotency Genes Oct4 and Nanog

AutochthonousMouseMelanomaandMammaryTumors donotExpressthePluripotencyGenesOct4andNanog CarolineSchreiber1*.,VanessaKuch1.,ViktorUmansky3,JonathanP.Sleeman1,2 1CentreforBiomedicineandMedicalTechnologyMannheim(CBTM),MedicalFacultyMannheim,UniversityHeidelberg,Mannheim,Germany,2InstituteforToxicology andGenetics,KITCampusNord,Karlsruhe,Germany,3SkinCancerUnit,GermanCancerResearchCenter,HeidelbergandDepartmentofDermatology,Venereologyand Allergology,UniversityMedicalCenterMannheim,Ruprecht-KarlUniversityofHeidelberg,Mannheim,Germany Abstract ThehomeodomaintranscriptionfactorsOct4andNanogmaintainpluripotencyandself-renewalinembryonicstemcells.In somatic cells, inappropriate expression of these genes has been associated with loss of differentiation, malignant transformation,andtheacquisitionofcancerstemcell-likeproperties.Ascancerstemcellshavebeensuggestedtounderlie thegrowthandmalignancyoftumors,Oct4andNanogmayrepresenttherapeutictargets.Theirexpressioncouldalsoact asamarkerofthecancerstemcellpopulation,permittingitsisolationandcharacterisation.Nevertheless,theexistenceof multiplepseudogenesandisoformsofthesegeneshascomplicatedtheinterpretationofthedatathatsupportsarolefor Oct4andNanoginthecancercontext.HereweaddressedthisissueusingknockinmiceinwhichIRESelementsareusedto allowGFPexpressionunderthecontroloftheendogenousOct4orNanogpromoters,whilemaintainingcorrectexpression oftheOct4orNanoggene.ThesemicewerecrossedwithMT/retmicethatdevelopmelanomas,andwithMMTV-PyMT mice and MMTV-Neu mice that develop mammary adenocarcinomas. We analysed the tumors that developed in these compoundmiceforGFPexpression.InthiswaywecouldassesstranscriptionofOct4andNanoginautochthonouscancers withoutthecomplicationoffactorssuchaspseudogeneexpression,alternativesplicingandantibodyspecificity.Boththe Oct4andNanogknockintumor-bearingmiceexpressedGFPinblastocystsandtestesasexpected.However,wecouldfind no evidence for expression of the GFP reporter above background levels in tumors using FACS,