Role of Vif in HIV-1 Replication/AIDS

HIV-1 vif基因对非允许细胞(NP)产生感染性病毒是必不可少的，非允许细胞包括CD4阳性T淋巴细胞和巨噬细胞以及一些白血病T细胞系，但在其他被称为允许细胞(P)的细胞系中无关。P细胞来源的HIV-1病毒可感染NP细胞，但感染的NP细胞只释放非感染性病毒粒子。这些非感染性病毒粒子不能通过在靶细胞中表达vif来挽救，而且它们被认为存在缺陷，降低了反转录的效率。我们和另一个实验室最近发现了PxNP异核体中NP表型的优势，这表明NP细胞中含有一种可以灭活HIV-1的因子，该因子可以被Vif抵消。利用酵母双杂交文库，以Vif为诱饵，我们现在已经确定干扰素诱导蛋白Sp140是该NP因子的一个很好的候选者。简而言之，Sp140在所有被测试的NP细胞中都存在，但在P细胞中不存在。Sp140存在于含有几种蛋白质的核体(NBS)中，这些蛋白质通过添加泛素相关蛋白SUMO-1而共价修饰。Sp140在Hela-CD4细胞中的表达诱导至少一种蛋白质的总和基化，而Vif的共表达阻断了这一诱导。HIV-1感染可诱导Sp140向胞浆迁移，并与Vif部分共存。我们建议：(I)通过遗传学、免疫学和生物化学方法验证Vif与Sp140和其他潜在合作伙伴的相互作用。(Ii)分析Sp140亚型和其他潜在的Vif合作伙伴使vif缺失的HIV-1失活的能力。(3)确定VIF与SP140以及与其他潜在合作伙伴互动的活跃地点。(4)通过双向电泳法，鉴定Sp140诱导的P细胞中的总和蛋白和非总和蛋白，并确定Vif对这些诱导的影响。同样，确定Vif在NP细胞中的表达是否改变了蛋白质的相加作用，以及在P或NP细胞中产生的Vif缺失的HIV-1是否含有任何相加蛋白。(Iv)有趣的是，1型单纯疱疹病毒ICPO蛋白以NBS为靶标，并特异性地消除一些SUMoyl化的Nb蛋白，而CMV、EBV、腺病毒和ArenaVirus编码的蛋白似乎具有类似的活性。确定这些其他病毒蛋白的表达是否会将NP细胞转化为P。这项工作可能会揭示艾滋病药物开发的新靶点。

The HIV-1 vif gene is essential for infectious virus production by nonpermissive cells (NP), which include CD4-positive T-lymphocytes and macrophages and some leukemic T-cell lines, but is irrelevant in other cell lines that are termed permissive (P). NP cells can be infected with vif-deleted HIV-1 derived from P cells, but the infected NP cells release only noninfectious virions. These noninfectious virions cannot be rescued by vif expression in target cells, and they are believed to have a defect that reduces efficiency of reverse transcription. We and another laboratory recently showed the dominance of the NP phenotype in PxNP heterokaryons, suggesting that NP cells contain a factor that can inactivate HIV-1 and that this factor is counteracted by Vif. Using a human lymphocyte cDNA library in a yeast two-hybrid screen with Vif as bait, we have now identified the interferon-inducible protein Sp140 as an excellent candidate for this NP factor. Briefly, Sp140 occurs in all tested NP cells but not in P cells. Sp140 occurs in nuclear bodies (NBs) that contain several proteins covalently modified by addition of the ubiquitin-related protein Sumo-1. Expression of Sp140 in Hela-CD4 cells induces sumoylation of at least one protein, and coexpression of Vif blocks this induction. HIV-1 infections induce Sp140 emigration to the cytosol and its partial colocalization with Vif. We propose: (i) Verify Vif interaction with Sp140 and with other potential partners by genetic, immunological, and biochemical methods. (ii) Analyze Sp140 isoforms and other potential Vif partners for their abilities to inactivate vif-deleted HIV-1. (iii) Identify active sites for interaction of Vif with Sp140 and with other potential partners. (iv) By two-dimensional electrophoresis, identify sumoylated and non-sumoylated proteins induced in P cells by Sp140, and determine how Vif influences these inductions. Similarly, determine whether Vif expression in NP cells alters protein sumoylation and whether vif-deleted HIV-1 made in P or NP cells contains any sumoylated protein. (iv) Interestingly, the herpes simplex virus type 1 ICPO protein targets NBs and specifically causes the elimination of some sumoylated NB proteins, and proteins encoded by CMV, EBV, adenoviruses and arenaviruses appear to have similar activities. Determine whether expression of these other viral proteins converts NP cells to P. This work may unveil a novel target for drug development in AIDS.