Transcriptional regulator CTCF controls human interleukin 1 receptor-associated kinase 2 promoter.
Transcriptional regulator CTCF controls human interleukin 1 receptor-associated kinase 2 promoter.
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转录调节因子 CTCF 控制人白细胞介素 1 受体相关激酶 2 启动子。
DOI：
10.1016/j.jmb.2004.11.066
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发表时间：
2005
影响因子：
5.6
通讯作者：
Lerman,MichaelI
中科院分区：
文献类型：
--
作者：
Kuzmin,Igor;Geil,Laura;Gibson,Lauren;Cavinato,Tiziana;Loukinov,Dmitry;Lobanenkov,Victor;Lerman,MichaelI
对入侵病原体的免疫应答主要通过跨膜toll样受体家族介导，并由许多下游效应器调节。特别是，一个由四个白介素1受体相关激酶（IRAK）组成的家族调节对细菌内毒素的反应性。针对特定的IRAK成分的药理学靶向可能有利于治疗细菌感染。在这里，我们研究了人类IRAK2基因的转录调控。对IRAK2启动子区域的分析揭示了几种转录因子的推定结合位点，包括ZIP （EGR1和SP1）、CTCF和ap -2 β。在naïve和内毒素处理的单核细胞、休眠和活化的Jurkat t细胞、肺和肾细胞中，ZIP或AP-2位点的缺失对IRAK2启动子活性没有显著影响。相反，我们发现CTCF在IRAK2转录中起主要作用。对含有IRAK2 CpG岛的DNA片段进行的电泳迁移迁移分析显示，转录调节因子和染色质绝缘体蛋白CTCF具有单一的高亲和力结合位点。该分析在小鼠Irak2启动子中发现了ctcf结合位点。用染色质免疫沉淀法证实了CTCF蛋白在人IRAK2启动子中的存在。通过甲基化干扰试验鉴定了与CTCF蛋白相互作用的特异性残基。在所分析的所有细胞系中，包括肺细胞、肾细胞、单核细胞和t细胞来源的细胞，IRAK2荧光素酶报告结构含有一个完整的ctcf结合位点，显示出很强的启动子活性。然而，对于ctcf结合位点突变的构建体，IRAK2启动子活性显著降低。
Immune responses to invading pathogens are mediated largely through a family of transmembrane Toll-like receptors and modulated by a number of downstream effectors. In particular, a family of four interleukin 1 receptor-associated kinases (IRAK) regulates responsiveness to bacterial endotoxins. Pharmacological targeting of particular IRAK components may be beneficial for treatment of bacterial infections. Here, we studied transcriptional regulation of the human IRAK2 gene. Analysis of the IRAK2 promoter region reveals putative binding sites for several transcriptional factors, including ZIP (EGR1 and SP1), CTCF and AP-2beta. Deletion of the ZIP or AP-2 sites did not significantly affect IRAK2 promoter activity in naïve and endotoxin-treated mononuclear cells, in dormant and activated Jurkat T-cells, in lung and kidney cells. In contrast, we found that CTCF plays a major role in IRAK2 transcription. An electrophoretic mobility shift assay of the DNA fragments containing the IRAK2 CpG island, revealed a single high-affinity binding site for the transcriptional regulator and a chromatin insulator protein, CTCF. This assay revealed a CTCF-binding site within the mouse Irak2 promoter. The presence of the CTCF protein in human IRAK2 promoter was confirmed by chromatin immunoprecipitation assay. Specific residues that interacted with the CTCF protein, were identified by methylation interference assay. In all cell lines analyzed, including cells of lung, renal, monocytic and T-cell origin, the IRAK2 luciferase reporter construct, containing an intact CTCF-binding site, showed strong promoter activity. However, IRAK2 promoter activity was decreased dramatically for the constructs with a mutated CTCF-binding site.