宿主蛋白PQBP1通过结合HIV-1复制复合体抑制病毒感染巨噬细胞的机理研究

Macrophages and T cells are major target cells of HIV-1 infection. Compared to T cells, macrophages can maintain long-term HIV-1 replication. Infected macrophages can also mediate highly efficient virus dissemination through cell-to-cell transmission and therefore play important roles in virus replication, dissemination and pathogenesis. HIV-1 infection of macrophages is largely determined by the virus-host interactions. Our preliminary results indicated that host factor PQBP1 (polyglutamine binding protein 1) binds to HIV-1 replication complexes in infected macrophages and inhibits virus infection; however the molecular mechanism of inhibition is unclear. Here, we propose 1) to further investigate the interaction between PQBP1 and HIV-1 replication complexes in infected macrophages with our recently developed visualization strategy; 2) to identify the viral component(s) that interact(s) with PQBP1 and their interaction interface; 3) to identify the HIV-1 replication event that is inhibited by PQBP1 through quantitative analysis of replication products during infection and the stability of viral replication complexes. Taken these results together, we will be able to elucidate the molecular mechanism of PQBP1 restriction of HIV-1 replication in macrophages. This work will not only shed light on the mechanism of HIV-1 infection of macrophages but also aid the development of novel anti-viral drug to block HIV-1 infection of macrophages.

巨噬细胞和T细胞是HIV-1感染的主要靶细胞。相比T细胞，被HIV-1感染的巨噬细胞能相对持久地维持病毒复制，并可通过细胞间感染介导HIV-1高效传播，因此在病毒增殖、传播和致病方面具有重要作用。宿主因子与病毒之间的相互作用决定了HIV-1感染巨噬细胞的进程。我们前期研究发现宿主蛋白PQBP1能结合巨噬细胞内的HIV-1复制复合体并抑制病毒复制，但其抑制机理尚不清楚。本项研究拟1）利用可视化方法在HIV-1感染的巨噬细胞内分析PQBP1与病毒复制复合体的相互作用；2）鉴定与PQBP1相互作用的病毒成分及相互作用结构域；3）定量检测PQBP1对病毒复制各阶段产物及复制复合体稳定性的影响，明确PQBP1抑制的病毒复制特定阶段。最终综合上述工作阐明PQBP1抑制巨噬细胞内HIV-1复制的分子机理。该项研究不仅将加深对HIV-1感染巨噬细胞机理的理解，也将为新型抗HIV-1药物设计提供新思路。

HIV-1感染者体内的主要靶细胞是巨噬细胞和T细胞。T细胞被HIV-1感染后会快速地发生细胞死亡，造成T细胞耗竭及感染者免疫功能衰竭。相比而言，被HIV-1感染的巨噬细胞能更持久地维持病毒复制，并可通过细胞间传播介导HIV-1高效扩增，因此在HIV-1的感染致病过程中扮演重要作用。宿主因子与病毒之间的相互作用调控了HIV-1对巨噬细胞的感染。在本项目中，我们发现敲降宿主蛋白PQBP1后HIV-1的复制受到显著抑制，且抑制主要发生在HIV-1核入侵阶段。进一步研究发现，PQBP1结合宿主因子CPSF6并介导其高效入核。在PQBP1敲降的细胞中，CPSF6出现在细胞质内的累积，从而结合细胞质内的HIV-1复制复合体，并抑制其入核。进一步研究鉴定到了PQBP1上65位络氨酸及75位色氨酸是与CPSF6相互作用的关键位点，是其介导CPSF6入核的潜在位点。本项研究首次揭示了HIV-1复制过程中两个重要的宿主因子之间的相互作用调控HIV-1核入侵事件的机理，为新型抗HIV-1药物设计提供了潜在靶点。