期刊
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
卷 98, 期 15, 页码 8638-8643出版社
NATL ACAD SCIENCES
DOI: 10.1073/pnas.151239498
关键词
signal transduction; NO synthase; iNOS
资金
- NIGMS NIH HHS [F32 GM-19877, R01-GM-52021, F32 GM019877, R01 GM052021] Funding Source: Medline
The human inducible nitric oxide synthase (hiNOS) gene is expressed in several disease states and is also important in the normal immune response. Previously, we described a cytokine-responsive enhancer between -5.2 and -6.1 kb in the 5 ' -flanking hiNOS promoter DNA, which contains multiple nuclear factor KP (NF-kappaB) elements. Here, we describe the role of the IFN-Jak kinase-Stat (signal transducer and activator of transcription) 1 pathway for regulation of hiNOS gene transcription, In A549 human lung epithelial cells, a combination of cytokines tumor necrosis factor-cu, interleukin-1 beta, and lFN-gamma (TNF-alpha, IL-1 beta, and IFN-gamma) function synergistically for induction of hiNOS transcription. Pharmacological inhibitors of Jak2 kinase inhibit cytokine-induced Stat 1 DNA-binding and hiNOS gene expression. Expression of a dominant-negative mutant Stat 1 inhibits cytokine-induced hiNOS reporter expression. Site-directed mutagenesis of a cis-acting DNA element at -5.8 kb in the hiNOS promoter identifies a bifunctional NF-kappaB/Stat 1 motif, In contrast, gel shift assays indicate that only Stat 1 binds to the DNA element at -5.2 kb in the hiNOS promoter. Interestingly, Stat 1 is repressive to basal and stimulated iNOS mRNA expression in 2fTGH human fibroblasts, which are refractory to iNOS induction. Overexpression of NF-KB activates hiNOS promoter-reporter expression in Stat 1 mutant fibroblasts, but not in the wild type, suggesting that Stat 1 inhibits NF-KB function in these cells. These results indicate that both Stat 1 and NF-KB are important in the regulation of hiNOS transcription by cytokines in a complex and cell type-specific manner.
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