Lipopolysaccharide induces 5-lipoxygenase-activating protein gene expression in THP-1 cells via a NF-κB and C/EBP-mediated mechanism
Lipopolysaccharide induces 5-lipoxygenase-activating protein gene expression in THP-1 cells via a NF-κB and C/EBP-mediated mechanism
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DOI：
10.1152/ajpcell.00296.2004
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发表时间：
2005-05-01
影响因子：
5.5
通讯作者：
Bigby, TD
中科院分区：
文献类型：
--
作者：
Serio, KJ;Reddy, KV;Bigby, TD
我们研究了5-脂氧合酶激活蛋白（FLAP）的诱导表达，这是在单核吞噬细胞的白三烯合成的关键。长时间暴露于细菌组分脂多糖（LPS），增加人单核细胞样THP-1细胞系中FLAP基因转录、mRNA表达和蛋白质表达。NF-κ B通路的激活和抑制可调节LPS诱导的FLAP基因表达。显性阴性I κ B α和p50/p65蛋白的过表达支持NF-κ B介导的作用机制。EMSA/supershift和DNase I足迹分析显示，p50结合到位于近端FLAP启动子的NF-κ B位点，而染色质免疫沉淀分析表明，LPS诱导p50结合，但不诱导p65结合。此外，EMSA/supershift分析表明，LPS诱导THP-1核提取物（含p50）的时间依赖性结合到这个启动子区域。NF-κ B位点的突变降低了基础启动子活性，并消除了p50和p65相关的诱导。EMSA/supershift分析也表明LPS诱导THP-1核提取物[含有CCAAT/增强子结合蛋白（C/EBP）-α、-δ和-β]与位于FLAP启动子中NF-κ B位点附近的C/EBP位点结合。我们的结论是LPS通过NF-κ B B和C/EBP介导的单核吞噬细胞转录机制增强FLAP基因表达。
We examined induced expression of the 5-lipoxygenase-activating protein (FLAP), which is critical for leukotriene synthesis in mononuclear phagocytes. Prolonged exposure to the bacterial component, lipopolysaccharide (LPS), increased FLAP gene transcription, mRNA expression, and protein expression in the human monocyte-like THP-1 cell line. Activation and inhibition of the NF-kappa B pathway modulated LPS induction of FLAP gene expression. An NF-kappa B-mediated mechanism of action was supported by overexpression of dominant-negative I kappa B alpha and p50/p65 proteins. EMSA/supershift and DNase I footprint analyses revealed that p50 binds to an NF-kappa B site located in the proximal FLAP promoter, while chromatin immunoprecipitation assays demonstrated that LPS induced binding of p50 but not of p65. Moreover, EMSA/supershift analyses demonstrated that LPS induced time-dependent binding of THP-1 nuclear extracts ( containing p50) to this promoter region. Mutation of the NF-kappa B site decreased basal promoter activity and abolished the p50- and p65-associated induction. EMSA/supershift analyses also demonstrated that LPS induced binding of THP-1 nuclear extracts [ containing CCAAT/enhancer binding protein (C/EBP)-alpha, -delta, and -epsilon] to a C/EBP site located adjacent to the NF-kappa B site in the FLAP promoter. We conclude that LPS enhances FLAP gene expression via both NF-kappa B- and C/EBP-mediated transcriptional mechanisms in mononuclear phagocytes.