Role of HIV-1 Integrase in the Late Stage of Viral Replication

描述(由申请人提供)：HIV-1整合酶(IN)是病毒复制所必需的，因此是一个重要的治疗目标。在病毒复制的早期阶段，HIV-1 IN的一个四聚体催化逆转录病毒DNA整合到宿主基因组中。病毒DNA存在下IN的有序多聚化是IN催化的关键 活动。细胞辅因子LEDGF/p75与预先组装的IN-病毒DNA复合体结合，并将核蛋白复合体与活性基因捆绑在一起，从而确保有效整合。变构整合酶抑制剂(ALLINI)是一类新型的整合酶抑制剂，它结合在IN二聚体界面的LEDGF/p75结合部位，能够引发IN的异常多聚化。此外，在ALLINI存在的情况下产生的HIV-1病毒粒子显示病毒粒子核心的异常形态，并且对于随后的反转录和在靶细胞中的整合是有缺陷的。有趣的是，这些表型与某些IN突变体相似，被称为II类突变体。总的来说，对ALLINI和精选IN II类突变体的研究表明，HIV-1 IN在HIV-1复制的后期发挥关键作用。然而，潜在的机制尚不清楚。目前的应用将检验以下假设：有序IN多聚体对于其在病毒复制后期与病毒RNA的相互作用是重要的。因此，我们提出以下两个具体目标。目的1探讨HIV-1IN与RNA相互作用在病毒复制后期形成功能性核糖核蛋白复合体中的意义。AIM 2将表征影响IN多聚化和/或IN-LEDGF/p75结合的IN突变体。从这些研究中发现的机制和结构细节将为药物发现工作提供信息，以开发改进的ALLINI用于临床应用。

DESCRIPTION (provided by applicant): HIV-1 integrase (IN) is essential for viral replication and thus is an important therapeutic target. During the early stages of viral replication, a tetramer of HIV-1 IN catalyzes integration of reverse transcribed viral DNA into the host genome. The ordered multimerization of IN in the presence of viral DNA is critical for IN catalytic activity. Cellular cofactor LEDGF/p75 binds to the pre-assembled IN- viral DNA complex and tethers the nucleoprotein complex to active genes, thus ensuring effective integration. Allosteric integrase inhibitors (ALLINIs) are a novel class of integrase inhibitors that bind at the LEDGF/p75 binding site at the IN dimer interface and are capable of triggering aberrant IN multimerization. In addition, HIV-1 virions produced in the presence of ALLINIs display abnormal morphology of the virion cores and are defective for subsequent reverse transcription and integration in target cells. Interestingly, these phenotypes are similar to certain IN mutants, which have been termed class II mutants. Collectively, the studies with ALLINIs and select IN class II mutants suggest that HIV-1 IN plays a key role during late stage HIV-1 replication. However, the underlying mechanism is not clear. The present application will test the following hypothesis: ordered IN multimerization is important for its interaction with viral RNA during the late stage viral replication. Therefore, we propose the following two specific aims. Aim 1 will explore the significance of HIV-1 IN interactions with RNA for formation of the functional ribonuleoprotein complexes during the late stage viral replication. Aim 2 will characterize IN mutants that affect IN multimerization and/or IN- LEDGF/p75 binding. Mechanistic and structural details that will emerge from these studies will inform drug discovery efforts to develop improved ALLINIs for their clinical application.