J Biol Chem, Vol. 274, Issue 31, 21714-21718, July 30, 1999

Human Tumor Necrosis Factor- Gene 3' Untranslated Region Confers Inducible Toxin Responsiveness to Homologous Promoter in Monocytic THP-1 Cells

From the Division of Immunology and Allergy, Department of Medicine, Centre Hospitalier Universitaire Vaudois, CH-1011 Lausanne, Switzerland

To better define the role of 3' untranslated region (3'UTR) on transcriptional regulation of the human tumor necrosis factor (TNF)- gene, monocytic human THP-1 cells were transfected with two TNF- promoter constructs spanning base pairs 1897/1 and 1214/1, respectively, and linked to the rabbit -globin gene. Quantitative globin gene expression of chimerae was measured by reverse transcription-polymerase chain reaction. A construct linking the chicken -actin promoter and a deleted portion of the -globin gene was cotransfected and used as internal standard. Unexpectedly, when THP-1 cells were stimulated with lipopolysaccharide or toxic shock syndrome toxin-1, gene regulation was hardly detected. In contrast, endogenous TNF- gene regulation measured by the same reverse transcription-polymerase chain reaction procedure was vigorous. Remarkably, ligation of 3'UTR to chimeric constructs led to a drastic drop in the basal level of chimeric gene expression, resulting in a 15- to 40-fold induction of the reporter gene. Consistently, when the TNF- promoter was replaced by the cytomegalovirus early immediate promoter, gene expression was also uniformly reduced but was no longer up-regulated upon stimulation with lipopolysaccharide and toxic shock syndrome toxin-1. These data provide the first line of evidence that, in addition to its role in TNF- transcript stability and translation, human TNF- 3'UTR also participates in modulating gene expression at the transcriptional level.