Genome editing of the HIV co-receptors CCR5 and CXCR4 by CRISPR-Cas9 protects CD4(+) T cells from HIV-1 infection.
Genome editing of the HIV co-receptors CCR5 and CXCR4 by CRISPR-Cas9 protects CD4(+) T cells from HIV-1 infection.
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通过 CRISPR-Cas9 对 HIV 辅助受体 CCR5 和 CXCR4 进行基因组编辑可保护 CD4 T 细胞免受 HIV-1 感染
DOI：
10.1186/s13578-017-0174-2
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发表时间：
2017
影响因子：
7.5
通讯作者：
Guo D
中科院分区：
文献类型：
--
作者：
Liu Z;Chen S;Jin X;Wang Q;Yang K;Li C;Xiao Q;Hou P;Liu S;Wu S;Hou W;Xiong Y;Kong C;Zhao X;Wu L;Li C;Sun G;Guo D
治疗HIV-1感染的主要方法是联合抗逆转录病毒治疗（cART）。尽管cART在降低HIV-1病毒载量和控制疾病进展方面是有效的，但它具有许多副作用，并且对于必须保持终身治疗的HIV-1感染患者来说是昂贵的。随着基因组编辑工具研究的进展，HIV-1基因治疗引起了人们的广泛关注。例如，锌指核酸酶（ZFN）、转录激活因子样效应物核酸酶（TALEN）和成簇的规则间隔短回文重复序列（CRISPR）-Cas9已被用于成功地破坏HIV-I共受体CCR 5或CXCR 4，从而限制HIV-I感染。然而，通过CRISPR-Cas9对CXCR 4和CCR 5进行同步基因组编辑在阻断原代CD 4 + T细胞中的HIV-1感染方面的作用很少报道。此外，CXCR 4和CCR 5的不同靶位点的组合也需要研究。在这份报告中，我们设计了两种不同的gRNA组合，靶向CXCR 4和CCR 5，在一个单一的载体。CRISPR-sgRNA-Cas9可以在各种细胞系和原代CD 4 + T细胞中成功诱导CXCR 4和CCR 5基因的编辑。使用HIV-1攻击试验，我们证明了CXCR 4-和CCR 5-修饰的细胞中CXCR 4-嗜性或CCR 5-嗜性HIV-1感染显著减少，并且修饰的细胞在HIV-1感染期间表现出比未修饰细胞的选择性优势。脱靶分析表明，在所有预测位点中均未发现非特异性编辑。此外，细胞凋亡测定表明，CRISPR-Cas9同时破坏原代CD 4 + T细胞中的CXCR 4和CCR 5对细胞活力没有明显的细胞毒性作用。我们的研究结果表明，通过CRISPR-Cas9对CXCR 4和CCR 5进行同步基因组编辑可能为HIV-1感染的功能性治疗提供有效和安全的策略。本文的在线版本（doi：10.1186/s13578-017-0174-2）包含补充材料，可供授权用户使用。
The main approach to treat HIV-1 infection is combination antiretroviral therapy (cART). Although cART is effective in reducing HIV-1 viral load and controlling disease progression, it has many side effects, and is expensive for HIV-1 infected patients who must remain on lifetime treatment. HIV-1 gene therapy has drawn much attention as studies of genome editing tools have progressed. For example, zinc finger nucleases (ZFN), transcription activator like effector nucleases (TALEN) and clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 have been utilized to successfully disrupt the HIV-1 co-receptors CCR5 or CXCR4, thereby restricting HIV-1 infection. However, the effects of simultaneous genome editing of CXCR4 and CCR5 by CRISPR-Cas9 in blocking HIV-1 infection in primary CD4+ T cells has been rarely reported. Furthermore, combination of different target sites of CXCR4 and CCR5 for disruption also need investigation. In this report, we designed two different gRNA combinations targeting both CXCR4 and CCR5, in a single vector. The CRISPR-sgRNAs-Cas9 could successfully induce editing of CXCR4 and CCR5 genes in various cell lines and primary CD4+ T cells. Using HIV-1 challenge assays, we demonstrated that CXCR4-tropic or CCR5-tropic HIV-1 infections were significantly reduced in CXCR4- and CCR5-modified cells, and the modified cells exhibited a selective advantage over unmodified cells during HIV-1 infection. The off-target analysis showed that no non-specific editing was identified in all predicted sites. In addition, apoptosis assays indicated that simultaneous disruption of CXCR4 and CCR5 in primary CD4+ T cells by CRISPR-Cas9 had no obvious cytotoxic effects on cell viability. Our results suggest that simultaneous genome editing of CXCR4 and CCR5 by CRISPR-Cas9 can potentially provide an effective and safe strategy towards a functional cure for HIV-1 infection. The online version of this article (doi:10.1186/s13578-017-0174-2) contains supplementary material, which is available to authorized users.
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DOI：
10.3390/v7082816
发表时间：
2015-07-27
期刊：
Viruses
影响因子：
--
作者：
Hütter G;Bodor J;Ledger S;Boyd M;Millington M;Tsie M;Symonds G
通讯作者：
Symonds G
影响因子：
64.5
作者：
Hsu PD;Lander ES;Zhang F
通讯作者：
Zhang F
影响因子：
56.9
作者：
Jinek, Martin;Chylinski, Krzysztof;Charpentier, Emmanuelle
通讯作者：
Charpentier, Emmanuelle
影响因子：
16.8
作者：
Durand CM;Blankson JN;Siliciano RF
通讯作者：
Siliciano RF
影响因子：
4.6
作者：
Hou P;Chen S;Wang S;Yu X;Chen Y;Jiang M;Zhuang K;Ho W;Hou W;Huang J;Guo D
通讯作者：
Guo D