New insights into the interplay between HIV and the autophagy machinery

项目总结 自噬是一种重要的细胞抗病毒防御机制，可能影响艾滋病毒感染和 变速箱。自噬的抗病毒特性有两个方面：(I)自噬以溶酶体的病毒粒子为靶标 (2)该途径有助于病毒病原体的抗原提呈。尽管它有抗病毒的潜力， 利用自噬来对抗艾滋病毒的努力一直受到限制，因为我们对自噬的作用的理解存在差距 HIV感染中的自噬。我们的长期目标是操纵自噬的抗病毒特性，以促进 清除艾滋病毒并改进治疗。我们的总体目标是确定分子间的相互作用 艾滋病毒和自噬机制。我们的中心假设是自噬限制了艾滋病毒，但病毒 使用Nef克服了这一障碍，Nef是一种臭名昭著的毒性因子，可以调节免疫逃避。这一假设已经 是基于我们发表的和新的工作证明自噬会耗尽HIV蛋白 Gag、Vif和Vpr，但仅当nef有缺陷时。17.Gag是病毒粒子组装的驱动程序；Vif抵消病毒粒子组装 限制因子APOBEC3G的破坏作用；VPR通过促进病毒增强传染性 转录并阻断细胞周期于G2期。因此，Gag、Vif和Vpr的减少将影响病毒粒子的产生 和传染性。然而，我们发现Nef阻止了自噬，反过来又恢复了HIV蛋白质和颗粒的释放。 我们的研究表明，Nef将自噬启动子BECN1隔离在内质网(ER) 通过促进其与自噬抑制因子BCL2的结合。从机制上讲，我们发现了Nef诱导 BCL2通过E3连接酶Parkin实现单一泛素化。这一翻译后的修饰增加了《基本法》第二章的S 对BECN1的抑制作用。因此，Nef的这种活性损害了自噬小体的形成。它的意义在于 这项研究是：(I)它将解决关于艾滋病毒和艾滋病之间相互作用的一个重要的知识差距。 自噬，并将揭示这一途径如何影响艾滋病毒感染和传播；以及(Ii)它将提供 设计旨在加强自噬治疗艾滋病毒感染和/或改善治愈的方法的基础 战略。我们将通过以下三个具体目标来实现总体目标：(1)识别艾滋病毒 易受自噬的分子；(2)剖析Nef阻止自噬的机制；以及(3)阐明 自噬拮抗对HIV感染性的影响。这项拟议的研究具有创新性，因为它 通过揭示HIV严重需要Nef来抵消，代表了对现状的实质性偏离 自噬限制。

PROJECT SUMMARY Autophagy is an important cellular antiviral defense mechanism that potentially affects HIV infection and transmission. The antiviral properties of autophagy are two-fold: (i) autophagy targets virions for lysosomal degradation and (ii) this pathway aids in antigen presentation of virus pathogens. Despite its antiviral potential, efforts to capitalize on autophagy against HIV have been limited due to a gap in our understanding of the role of autophagy in HIV infection. Our long-term goal is to manipulate the antiviral properties of autophagy to facilitate HIV clearance and improve treatments. Our overall objective is to identify the molecular interactions between HIV and the autophagy machinery. Our central hypothesis is that autophagy restricts HIV, but the virus overcomes this block using Nef, a notorious virulence factor that mediates immune evasion. This hypothesis has been formulated based on our published and novel work demonstrating that autophagy depletes the HIV proteins Gag, Vif and Vpr, but only when nef is defective 17. Gag is the driver of virion assembly; Vif counteracts the damaging effects of the restriction factor APOBEC3G; and Vpr enhances infectivity by facilitating viral transcription and arresting cell cycle at G2. Thus, a decrease in Gag, Vif and Vpr will impact virion production and infectivity. However, we found that Nef blocks autophagy, restoring in turn HIV proteins and particle release. Our studies showed that Nef sequesters the autophagosome initiator BECN1 at the endoplasmic reticulum (ER) by facilitating its association with the autophagy inhibitor BCL2. Mechanistically, we uncovered that Nef induces BCL2 mono-ubiquitination via the E3 ligase Parkin. This post-translational modification increases BCL2's inhibitory effect over BECN1. Hence, this activity of Nef impairs autophagosome formation. The significance for this research is that: (i) it will address an important knowledge gap concerning the interplay between HIV and autophagy, and will reveal how this pathway influences HIV infection and transmission; and (ii) it will provide the basis for the design of approaches aimed at enhancing autophagy to treat HIV infection and/or improve cure strategies. We will attain the overall objective by pursuing the following three specific aims: (1) Identify HIV molecules vulnerable to autophagy; (2) Dissect the mechanism by which Nef blocks autophagy; and (3) Elucidate the impact of autophagy antagonism on HIV infectivity. The proposed research is innovative because it represents a substantive departure from the status quo by revealing that HIV critically needs Nef to counteract autophagy restriction.