Inhibition of tumor necrosis factor-α through selective blockade of Pre-mRNA splicing by shikonin
Inhibition of tumor necrosis factor-α through selective blockade of Pre-mRNA splicing by shikonin
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DOI：
10.1124/mol.106.032821
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发表时间：
2007-06-01
影响因子：
3.6
通讯作者：
Yang, Ning-Sun
中科院分区：
文献类型：
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作者：
Chiu, Shao-Chih;Yang, Ning-Sun
我们之前开发了一种基于基因枪的体内筛选系统，并鉴定出紫草素是肿瘤坏死因子-α（TNF-alpha）基因表达的有效抑制剂。在这里，我们表明，紫草素选择性抑制TNF-α的表达在RNA剪接水平。用紫草素处理脂多糖刺激的人原代单核细胞和THP-1细胞导致TNF-α的正常转录诱导，但未剪接的前mRNA以功能mRNA为代价积累。这种效应发生与非细胞毒性剂量的紫草素，是高度特异性的，因为既不是管家基因，也不是另一种炎症细胞因子基因，白细胞介素-8（IL-8）的mRNA的生产，受到影响。此外，与脂多糖（LPS）和紫草素的共处理增加了IL-8的终点蛋白的产生，伴随着抑制双链RNA激活的蛋白激酶（PKR）途径的激活。由于PKR失活已被证明下调TNF-α RNA的剪接过程并干扰翻译，我们的研究结果表明，紫草素可能通过PKR通路的失活实现细胞因子蛋白表达的差异调节，并揭示TNF-α前体mRNA剪接的调节可能构成未来抗炎应用的一个有前途的靶点。
We previously developed a gene-gun-based in vivo screening system and identified shikonin as a potent suppressor of tumor necrosis factor-alpha(TNF-alpha) gene expression. Here, we show that shikonin selectively inhibits the expression of TNF-alpha at the RNA splicing level. Treatment of lipopolysaccharide-stimulated human primary monocytes and THP-1 cells with shikonin resulted in normal transcriptional induction of TNF-alpha, but unspliced pre-mRNA accumulated at the expense of functional mRNA. This effect occurred with noncytotoxic doses of shikonin and was highly specific, because mRNA production of neither a housekeeping gene nor another inflammatory cytokine gene, interleukin-8 (IL-8), was affected. Moreover, cotreatment with lipopolysaccharide (LPS) and shikonin increased the endpoint protein production of IL-8, accompanied by suppressed activation of the double-stranded RNA-activated protein kinase (PKR) pathway. Because PKR inactivation has been shown to down-regulate the splicing process of TNF-alpha RNA and interfere with translation, our findings suggest that shikonin may achieve differential modulation of cytokine protein expression through inactivation of the PKR pathway and reveal that regulation of TNF-alpha pre-mRNA splicing may constitute a promising target for future anti-inflammatory application.