Relevance of the tumor necrosis factor alpha (TNF alpha) -308 promoter polymorphism in TNF alpha gene regulation.
Tumor necrosis factor alpha (TNF alpha) is a central mediator of the immunological response and the location of the gene within the major histocompatibility complex (MHC) has prompted much speculation about the role of TNF alpha alleles in inflammatory and MHC-associated autoimmune diseases. A G to A transition polymorphism at position -308 of the TNF alpha promoter/enhancer region has been described. The uncommon -308A allele was shown to be strongly associated with human leukocyte antigen (HLA)-DR3, known to be related to a TNF alpha "high producer" phenotype. In support for a clinical relevance, the -308A allele is implicated in susceptibility for cerebral malaria. In this study, we determined the junctional consequences of the TNF -308 polymorphism. Therefore, we analyzed both allelic forms (TNF alpha(-308G) and TNF alpha(-3O8A)) of the TNF alpha enhancer/promoter region (-598/+108) in a transient transfection system, using chloramphenicol acetyltransferase (CAT) as reporter gene. The T cell line Jurkat and the B cell line Raji served as hosts in these experiments. The results showed no differences in the level of inducible reporter gene expression between the TNF(-3O8G)/CAT and the TNF(-308A)/CAT constructs. These data were confirmed by allele specific TNF alpha transcript quantification (ASTQ) analysis, which demonstrated that both TNF alleles contribute equally to the total amount of mRNA in peripheral blood mononuclear cells (PBMCs) stimulated with phorbol 12-myristate 13-acetate (PMA)/anti-CD3. In analogy, no difference between the level of transcription of the -308A and -308G alleles was observed in lipopolysaccharide (LPS)-stimulated peripheral blood monocytes. This study indicates that the TNF alpha -308 G to A transition is not responsible for differential TNF alpha production induced by standard in vitro stimuli.
Brinkman
B M
BM
Department of Rheumatology, University Hospital, Leiden, The Netherlands.
Zuijdeest
D
D
Kaijzel
E L
EL
Breedveld
F C
FC
Verweij
C L
CL
D000483:Alleles; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D004742:Enhancer Elements, Genetic; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000737:chemistry; D008070:Lipopolysaccharides / Q000494:pharmacology; D010957:Plasmids; D011110:Polymorphism, Genetic; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000032:analysis / Q000378:metabolism; D013601:T-Lymphocytes / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics*
D008070:Lipopolysaccharides; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D015500:Chloramphenicol O-Acetyltransferase; D013755:Tetradecanoylphorbol Acetate