Lipopolysaccharide induction of the tumor necrosis factor-alpha promoter in human monocytic cells. Regulation by Egr-1, c-Jun, and NF-kappaB 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-kappaB (designated kappaB3) 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 kappaB3 sites, which were identified as Egr-1 and p50/p65, respectively. The CRE and kappaB3 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 kappaB3 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.
Yao
J
J
Department of Immunology, The Scripps Research Institute, La Jolla, California 92037, USA.
Mackman
N
N
Edgington
T S
TS
Fan
S T
ST
D051698:Activating Transcription Factor 2; D001665:Binding Sites; D002460:Cell Line; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000502:physiology*; D051766:Early Growth Response Protein 1; D004591:Electrophoresis, Polyacrylamide Gel; D005786:Gene Expression Regulation; D006801:Humans; D017874:Immediate-Early Proteins; D008070:Lipopolysaccharides / Q000494:pharmacology*; D009000:Monocytes / Q000378:metabolism*; D016328:NF-kappa B / Q000502:physiology*; D011401:Promoter Regions, Genetic / Q000187:drug effects*; D016755:Proto-Oncogene Proteins c-jun / Q000502:physiology*; D014157:Transcription Factors / Q000378:metabolism / Q000502:physiology*; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis / Q000235:genetics*; D016335:Zinc Fingers
D051698:Activating Transcription Factor 2; D017362:Cyclic AMP Response Element-Binding Protein; D004268:DNA-Binding Proteins; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; D017874:Immediate-Early Proteins; D008070:Lipopolysaccharides; D016328:NF-kappa B; D016755:Proto-Oncogene Proteins c-jun; D014157:Transcription Factors; D014409:Tumor Necrosis Factor-alpha; D004247:DNA