DNA Triplex Formation Selectively Inhibits Granulocyte-Macrophage Colony-stimulating Factor Gene Expression in Human T Cells*
DNA Triplex Formation Selectively Inhibits Granulocyte-Macrophage Colony-stimulating Factor Gene Expression in Human T Cells*
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DNA 三链体的形成选择性抑制人类 T 细胞中的粒细胞-巨噬细胞集落刺激因子基因表达*
DOI：
10.1074/jbc.271.24.14438
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发表时间：
1996
期刊：
影响因子：
--
通讯作者：
M. Frances Shannon
中科院分区：
文献类型：
--
作者：
M. Kochetkova;M. Frances Shannon
粒细胞-巨噬细胞集落刺激因子（GM-CSF）是一种造血生长因子，在活化的T细胞、成纤维细胞、巨噬细胞和内皮细胞中表达。虽然GM-CSF似乎不是正常造血所必需的，但GM-CSF的过表达已涉及某些疾病如髓性白血病和慢性炎症的发病机制。GM-CSF启动子内的NF-κB/Rel结合位点（称为κB元件）似乎对于响应T细胞中的许多刺激而在报告基因测定中控制表达是重要的。我们研究了在这个调控序列中，作为抑制GM-CSF基因转录的潜在工具的三螺旋形成。将15个碱基的寡核苷酸GM 3靶向GM-CSF近端启动子中的富含嘌呤的区域，该区域与κB元件重叠。凝胶迁移率变动分析和DNA酶I足迹法表明，GM 3形成了一个序列特异性的共线三链体与其双链DNA靶。GM 3形成的三链体阻断了重组和核NF-κB蛋白与GM-CSF元件的结合。GM 3还引起对人T细胞嗜淋巴细胞病毒-1 Tax反式激活因子诱导的荧光素酶活性的选择性抑制，所述荧光素酶活性来自由Jurkat T细胞中的GM-CSF启动子驱动的报告构建体。最后，GM 3大大降低了Jurkat T细胞中由不同刺激诱导的内源性GM-CSF mRNA的浓度，但不影响相同细胞中白细胞介素3 mRNA的水平。结论：GM-CSF启动子中的κB元件在内源性GM-CSF基因的转录激活中起着重要作用。共线三链体形成作为GM-CSF基因的选择性转录抑制因子，并且在该蛋白的不期望的过表达的情况下可能具有潜在的治疗应用。
Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a hemopoietic growth factor that is expressed in activated T cells, fibroblasts, macrophages, and endothelial cells. Although GM-CSF does not appear to be essential for normal hemopoiesis, overexpression of GM-CSF has been implicated in the pathogenesis of some diseases such as myeloid leukemia and chronic inflammation. An NF-κB/Rel binding site within the GM-CSF promoter, termed the κB element appears to be important for controlling expression in reporter gene assays in response to a number of stimuli in T cells. We investigated oligonucleotide-directed triple helix formation across this regulatory sequence as a potential tool to inhibit GM-CSF gene transcription. A 15-base oligonucleotide, GM3, was targeted to a purine-rich region in the GM-CSF proximal promoter, which overlaps the κB element. Gel mobility shift assays and DNase I footprinting demonstrated that GM3 formed a sequence-specific collinear triplex with its double-stranded DNA target. Triplex formation by GM3 blocked recombinant and nuclear NF-κB proteins binding to the GM-CSF element. GM3 also caused selective inhibition of the human T-cell lymphotrophic virus-1 Tax transactivator-induced luciferase activity from a reporter construct driven by the GM-CSF promoter in Jurkat T cells. Finally, GM3 greatly reduced the concentration of endogenous GM-CSF mRNA induced by different stimuli in Jurkat T cells but did not affect interleukin 3 mRNA levels in the same cells. We conclude that the κB element in the GM-CSF promoter plays a central role in the transcriptional activation of the endogenous GM-CSF gene. Colinear triplex formation acts as a selective transcriptional repressor of the GM-CSF gene and may have potential therapeutic application in cases of undesirable overexpression of this protein.
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影响因子：
14.9
作者：
Thekkumkat Thomas;C. Faaland;Michael A. Gallo;Thresla Thomas
通讯作者：
Thekkumkat Thomas;C. Faaland;Michael A. Gallo;Thresla Thomas
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影响因子：
14.9
作者：
Mayfield,C;Miller,D
通讯作者：
Miller,D
影响因子：
14.9
作者：
Frank M. Orson;Douglas W. Thomas;W. Michael McShan;D. Kessler;Michael E. Hogan
通讯作者：
Frank M. Orson;Douglas W. Thomas;W. Michael McShan;D. Kessler;Michael E. Hogan
急性髓细胞白血病中 GM-CSF 的自分泌。
影响因子：
20.3
作者：
Young,DC;Griffin,JD
通讯作者：
Griffin,JD
影响因子：
56.9
作者：
MOSER, HE;DERVAN, PB
通讯作者：
DERVAN, PB