Design of extended short hairpin RNAs for HIV-1 inhibition.
Design of extended short hairpin RNAs for HIV-1 inhibition.
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DOI：
10.1093/nar/gkm596
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发表时间：
2007
影响因子：
14.9
通讯作者：
Berkhout B
中科院分区：
文献类型：
--
作者：
Liu YP;Haasnoot J;Berkhout B
针对病毒mRNAs的RNA干扰(RNAi)被广泛用于阻断病毒在哺乳动物细胞中的复制。特异性的抗病毒RNAi反应可以通过转染合成的小干扰RNA(SiRNAs)或通过细胞内表达短发夹状RNAs(ShRNAs)来诱导。对于HIV-1，这两种方法都极大地抑制了病毒复制。然而，由于RNAi耐药逃逸病毒的迅速出现，单一siRNA/shRNA的治疗应用似乎有限。这些变异体在靶序列中包含缺失或点突变，从而消除了抗病毒作用。为了避免逃避RNAi，病毒应该同时以多个shRNA为靶点。或者，可以使用长发夹状RNA，从其可以产生多个有效的siRNA。在本研究中，我们构建了扩展的shRNAs(e-shRNAs)，它编码两个有效的针对HIV-1保守序列的siRNAs。活性分析和RNA加工分析表明，两个siRNAs在发夹茎中的定位对于两个功能siRNAs的产生至关重要。高效地加工成两个有效的siRNAs的e-shRNAs比加工较差的e-shRNAs显示出更好的抑制病毒产生的作用，而不会诱导干扰素反应。这些结果为设计多个siRNA发夹结构提供了构建原则。
RNA interference (RNAi) targeted towards viral mRNAs is widely used to block virus replication in mammalian cells. The specific antiviral RNAi response can be induced via transfection of synthetic small interfering RNAs (siRNAs) or via intracellular expression of short hairpin RNAs (shRNAs). For HIV-1, both approaches resulted in profound inhibition of virus replication. However, the therapeutic use of a single siRNA/shRNA appears limited due to the rapid emergence of RNAi-resistant escape viruses. These variants contain deletions or point mutations within the target sequence that abolish the antiviral effect. To avoid escape from RNAi, the virus should be simultaneously targeted with multiple shRNAs. Alternatively, long hairpin RNAs can be used from which multiple effective siRNAs may be produced. In this study, we constructed extended shRNAs (e-shRNAs) that encode two effective siRNAs against conserved HIV-1 sequences. Activity assays and RNA processing analyses indicate that the positioning of the two siRNAs within the hairpin stem is critical for the generation of two functional siRNAs. E-shRNAs that are efficiently processed into two effective siRNAs showed better inhibition of virus production than the poorly processed e-shRNAs, without inducing the interferon response. These results provide building principles for the design of multi-siRNA hairpin constructs.
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DOI：
10.1073/pnas.232688199
发表时间：
2003-01-07
影响因子：
11.1
作者：
Qin, XF;An, DS;Baltimore, D
通讯作者：
Baltimore, D
影响因子：
3.7
作者：
Verma, S;Ziegler, K;Nerurkar, VR
通讯作者：
Nerurkar, VR
影响因子：
12.4
作者：
ter Brake, Olivier;Konstantinova, Pavlina;Berkhout, Ben
通讯作者：
Berkhout, Ben
影响因子：
64.8
作者：
Grimm, Dirk;Streetz, Konrad L.;Kay, Mark A.
通讯作者：
Kay, Mark A.
影响因子：
5.4
作者：
Coburn, GA;Cullen, BR
通讯作者：
Cullen, BR