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Localization of the genes for tumor necrosis factor and lymphotoxin between the HLA class I and III regions by field inversion gel electrophoresis.

Medizinische Klinik, University of Tübingen, Federal Republic of Germany.
Immunogenetics (Impact Factor: 2.49). 02/1988; 27(1):66-9. DOI: 10.1007/BF00404447
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    ABSTRACT: Summary Sincea dysregulatedsynthesisoftumor necrosisfactora (TNF-a) may be involvedin the pathogenesisofautoimmune diseases, itwas ofinterest toprecisely locatetherecentlyreported Ncol restriction fragmentlengthpolymorphism (RFLP) oftheTNF-a region.However,by mapping of56.8kb ofoverlappingcosmidclonesanddirectsequencing,we couldlocalize the polymorphicNcol restriction sitewithinthefirstintronoftheTNF-0 gene and notinthe TNF-a gene.To studywhetherregulatorymechanismsareaffected bythispolymorphism,we analyzedtheTNF-a/TNF-0 productionof phytohemagglutinin-stimuated peripheralblood mononuclearcells ofindividuals homozygousfortheTNF-0 NcolRFLP byELISA andconcomitant Northern blotanalysis . On days2-4 afterstimulationwith mitogen,the TNFB*1 allele correspondingtoa5.3-kbNcol fragmentpresentedwithasignificantly higherTNF-0 response. A mRNA analysis demonstratedthathigherproteinlevels ofTNF-0 correlate alsowithincreased amountsofTNF-0 transcripts .No allelic association wasfoundinrespecttoTNF-a production. Tofurtherinvestigate apossible allelic influence on transcription, we determinedtheDNA sequence of2kb ofthe5'portionofourclonedTNFB*2 allele andcompareditwiththeavailable TNF-0 sequences .By computer-aided recognition motifsearchofDNA bindingfactors, we reportputative bindingsitesconservedbetweenmouse and man inthe5'flankingregionaswellasinintron 1oftheTNF-0 gene,foundalsoinothercytokinepromotersequences .Inaddition, bypolymerase chainreactionamplification andsequencingof740by ofthe5'partofTNF-0 ofindividuals typedhomozygouslyfortheNcol RFLP, we couldshow thataminoacidposition26isconserved asasparagineintheTNFB *1andasthreonineintheTNFB *2sequence.A previouslyreported, EcoRI RFLP inthe3'untranslated regionofTNF-0 doesnotsegregatewitheitherofthetwo alleles .Thus,fourTNFB alleles cande definedattheDNA level .
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    Scandinavian Journal of Immunology 12/1988; 28(5):599-606. DOI:10.1111/j.1365-3083.1988.tb01492.x · 1.88 Impact Factor