Polymorphic structure of the tumor necrosis factor (TNF) locus: an NcoI polymorphism in the first intron of the human TNF-beta gene correlates with a variant amino acid in position 26 and a reduced level of TNF-beta production.
Open Access
- 1 January 1991
- journal article
- Published by Rockefeller University Press in The Journal of Experimental Medicine
- Vol. 173 (1), 209-219
- https://doi.org/10.1084/jem.173.1.209
Abstract
Since a dysregulated synthesis of tumor necrosis factor alpha (TNF-alpha) may be involved in the pathogenesis of autoimmune diseases, it was of interest to precisely locate the recently reported NcoI restriction fragment length polymorphism (RFLP) of the TNF-alpha region. However, by mapping of 56.8 kb of overlapping cosmid clones and direct sequencing, we could localize the polymorphic NcoI restriction site within the first intron of the TNF-beta gene and not in the TNF-alpha gene. To study whether regulatory mechanisms are affected by this polymorphism, we analyzed the TNF-alpha/TNF-beta production of phytohemagglutinin-stimulated peripheral blood mononuclear cells of individuals homozygous for the TNF-beta NcoI RFLP by ELISA and concomitant Northern blot analysis. On days 2-4 after stimulation with mitogen, the TNFB*1 allele corresponding to a 5.3-kb NcoI fragment presented with a significantly higher TNF-beta response. A mRNA analysis demonstrated that higher protein levels of TNF-beta correlate also with increased amounts of TNF-beta transcripts. No allelic association was found in respect to TNF-alpha production. To further investigate a possible allelic influence on transcription, we determined the DNA sequence of 2 kb of the 5' portion of our cloned TNFB*2 allele and compared it with the available TNF-beta sequences. By computer-aided recognition motif search of DNA binding factors, we report putative binding sites conserved between mouse and man in the 5' flanking region as well as in intron 1 of the TNF-beta gene, found also in other cytokine promoter sequences. In addition, by polymerase chain reaction amplification and sequencing of 740 bp of the 5' part of TNF-beta of individuals typed homozygously for the NcoI RFLP, we could show that amino acid position 26 is conserved as asparagine in the TNFB*1 and as threonine in the TNFB*2 sequence. A previously reported, EcoRI RFLP in the 3' untranslated region of TNF-beta does not segregate with either of the two alleles. Thus, four TNFB alleles can be defined at the DNA level.Keywords
This publication has 50 references indexed in Scilit:
- Regulation of tumor necrosis factor alpha transcription in macrophages: involvement of four kappa B-like motifs and of constitutive and inducible forms of NF-kappa B.Molecular and Cellular Biology, 1990
- Molecular cloning and expression of a receptor for human tumor necrosis factorCell, 1990
- Molecular cloning and expression of the human 55 kd tumor necrosis factor receptorCell, 1990
- Heritable major histocompatibility complex class II-associated differences in production of tumor necrosis factor alpha: relevance to genetic predisposition to systemic lupus erythematosus.Proceedings of the National Academy of Sciences, 1990
- Primer-Directed Enzymatic Amplification of DNA with a Thermostable DNA PolymeraseScience, 1988
- A conserved AU sequence from the 3′ untranslated region of GM-CSF mRNA mediates selective mRNA degradationCell, 1986
- Genomic sequencing.Proceedings of the National Academy of Sciences, 1984
- A technique for radiolabeling DNA restriction endonuclease fragments to high specific activityAnalytical Biochemistry, 1983
- Isolation of biologically active ribonucleic acid from sources enriched in ribonucleaseBiochemistry, 1979
- DNA sequencing with chain-terminating inhibitorsProceedings of the National Academy of Sciences, 1977