Lipopolysaccharide induction of the tumor necrosis factor-alpha promoter in human monocytic cells - Regulation by Egr-1, c-Jun, and NF-kappa B transcription factors

Biosynthesis of tumor necrosis factor-alpha (TNF-alpha) is predominantly by cells of the monocytic lineage. This study examined the role of various cis-acting regulatory elements in the lipopolysaccharide (LPS) induction of the human TNF-alpha promoter in cells of monocytic lineage, Functional analysis of monocytic THP-1 cells transfected with plasmids containing various lengths of TNF-alpha promoter localized enhancer elements in a region (-182 to -37 base pairs (bp)) that were required for optimal transcription of the TNF-alpha gene in response to LPS, Two regions were identified: region I (-182 to -162 bp) contained an overlapping Sp1/Egr-1 site, and region II (-119 to -88) contained CRE and NF-kappa B (designated kappa B3) sites, In unstimulated THP-1, CRE-binding protein and, to a lesser extent, c-Jun complexes were found to bind to the CRE site, LPS stimulation increased the binding of c-Jun-containing complexes, In addition, LPS stimulation induced the binding of cognate nuclear factors to the Egr-1 and kappa B3 sites, which were identified as Egr-1 and p50/p65, respectively, The CRE and kappa B3 sites in region II together conferred strong LPS responsiveness to a heterologous promoter, whereas individually they failed to provide transcriptional activation, Furthermore, increasing the spacing between the CRE and the kappa B3 sites completely abolished LPS induction, suggesting a cooperative interaction between c-Jun complexes and p50/p65, These studies indicate that maximal LPS induction of the TNF-alpha promoter is mediated by concerted participation of at least two separate cis-acting regulatory elements.