FUNCTIONAL ANALYSIS OF LINKER-SCAN MUTANTS SPANNING THE −376, −308, −244, AND −238 POLYMORPHIC SITES OF THE TNF-α PROMOTER

• Published in: Cytokine (Philadelphia, Pa.) Pa.), 2001-06, Vol.14 (6), p.316-323
• Main Authors: Bayley, Jean-Pierre, de Rooij, Henny, van Den Elsen, Peter J., Huizinga, Tom W.J., Verweij, Cornelis L.
• Format: Article
• Language: English
• Subjects: B-Lymphocytes - cytology; Base Sequence; Humans; Jurkat Cells; Lymphocyte Activation; Molecular Sequence Data; Mutation; Polymorphism, Genetic; Polymorphism, Single Nucleotide; Promoter Regions, Genetic; T-Lymphocytes - cytology; TNF-α/polymorphisms/functional/TRS/repressor; Transcription, Genetic; Transfection; Tumor Necrosis Factor-alpha - chemistry; Tumor Necrosis Factor-alpha - genetics; Tumor Necrosis Factor-alpha - metabolism; U937 Cells
• ISSN: 1043-4666; 1096-0023
• DOI: 10.1006/cyto.2001.0902

Tumor necrosis factor alpha (TNF-α) promoter polymorphisms have been linked to a large number of diseases but studies examining the possible direct functional effects of these polymorphisms have been contradictory. Previous studies compared TNF-α promoter constructs containing single nucleotide changes. We have now made a series of mutant constructs in which regions of the TNF-α promoter containing suspected functional single nucleotide polymorphisms, including −376, −308, −244 and −238, were replaced by a 10bp linker scan sequence. These constructs were transiently transfected into the T cell line Jurkat, the B cell line Raji, and the monocytic cell line U937, and tested for basal and induced transcriptional activity. Mutant constructs covering both the −308 and −376 polymorphisms showed no significant differences in either basal or induced transcriptional activity. Constructs covering the −244/−238 region showed a small increase in basal activity in the U937 cell line. These results indicate (i) that the −308 and −376 regions are of no functional relevance for TNF-α promoter transcription, and (ii) that the −244/−238 region does not influence transcription in some cell lines but may have some role in transcription in others.