Transcriptional regulation of human inducible nitric oxide synthase (NOS2) gene by cytokines: initial analysis of the human NOS2 promoter.

Abstract

The expression of inducible nitric oxide synthase (NOS2) is complex and is regulated in part by gene transcription. In this investigation we studied the regulation of NOS2 in a human liver epithelial cell line (AKN-1) which expresses high levels of NOS2 mRNA and protein in response to tumor necrosis factor a, interleukin 113, and interferon y (cytokine mix, CM). Nuclear run-on analysis revealed that CM transcript"onally activated the human NOS2 gene. To delineate the cytokine-responsive regions of the human NOS2 promoter, we stimulated AKN-1 cells with CM following transfection of NOS2 luciferase constructs. Analysis of the first 3.8 kb upstream of the NOS2 gene demonstrated basal promoter activity but failed to show any cytokine-inducible activity. However, 3-to 5-fold inductions of luciferase activity were seen in constructs extending up to -5.8 and -7.0 kb, and a 10-fold increase was seen upon transfection of a -16 kb construct.' Further analysis of various NOS2 luciferase constructs ligated upstream of the thymidine kinase promoter identified three regions containing cytokine-responsive elements in the human NOS2 gene: -3.8 to -5.8, -5.8 to -7.0, and -7.0 to -16 kb. These results are in marked contrast with the murine macrophage NOS2 promoter in which only 1 kb of the proximal 5' flanking region is necessary to confer inducibility to lipopolysaccharide and interferon y. These data demonstrate that the human NOS2 gene is transcriptionally regulated by cytokines and identify multiple cytokineresponsive regions in the 5' flanking region of the human NOS2 gene.

Central Role of Double-Stranded RNA-Activated Protein Kinase in Microbial Induction of Nitric Oxide Synthase

“…The molecular basis for induction of the human NOS2 gene is only partially understood (33,43). In contrast, regions in the murine macrophage NOS2 promotor essential for conferring inducibility of NOS2 to LPS and IFN-␥ have been well defined (44,45).…”

TP53 mutations and hepatocellular carcinoma: insights into the etiology and pathogenesis of liver cancer

“…During chronic viral hepatitis, the upregulation of certain proinflammatory cytokines, like tumor necrosis factor-a (TNF-a) and interferon-g (IFN-g), has been repeatedly demonstrated (Gonzalez-Amaro et al, 1994;Mihm et al, 1996). These proinflammatory cytokines, TNF-a, IFN-g and IL-1, induce NOS2 gene expression, which leads to increased NO concentrations in human hepatocytes (de Vera et al, 1996;Laskin et al, 1998). In addition, NOS2 is also induced directly by the HBV and HCV.…”

Haplotypic relationship between SNP and microsatellite markers at the NOS2A locus in two populations

“…The NOS2A gene is predominantly transcriptionally regulated 35,36 and so genetic variation within the 5 0 region may influence gene expression. The proximal NOS2A promoter contains several SNPs 37,38 and a pentanucleotide microsatellite (the 'NOS2A À2.6 microsatellite').…”