Targets of small interfering RNA restriction during human immunodeficiency virus type 1 replication
Targets of small interfering RNA restriction during human immunodeficiency virus type 1 replication
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DOI：
10.1128/jvi.02126-07
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发表时间：
2008-03-01
影响因子：
5.4
通讯作者：
Arts, Eric J.
中科院分区：
文献类型：
--
作者：
Gao, Yong;Lobritz, Michael A.;Arts, Eric J.
小干扰RNA(SiRNAs)在体外能有效抑制人类免疫缺陷病毒1型(HIV-1)的复制。这种抑制的机制(S)尚不清楚，因为在逆转录病毒生命周期的不同阶段，siRNA可能与多个艾滋病毒-1RNA物种相互作用。为了确定敏感的HIV-1RNA物种，siRNAs首先被设计成通过env和gag基因靶点特异性地抑制两个不同的HIV-1初级分离株。携带这些靶序列的自失活慢病毒载体证实，siRNA不能降解传入的基因组RNA。然而，猕猴TRIM5α对进入的核心结构的破坏确实提供了siRNA-RNA诱导的沉默复合体访问HIV-1基因组RNA并促进了降解。在没有加速核心破坏的情况下，只有细胞质中新转录的HIV-1mRNA对Arna降解敏感。HIV-1mRNA核输出的抑制剂，如软霉素B和喜树碱，可阻断siRNA限制。通过3‘-5’-siRNA的单向扩增和扩散，所有HIV-1RNA区域和转录本都被降解，包括多个剪接的HIV-1RNA。相反，靶序列的HIV-1RNA3‘对Arna不敏感。即使在Arna存在的情况下，全长的HIV-1RNA仍然被封装到新组装的病毒中。这些发现表明，尽管病毒RNA几乎参与了逆转录病毒生命周期的每一步，但ARNA只能针对相对“裸露的”细胞质HIV-1RNA。在病毒进入后、在包膜/病毒组装期间或在核内对HIV-1RNA的保护可能反映了病毒对siRNA、TRIM5α或其他宿主限制因素的反应。
Small interfering RNAs (siRNAs) have been shown to effectively inhibit human immunodeficiency virus type 1 (HIV-1) replication in vitro. The mechanism(s) for this inhibition is poorly understood, as siRNAs may interact with multiple HIV-1 RNA species during different steps of the retroviral life cycle. To define susceptible HIV-1 RNA species, siRNAs were first designed to specifically inhibit two divergent primary HIV-1 isolates via env and gag gene targets. A self-inactivating lentiviral vector harboring these target sequences confirmed that siRNA cannot degrade incoming genomic RNA. Disruption of the incoming core structure by rhesus macaque TRIM5 alpha did, however, provide siRNA-RNA-induced silencing complex access to HIV-1 genomic RNA and promoted degradation. In the absence of accelerated core disruption, only newly transcribed HIV-1 mRNA in the cytoplasm is sensitive to ARNA degradation. Inhibitors of HIV-1 mRNA nuclear export, such as leptomycin B and camptothecin, blocked siRNA restriction. All HIV-1 RNA regions and transcripts found 5' of the target sequence, including multiply spliced HIV-1 RNA, were degraded by unidirectional 3'-to-5' siRNA amplification and spreading. In contrast, HIV-1 RNA 3' of the target sequence was not susceptible to ARNA. Even in the presence of ARNA, full-length HIV-1 RNA is still encapsidated into newly assembled viruses. These findings suggest that ARNA can target only a relatively "naked" cytoplasmic HIV-1 RNA despite the involvement of viral RNA at nearly every step in the retroviral life cycle. Protection of HIV-1 RNA within the core following virus entry, during encapsidation/virus assembly, or within the nucleus may reflect virus evolution in response to siRNA, TRIM5 alpha, or other host restriction factors.