Modulation of Retroviral Uncoating by Cellular Factors

 描述（由申请人提供）：在上一个资助期内，我们在我们对HIV-1脱壳和逆转录的理解方面取得了重大进展，发表了36篇手稿，其中19篇是最后一位作者。我们的突出发现是，细胞蛋白质和药物使HIV-1核心不稳定，从而加速脱壳，阻止逆转录的发生并破坏感染。总的来说，我们的研究结果使我们提出了一个模型，其中HIV-1核心是一个“容器”，确保在脱壳之前完成逆转录。TRIM 5 α或小分子抑制剂PF 74和Bi 2等药物可使HIV-1核心不稳定，从而阻止逆转录的发生。然而，稳定HIV-1核心的蛋白质和药物也会破坏感染。为了了解HIV-1核心的稳定性阻断HIV-1感染的机制，我们将利用最近发现的人类MxB蛋白。MxB是一种干扰素-α-诱导蛋白，在逆转录后但整合前阻断HIV-1感染。遗传实验表明，衣壳是MxB阻断HIV-1感染能力的病毒决定因素;与此一致，我们的初步结果表明，MxB与HIV-1核心结合，并在感染期间稳定HIV-1核心。MxB是已知的第一种天然表达的稳定HIV-1核心的蛋白质，使MxB成为了解核心稳定如何导致HIV-1感染阻断的理想候选者。本申请将测试MxB与HIV-1核心结合并在其表面形成更高级结构，从而防止脱壳的假设。以下目的将用于检验该假设：1）检查Mx B结合衣壳对限制的作用，2）检查寡聚化和高阶自缔合在Mx B结合HIV-1核心能力中的作用，3）检查限制B Mx B的亚细胞定位，和4）检查磷酸化在Mx B阻断HIV-1感染能力中的作用。通过这项提议获得的知识将有助于理解HIV-1的基本脱壳过程和MxB阻断感染的机制。这里获得的知识是开发针对HIV-1的新疗法的潜在基础。

 DESCRIPTION (provided by applicant): During the previous funding period we made major advances in our understanding of HIV-1 uncoating and reverse transcription, publishing 36 manuscripts, including 19 with senior last authorship. Our salient finding was that cellular proteins and drugs that destabilize the HIV-1 core, thereby accelerating uncoating, prevent the occurrence of reverse transcription and disrupt infection. Overall our findings lead us to propose a model in which the HIV-1 core is a "container" that ensures the completion of reverse transcription before uncoating. Agents such as TRIM5α or the small molecule inhibitors PF74 and Bi2, which destabilize the HIV-1 core, prevent the occurrence of reverse transcription. Proteins and drugs that stabilize the HIV-1 core, however, also disrupt infection. To understand the mechanism by which stabilization of the HIV-1 core blocks HIV-1 infection, we are going to take advantage of the recently discovered human MxB protein. MxB is an interferon-α-inducible protein that blocks HIV-1 infection after reverse transcription but prior to integration. Genetic experiments suggested that capsid is the viral determinant for the ability of MxB to block HIV-1 infection; In agreement, our preliminary results indicated that MxB binds to the HIV-1 core and stabilizes the HIV-1 core during infection. MxB is the first naturally expressed protein known to stabilize the HIV-1 core, making MxB an ideal candidate to understand how core stabilization leads to a block on HIV-1 infection. This application will test the hypothesis that MxB binds to the HIV-1 core and forms higher order structures on its surface, thereby preventing uncoating. The following aims will be used to test this hypothesis: 1) Examine the role of MxB binding to capsid on restriction, 2) Examine the role of oligomerization and higher-order self-association in the ability of MxB to bind the HIV-1 core, 3) Examine the subcellular localization of restriction b MxB, and 4) Examine the role of phosphorylation in the ability of MxB to block HIV-1 infection. The knowledge gain by this proposal will be instrumental for understanding the basic uncoating process of HIV-1 and the mechanism by which MxB blocks infection. The knowledge gain here is the potential basis for the development of novel treatments against HIV-1.