A Doubly Fluorescent HIV-1 Reporter Shows that the Majority of Integrated HIV-1 Is Latent Shortly after Infection
A Doubly Fluorescent HIV-1 Reporter Shows that the Majority of Integrated HIV-1 Is Latent Shortly after Infection
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DOI：
10.1128/jvi.03478-12
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发表时间：
2013-04-01
影响因子：
5.4
通讯作者：
Sadowski, Ivan
中科院分区：
文献类型：
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作者：
Dahabieh, Matthew S.;Ooms, Marcel;Sadowski, Ivan
HIV-1潜伏期是根除病毒的主要障碍。典型地，潜伏期被认为是由活动性感染的渐进性表观遗传沉默引起的。然而，由于目前的大多数潜伏期模型不能区分初始潜伏期(LTR-静止期)和继发性(渐进性沉默)潜伏期，因此对末端重复(LTR)沉默感染的发生时间和持续时间知之甚少。在这项研究中，我们构建并鉴定了一种新型的双标记HIV-1载体(红-绿-HIV-1[RGH])，它允许独立于LTR活性检测感染细胞。Jurkat T细胞和其他细胞系感染RGH表明，大多数整合前病毒在感染后早期是LTR沉默的。此外，由于前病毒可以被各种T细胞信号激动剂重新激活，因此LTR沉默的感染在转录水平上是有效的。此外，我们使用双标记载体系统比较了来自七个不同亚型的LTRs在LTR沉默和重新激活方面的作用。这些实验表明D亚型和F ltrs对沉默更敏感，而AELtr亚型对沉默基本不敏感。最后，感染激活的人类原代CD4(+)T细胞会产生LTR沉默感染和生产性感染。综上所述，我们使用新开发的RGH载体作为在单细胞水平上分析HIV-1潜伏期的敏感工具产生的数据表明，大多数HIV-1感染是潜在的早期感染。
HIV-1 latency poses a major barrier to viral eradication. Canonically, latency is thought to arise from progressive epigenetic silencing of active infections. However, little is known about when and how long terminal repeat (LTR)-silent infections arise since the majority of the current latency models cannot differentiate between initial (LTR-silent) and secondary (progressive silencing) latency. In this study, we constructed and characterized a novel, double-labeled HIV-1 vector (Red-Green-HIV-1 [RGH]) that allows for detection of infected cells independently of LTR activity. Infection of Jurkat T cells and other cell lines with RGH suggests that the majority of integrated proviruses were LTR-silent early postinfection. Furthermore, the LTR-silent infections were transcriptionally competent, as the proviruses could be reactivated by a variety of T cell signaling agonists. Moreover, we used the double-labeled vector system to compare LTRs from seven different subtypes with respect to LTR silencing and reactivation. These experiments indicated that subtype D and F LTRs were more sensitive to silencing, whereas the subtype AE LTR was largely insensitive. Lastly, infection of activated human primary CD4(+) T cells yielded LTR-silent as well as productive infections. Taken together, our data, generated using the newly developed RGH vector as a sensitive tool to analyze HIV-1 latency on a single-cell level, show that the majority of HIV-1 infections are latent early postinfection.