核转运蛋白KPNA2在HIV-1整合前复合体入核过程中的作用机制研究

In the process of HIV replication, the penetration of viral pre-integration complex (PIC) through nuclear pore is an important step in the replication cycle. Although some of the nuclear pore proteins and viral proteins have been reported to participate in the PIC transmission, the molecular mechanism of HIV-1 nuclear transport is more controversial because it involves many kinds of viral and host interaction. Recently, we found that karyopherin α2 (KPNA2) in human monocytes was significantly up-regulated in macrophage differentiation and associated with HIV replication. We further confirmed that down-regulation of KPNA2 inhibits HIV replication. The inhibition occurs mainly in the nuclear import steps. HIV replication complex RTC/PIC co-localized with KPNA2 in cytoplasm. Based on these evidences, we speculated that KPNA2 play an important role in the process of HIV nuclear import. We propose to further study of HIV nuclear import molecular mechanism by   report virus infection, the fluorescent microscopy, co-immune precipitation and other technical methods. These include the interaction of KPNA2 with viral protein and molecular pathways downstream of transport. This study will elucidate the new mechanism of KPNA2 for HIV nuclear import, and also provide theoretical basis for potential targets in HIV anti-viral drug design.

HIV复制过程中，病毒整合前复合体PIC通过细胞核孔是复制周期中一个重要步骤。虽然部分细胞核孔蛋白和病毒蛋白已经被报道参与PIC入核过程，但是HIV-1核转运涉及多种病毒蛋白与宿主相互作用，其分子机制还存在较多争议。我们近期研究发现，核转运蛋白KPNA2在人单核细胞到巨噬细胞分化过程中显著上调并和HIV复制相关；进一步我们确认下调KPNA2抑制HIV复制；复制抑制主要发生在入核步骤；HIV复制复合体RTC/PIC在细胞质与KPNA2有显著共定位，因此我们推测KPNA2在HIV核转运过程中起重要作用。本研究拟采用报告病毒感染、荧光标记显微技术、免疫共沉淀等多种技术手段，深入研究KPNA2在HIV PIC入核转运过程中的分子机制，包括与KPNA2相互作用的病毒蛋白以及下游转运分子通路。本研究将阐明KPNA2调控HIV复制入核的全新机制，也有望为设计HIV抗病毒药物的潜在作用靶点提供依据。

HIV复制过程中，病毒整合前复合体PIC通过细胞核孔是复制周期中一个重要步骤。虽然部分细胞核孔蛋白和病毒蛋白已经被报道参与PIC入核过程，但是HIV-1核转运涉及多种病毒蛋白与宿主相互作用，其分子机制还存在较多争议。通过此项目的工作，我们开展了核转运蛋白KPNA2在HIV-1病毒复制过程中PIC入核的作用机制研究。我们的结果发现，核转运蛋白KPNA2在人单核细胞到巨噬细胞分化过程中显著上调并和HIV复制相关；进一步我们确认下调KPNA2抑制HIV复制；复制抑制主要发生在入核步骤；HIV复制复合体RTC/PIC在细胞质与KPNA2有显著共定位，更为重要的是我们发现KPNA2和HIV-1衣壳蛋白p24相互作用，过表达N端 p24可以显著抑制HIV-1 PIC的入核，而且PIC KPNA2介导的入核是KPNB1依赖的，结合我们先前研究我们揭示了一条新的HIV-1 PIC入核的途径：KPNA2-KPNB1-pom121, 同时本研究也发现了一个潜在的抑制HIV-1 PIC入核的靶点，即N端p24, 后续研究我们将集中力量设计HIV-1 入核的抑制肽，开发一种抗HIV-1感染，作用于入核阶段的抗病毒药物。