title: Functional analysis of linker-scan mutants spanning the -376, -308, -244, and -238 polymorphic sites of the TNF-alpha promoter.
abstract: Tumor necrosis factor alpha (TNF-alpha) 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-alpha promoter constructs containing single nucleotide changes. We have now made a series of mutant constructs in which regions of the TNF-alpha promoter containing suspected functional single nucleotide polymorphisms, including -376, -308, -244 and -238, were replaced by a 10 bp 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-alpha 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.
authors.affiliation: Department of Rheumatology, Leiden University Medical Center, 2300 RC Leiden, The Netherlands.
authors.forename: J P
authors.initials: JP
authors.lastname: Bayley
authors.forename: H
authors.initials: H
authors.lastname: de Rooij
authors.forename: P J
authors.initials: PJ
authors.lastname: van den Elsen
authors.forename: T W
authors.initials: TW
authors.lastname: Huizinga
authors.forename: C L
authors.initials: CL
authors.lastname: Verweij
mesh_terms: D001402:B-Lymphocytes / Q000166:cytology; D001483:Base Sequence; D006801:Humans; D019169:Jurkat Cells; D008213:Lymphocyte Activation; D008969:Molecular Sequence Data; D009154:Mutation; D011110:Polymorphism, Genetic; D020641:Polymorphism, Single Nucleotide; D011401:Promoter Regions, Genetic; D013601:T-Lymphocytes / Q000166:cytology; D014158:Transcription, Genetic; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D020298:U937 Cells
chemical_list: D014409:Tumor Necrosis Factor-alpha