Human H1 promoter expressed short hairpin RNAs (shRNAs) suppress avian influenza virus replication in chicken CH-SAH and canine MDCK cells
Human H1 promoter expressed short hairpin RNAs (shRNAs) suppress avian influenza virus replication in chicken CH-SAH and canine MDCK cells
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DOI：
10.1016/j.antiviral.2009.08.009
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发表时间：
2009-11-01
影响因子：
7.6
通讯作者：
Golovan, Serguei P.
中科院分区：
文献类型：
--
作者：
Abrahamyan, Arusyak;Nagy, Eva;Golovan, Serguei P.
甲型流感病毒对世界各地的人类和动物都构成了重大的健康和经济威胁。利用聚合酶III人H1启动子表达针对禽流感病毒核衣壳蛋白(NP)或聚合酶复合体(PA)酸性成分的9个短发夹状RNA(ShRNAs)。被测试的shRNAs使鸡CH-SAH细胞中的感染性滴度下降了400倍，病毒NP和PA的mRNAs水平下降了80%(p<；0.05)。在哺乳动物的MDCK细胞中，检测到感染滴度下降了10(6)倍，病毒mRNAs水平下降了90%(p<；0.05)。虽然RNAi沉默在病毒抑制中发挥了主要作用，但任何一种质粒的转染都能在MDCK细胞中诱导抗病毒干扰素反应。在CH-SAH细胞中，构建体未诱导干扰素应答，流感病毒感染可抑制干扰素应答。同时在CH-SAH细胞中诱导RNAi和干扰素反应导致感染滴度下降18,500倍(p<；0.05)。我们还在NP基因中发现了新的高效和保守的RNAi靶点，可用于shRNAs的抗病毒鸡尾酒，以阻止流感病毒的逃逸。(C)2009爱思唯尔B.V.保留所有权利。
Influenza A virus represents a significant health and economic threat to both humans and animals worldwide. The polymerase III human H1 promoter was used to express nine short hairpin RNAs (shRNAs) targeting nucleocapsid protein (NP) or the acidic component of polymerase complex (PA) genes of avian influenza virus. Tested shRNAs led to a decrease of up to 400 fold in infective titre and up to 80% in the level of viral NP and PA mRNAs in chicken CH-SAH cells (p < 0.05). In mammalian MDCK cells a decrease of up to 10(6)-fold in infective titre, and up to 90% in the level of viral mRNAs (p < 0.05) was detected. While RNAi silencing played a major role in viral inhibition, transfection of any plasmid induced an antiviral interferon response in MDCK cells. In CH-SAH cells, no induction of IFN response was observed by constructs, and the IFN response was suppressed by influenza virus infection. Simultaneous induction of RNAi and IFN responses in CH-SAH cells resulted in 18,500-fold decrease in infective titre (p < 0.05). We also identified novel efficient and conserved RNAi target site in the NP gene which can be used in antiviral cocktails of shRNAs to block the escape of influenza virus. (C) 2009 Elsevier B.V. All rights reserved.