Alternating 2′-O-ribose methylation is a universal approach for generating non-stimulatory siRNA by acting as TLR7 antagonist
Alternating 2′-O-ribose methylation is a universal approach for generating non-stimulatory siRNA by acting as TLR7 antagonist
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DOI：
10.1016/j.imbio.2009.09.003
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发表时间：
2010-07-01
期刊：
影响因子：
2.8
通讯作者：
Bauer, Stefan
中科院分区：
文献类型：
--
作者：
Hamm, Svetlana;Latz, Eicke;Bauer, Stefan
小干扰RNA（siRNA）被广泛用于调节基因表达，但其通过Toll样受体（TLR）诱导细胞因子的可能性严重损害了其使用。选择性的2 '-O-核糖正义或反义链甲基化可以消除免疫刺激潜力，然而，没有通用的方法可用，并且作用机制未知。在这里，我们证明了eGFP或β 2-微球蛋白特异性siRNA双链体内有义链的交替2 '-O-核糖甲基化破坏了其在原代免疫细胞中的免疫刺激功能，而靶基因表达的减少是功能性的。此外，将含有2 '-O-核糖甲基化正义链的siRNA添加到免疫刺激siRNA中消除了其刺激活性，并且结合研究显示2 '-O-核糖甲基化RNA比未修饰的RNA更强地结合TLR 7。我们的结论是，2 '-O-核糖甲基化作为抑制剂的RNA驱动的免疫刺激通过TLR 7，并建议交替2 '-O-核糖甲基化的正义链作为一种通用的方法，用于产生非免疫刺激siRNA。(C)2009年Elsevier GmbH。All rights reserved.
Small interfering RNA (siRNA) is widely used to modulate gene expression, but its potential induction of cytokines via Toll-like receptors (TLR) strongly impairs its use. Selective 2 '-O-ribose methylation of sense or antisense strand can abolish the immunostimulatory potential, however, no universal approach is available and the mechanism of action is unknown. Here, we demonstrate that alternating 2 '-O-ribose methylation of the sense strand within a siRNA duplex specific for eGFP or beta(2)-microglobulin destroyed its immunostimulatory function in primary immune cells, while reduction in target gene expression was functional. Furthermore, addition of siRNA containing a 2 '-O-ribose-methylated sense strand to immunostimulatory siRNA abolished its stimulatory activity and binding studies revealed that 2 '-O-ribose-methylated RNA bound stronger to TLR7 than unmodified RNA. We conclude that 2 '-O-ribose methylation acts as inhibitor for RNA-driven immune stimulation via TLR7 and recommend alternating 2 '-O-ribose methylation of the sense strand as a universal approach for the generation of non-immunostimulatory siRNA. (C) 2009 Elsevier GmbH. All rights reserved.