HIV-1 INDUCES G2/M CELL CYCLE ARREST

已经提出了各种机制来解释这种免疫 艾滋病的功能障碍最终会导致CD4+T细胞的丧失，包括 包膜介导的合胞体形成、凋亡和细胞因子调节。 我们最近的研究结果提出了一个新的假说--细胞功能障碍。我们 使用携带新细胞的人类免疫缺陷病毒1型(HIV-1) 表面报告基因，小鼠THY1.2，以区分感染和 同一种群中未感染的细胞。我们发现被感染的细胞， 无论是来自细胞系还是原代T细胞，都不能正常进行 通过细胞周期，并被阻止在G2/M期。其中一个 HIV-1基因vpr与许多功能有关，包括 潜伏的HIV-1的反式激活、核靶向和重新激活。 我们证明，细胞生长受阻可以归因于 HIV-1的vpr基因产物，因为该基因的突变允许细胞周期 感染细胞的进展。此外，通过瞬时转染法 分析表明，单独表达vpr可以诱导细胞周期。 停滞于SupT1 T细胞和HeLa细胞。使用不同的记者 系统，荧光素酶，我们调查G2被捕对艾滋病毒的后果- 1个复制。我们的结果表明，HIV-1前病毒不能维持 由于感染细胞的选择性生长劣势。基因突变 VPR导致感染的维持。因此，VPR的表达防止了 建立慢性HIV-1感染。这些结果有两个 与HIV-1致病机制相关的预测：1)HIV-1 感染一般不会导致持续的慢性感染 以及2)被感染的T细胞将不能 G2/M期停滞导致的克隆性扩张，并对免疫产生影响 功能障碍。我们建议在此扩展我们的初步发现，以开发 对其作用机制的进一步洞察。 具体目标是： 1.确定G2/M期细胞停滞表型与 Vpr基因的其他观察到的功能。结构-功能 关系将通过遗传方法进行调查。 2.比较HIV-1、HIV-2和VPR相关基因的功能 猴免疫缺陷病毒(SIV)与细胞周期停滞有关。我们 我还将比较VPR的功能与进化上的 保守的VPX基因。

Various mechanisms have been proposed to account for the immune dysfunction in AIDS ultimately leading to loss of CD4+ T-cells, including envelope-mediated syncytium formation, apoptosis and cytokine modulation. Our recent results suggest a novel hypothesis forT-cell dysfunction. We used a human immunodeficiency virus type 1 (HIV-1) bearing a novel cell surface reporter gene, murine thy 1.2, to distinguish infected from uninfected cells in the same population. We show that infected cells, both from cell lines and primary T-cells, are unable to progress normally through the cell cycle and become arrested in the G2/M phase. One of the HIV-1 genes, vpr, has been implicated in a number of functions, including trans-activation, nuclear targeting and re-activation of latent HIV-1. We demonstrate that the arrest of cell growth can be attributed to the vpr gene product of HIV-1, as mutation of this gene allows cell cycle progression of the infected cells. Furthermore, by transient transfection assays, we show that expression of vpr alone will induce cell cycle arrest in SupT1 T-cells and HeLa cells. Using a different reporter system, luciferase, we investigate the consequences of G2 arrest for HIV- 1 replication. Our results show that the HIV-1 provirus is not maintained due to the selective growth disadvantage of infected cells. Mutation of vpr results in maintenance of infection. Thus, expression of vpr prevents the establishment of a chronic HIV-1 infection. These results have two predictions that are relevant to HIV-1 pathogenesis: 1) that HIV-1 infection does not generally lead to a chronic infection with persistence of provirus; and 2) that infected T-cells would be unable to undergo clonal expansion as a result of G2/M arrest, with consequences for immune dysfunction. We propose here to extend our initial findings to develop further insights into its mechanism of action. The Specific Aims are to: 1. Determine the relationship between the G2/M arrest phenotype and other observed functions of the vpr gene. Structure-function relationships will be investigated through genetic approaches. 2. Compare the functions of vpr-related genes from HIV-1, HIV-2 and simian immunodeficiency virus (SIV) in regards to cell cycle arrest. We will also compare the functions of Vpr with those of the evolutionarily conserved vpx gene.