Transcriptional regulation of human inducible nitric oxide synthase (NOS2) gene by cytokines: Initial analysis of the human NOS2 promoter
Transcriptional regulation of human inducible nitric oxide synthase (NOS2) gene by cytokines: Initial analysis of the human NOS2 promoter
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细胞因子对人类诱导型一氧化氮合酶（NOS2）基因的转录调控： 对人类 NOS2 启动子的初步分析
DOI：
10.1073/pnas.93.3.1054
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发表时间：
1996-02-06
影响因子：
11.1
通讯作者：
Geller, DA
中科院分区：
文献类型：
--
作者：
deVera, ME;Shapiro, RA;Geller, DA
诱导型一氧化氮合酶（inducible nitric oxide synthase，NOS 2）的表达是复杂的，部分受基因转录调控。本研究中，我们研究了人肝上皮细胞系（AKN-1）中NOS 2的调控，该细胞系在肿瘤坏死因子α、白细胞介素1 β和干扰素γ（cytokine mix，CM）的作用下表达高水平的NOS 2 mRNA和蛋白。为了描述人NOS 2启动子的精氨酸应答区，我们在转染NOS 2荧光素酶构建体后用CM刺激AKN-1细胞，对NOS 2基因上游前3.8kb的分析显示了基础启动子活性，但没有显示任何精氨酸诱导的活性。然而，在延伸至约5.8和约7.0 kb的构建体中观察到荧光素酶活性的3至5倍诱导，并且在转染约16 kb构建体时观察到10倍增加。对连接在胸苷激酶启动子上游的各种NOS 2荧光素酶构建体的进一步分析鉴定了人NOS 2基因中含有精氨酸应答元件的三个区域：这些结果与鼠巨噬细胞NOS 2启动子形成鲜明对比，在鼠巨噬细胞NOS 2启动子中，只有1 kb的近端5 ′侧翼区是赋予脂多糖和干扰素γ诱导所必需的。这些数据表明，人NOS 2基因是由细胞因子转录调节，并确定在人NOS 2基因的5'侧翼区的多个精氨酸反应区。
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 alpha, interleukin 1 beta, and interferon gamma (cytokine mix, CM). Nuclear run-on analysis revealed that CM transcriptionally 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 gamma. These data demonstrate that the human NOS2 gene is transcriptionally regulated by cytokines and identify multiple cytokine-responsive regions in the 5' flanking region of the human NOS2 gene.