现代泌尿外科杂志·基础研究·TFE3基因易位相关性肾癌中TFE3融合蛋白亚细胞定位失调的机制研究潘骏，陈怡2.3.4，杨磊2.3.4，刘宁5，逐艳文，马文亮"，甘卫东"，李冬梅2.3.4（1.南京大学医学院附属鼓楼医院泌尿外科，江苏南京210008；2.南京大学医学院免疫与生殖生物学实验室，江苏南京210093；3.南京市江苏省医学分子技术重点实验室，江苏南京210093；4.南京大学生命分析化学国家重点实验室，江苏南京210093；5.南京医科大学附属南京医院泌尿外科，江苏南京210006）MechanismofdisorderedsubcellularlocalizationofTFE3fusionproteininTFE3translocationrenalcellcarcinomaPANJun',CHENYi?,YANGLei?,LIUNing",LUYanwen',MAWenliang',GANWeidong',LIDongmei?(1.DepartmentofUrology,AffiliatedDrumTowerHospitalofMedicalSchoolofNanjingUniversity,Nanjing210008;2.LaboratoryofImmunologyandReproductionBiology,NanjingUniversitySchoolofMedicine,Nanjing210093;3.JiangsuKeyLaboratoryofMolecularMedicine,NanjingUniversity,Nanjing210093;4.StateKeyLaboratoryofAnalyticalChemistryforLifeScience,NanjingUniversity,Nanjing210093;5.DepartmentofUrology,NanjingFirstHospital,NanjingMedicalUniversity,Nanjing210006,China)ABSTRACT:ObjectiveToinvestigatetheeffectsofthelossofexon1ofTFE3onnuclearlocalizationofchimericTFE3proteininTFE3translocationrenalcellcarcinoma(TFE3tRCC).MethodsThelocalizationofTFE3proteininTFE3tRCCandclearcellrenalcellcarcinoma(ccRCC)weredetectedwithimmunochemistry.TheexonretentionofTFE3geneinTFE3tRCCwasanalyzedindatabasesandliteratures.TheplasmidscontainingTFE3full-lengthanddifferent-lengthofTFE3exonswhichwereconstructedtopCDH-MCS-EGFP-PuroweretransfectedintoHEK293TusingLipoFiterTM.ThelocalizationofEGFPproteininHEK293Tcellsweredetectedwithconfocalmicroscopy.ThelocalizationofTFE3proteinandtruncatedTFE3proteinweredetectedwithWesternblotting.ThemRNAexpressionofthedownstreamgenesofTFE3proteinweredetectedwithq-PCR.ResultsStrongnuclearsignalofTFE3proteinwasobservedinTFE3tRCC,whereasTFE3proteininccRCCwasmainlylocalizedincytoplasm.TheresultsoffluorescenceimagingandWesternblottingshowedthatTFE3full-lengthproteinwasexpressedbothinnucleus...
