Functional consequences of a polymorphism affecting NF-kappa B p50-p50 binding to the TNF promoter region

Stimulation of the NF-kappaB pathway often causes p65-p50 and p50-p50 dimer s to be simultaneously present in the cell nucleus. A natural polymorphism at nucleotide -863 in the human TNF promoter (encoding tumor necrosis facto r [TNF]) region provides an opportunity to dissect the functional interacti on of p65-p50 and p50-p50 at a single NF-kappaB binding site, We found that this site normally binds both p65-p50 and p50-p50, but a single base chang e specifically inhibits p50-p50 binding. Reporter gene analysis in COS-7 ce lls expressing both p65-p50 and p50-p50 shows that the ability to bind p50- p50 reduces the enhancer effect of this NF-kappaB site. Using an adenoviral reporter assay, we found that the variant which binds p50-p50 results in a reduction of lipopolysaccharide-inducible gene expression in primary human monocytes. This finding adds to a growing body of experimental evidence th at p50-p50 can inhibit the transactivating effects of p65-p50 and illustrat es the potential for genetic modulation of inflammatory gene regulation in humans by subtle nucleotide changes that alter the relative binding affinit ies of different forms of the NF-kappaB complex.