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

• Published in: The Journal of biological chemistry 1997-07, Vol.272 (28), p.17795-17801
• Main Authors: Yao, Jin, Mackman, N, Edgington, T S, Fan, Sao-Tah
• Format: Article
• Language: English
• ISSN: 0021-9258
• DOI: 10.1074/jbc.272.28.17795

Biosynthesis of tumor necrosis factor- alpha (TNF- alpha ) is pre-dominantly 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.