Nuclear export of the HIV genome: A Molecular Dynamics Study of the Interactions

HIV基因组核输出：输出过程中HIV Rev和宿主CRM1相互作用和结构的分子动力学研究摘要HIV病毒是一种大流行慢病毒，目前尚无治愈方法或疫苗。目前的治疗方法包括强大的药物鸡尾酒，包含针对病毒生命周期的多个方面的多种药物。除了许多这些药物引起的毒副作用外，在大多数情况下，病毒对所使用的药物产生耐药性，迫使改用毒性更大、效果更差的药物，最终导致病毒逃脱药物控制。尽管可用的药物种类繁多，但目前没有药物靶向病毒生命周期的关键部分，Rev和出口蛋白-1 （Crm1）在活跃的病毒复制过程中介导了整个病毒基因组的输出。在这里，病毒蛋白Rev在病毒基因组的Rev响应元件（RRE）上形成一个六部分的低聚物。然后，这种低聚物招募宿主蛋白crm1的拷贝，绕过mRNA剪接和输出的标准途径，将基因组输出到细胞核外。由于对所涉及的蛋白质的结构和机制理解不佳，针对这一输出途径的尝试受到限制。虽然实验方法已经成功地阐明了该过程的广泛细节，所涉及的蛋白质和RNA的结合区域，甚至一些结构和机制的细节，但迄今为止得出的理解是不完整和不确定的，不足以用于真正深入的研究或支持修改该途径的尝试。利用计算和分子动力学方法的精细时空分辨率，可以克服这些问题。在本项目中，这些方法将用于研究出口复合物形成的整个过程，从第一个Rev单体的附着到RRE一直到多个crm1副本的招募，并将使用它们来解决该领域的突出问题，并为进一步的研究开辟道路。具体而言，本研究将验证先前提出的机制和结构，深入研究Rev寡聚物- rre组装的机制，包括协同性的重要性，并提供该组装的完整结构。此外，还将研究该装配体与crm1的结合，其结果将用于解决诸如结合crm1蛋白的最大可能数量以及所有六个Rev拷贝在结合中的重要性等问题。最后，也是最重要的是，将生成整个出口综合体的结构，可用于未来的机械和计算研究，或作为未来实验研究的路标。从长远来看，甚至有可能将这些结果付诸实践。

DESCRIPTION (provided by applicant): Nuclear export of the HIV genome: A Molecular Dynamics Study of the Interactions and Structure of HIV Rev and Host CRM1 during Export Summary The HIV virus is a pandemic lentivirus which has no existing cure or vaccine. Current treatments consist of powerful drug cocktails containing multiple drugs targeting multiple aspects of the viral lifecycle. Besides the toxic side effects caused by many of these drugs, in the majority of cases the virus develops resistance to the drugs used, forcing a switch to more toxic and less effective drugs, culminating in the escape of the virus from drug control. Despite the variety of drugs available, no current drugs target a critical part of the viral lifecycle, the Rev and Exportin-1 (Crm1) mediated export of the full viral genome during active viral replication. Here, the viral protein Rev forms a six-part oligomer on the Rev Response Element (RRE) of the viral genome. This oligomer then recruits copies of the host protein crm1 to export the genome out of the cell nucleus, bypassing the standard pathways of mRNA splicing and export. Attempts to target this export pathway are limited by a combination of poor structural and mechanistic understandings of the proteins involved. While experimental methods have been successful in elucidating broad details of the process, the binding regions of the proteins and RNA involved, and even some details of structure and mechanism, the understanding derived so far is incomplete and uncertain, inadequate for use in truly in-depth studies or to support attempts to modify the pathway. Utilizing the fine spatiotemporal resolution of computational and molecular dynamics approaches, these problems can be overcome. In this project, these methods will be used to investigate the entire process of export complex formation, from the attachment of the first Rev monomer to the RRE all the way up the recruitment of multiple crm1 copies, and will use them to address outstanding questions in the field and to open the way for further investigation. Specifically, this study will validate previous proposed mechanisms and structures, deeply investigate the mechanism of Rev oligomer-RRE assembly, including the importance of cooperativity, and provide a complete structure of this assembly. In addition, binding of the assembly with crm1 will be investigated, and the results used to address such topics as the maximal plausible number of bound crm1 proteins and the importance of all six Rev copies in binding. Finally, and most importantly, a structure of the entire export complex will be generated, which can be used in future mechanistic and computational studies, or as a guidepost for future experimental studies. In the long-run it may even be possible to put these results to practical use. PUBLIC HEALTH RELEVANCE: Nuclear export of the HIV genome: A Molecular Dynamics Study of the Interactions and Structure of HIV Rev and Host CRM1 during Export Narrative Despite extensive research, the process by which HIV Rev oligomerizes on the Rev Response Element (RRE) sequence of the viral genome and recruits crm1 to export this genome is not well-characterized. Many of the obstacles to experimental studies that frustrate attempts to fully understand the process fall away when computational methods are considered instead. The greater understanding of the Rev export pathway that will be obtained when such methods are used will prove invaluable to the field of study, and may provide intriguing avenues of exploration into one of the few steps of HIV replication which is not yet the target of therapy.