Mouse interferon-inducible RNA-dependent protein kinase Pkr gene: cloning and sequence of the 5'-flanking region and functional identification of the minimal inducible promoter.
Mouse interferon-inducible RNA-dependent protein kinase Pkr gene: cloning and sequence of the 5'-flanking region and functional identification of the minimal inducible promoter.
复制标题
小鼠干扰素诱导型 RNA 依赖性蛋白激酶 Pkr 基因：5 侧翼区域的克隆和序列以及最小诱导型启动子的功能鉴定。
DOI：
10.1016/s0378-1119(00)00080-9
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发表时间：
2000
期刊：
影响因子：
3.5
通讯作者：
Samuel,CE
中科院分区：
文献类型：
--
作者：
Tanaka,H;Samuel,CE
RNA依赖的蛋白激酶(PKR)参与干扰素的抗病毒和抗增殖作用。作为我们对小鼠PKR基因外显子-内含子组织结构特征的扩展，我们现在已经分离并鉴定了干扰素诱导转录所需的小鼠PKR启动子区域。瞬时转染分析，使用具有PKR基因5‘侧翼不同片段的报告构建体，导致鉴定出一个功能性的干扰素诱导启动子。缺失分析表明，在最小的44个核苷酸TATA缺失的启动子中存在一个单一的干扰素刺激反应元件(ISRE)；13个核苷酸的ISRE不同于先前描述的ISRE元件，因为3‘-NT是嘌呤而不是嘧啶。紧接在ISRE上游的序列具有15个碱基的KCS元件，该元件在序列和位置上完全保守于鼠和人的PKR启动子之间。在5‘-侧翼区域还存在单个干扰素激活序列(GAS)样元件和参与炎症反应中对各种细胞因子和激素信号的反应的因子的多个识别位点，包括核因子-κB和核因子-IL 6。Northern印迹分析表明，干扰素-α有效地诱导了2.4kb、4.5kb和约4kb的积累。6kb的PKR转录产物，但干扰素-γ和IL-6均不能诱导L细胞中PKR基因的表达。
The RNA-dependent protein kinase (PKR) is implicated in the antiviral and antiproliferative actions of interferon (IFN). As an extension of our structural characterization of the exon–intron organization of the mouse Pkr gene, we now have isolated and characterized the mouse Pkr promoter region required for IFN-inducible transcription. Transient transfection analyses, using reporter constructs possessing various 5′-flanking fragments of the Pkr gene, led to the identification of a functional IFN-inducible promoter. A single IFN-stimulated response element (ISRE) was present in a minimal 44-nt TATA-less promoter identified by deletion analysis; the 13-nt ISRE differed from previously described ISRE elements in that the 3′-nt was a purine instead of a pyrimidine. The sequence immediately upstream of the ISRE possessed the 15-nt KCS element that was exactly conserved in sequence and position between the mouse and human Pkr promoters. A single gamma IFN-activated sequence (GAS)-like element and multiple recognition sites for factors including NF-κB and NF-IL6 involved in responses to various cytokine and hormone signals in inflammatory responses were also present in the 5′-flanking region. Northern blot analysis showed efficient IFN-α induced accumulation of 2.4kb, 4.5kb and approx. 6kb Pkr transcripts, but neither IFN-γ nor IL-6 induced detectable Pkr mRNA accumulation in L cells.