Ryan-Graham M A, Peden K W
Laboratory of Molecular Microbiology, NIAID, NIH, Bethesda, Maryland 20892, USA.
Virology. 1995 Oct 20;213(1):158-68. doi: 10.1006/viro.1995.1556.
While it has been demonstrated that the Nef protein of simian immunodeficiency virus is obligatory for the establishment of high viral loads and the development of simian AIDS in rhesus macaques, demonstrating a critical role for the human immunodeficiency virus (HIV) Nef protein in tissue culture has been elusive. Data have been contradictory as to whether Nef has a negative or positive influence on in vitro virus replication. In an attempt to define a role for Nef during virus propagation in tissue culture and to obtain virus-host systems that could distinguish between the Nef mutant and wild-type viruses, we have introduced mutations into the nef genes of infectious molecular clones of three HIV-1 strains and two isolates of the HIV-2ROD strain and have investigated the capacity of viruses derived from them to infect a number of CD4-positive T-cell lines and peripheral blood mononuclear cells (PBMC). Mutating the nef gene of all viruses had a modest negative effect on virus production in activated PBMC. In some T-cell lines with some viruses, the effects were severe, and little or no Nef mutant virus could be detected. In other cell lines, the result of mutating the nef gene either had no effect or had a slight negative effect on the replication kinetics. Therefore, whether the consequences of loss of Nef activity can be demonstrated in vitro depends on both the particular virus and the host cell used, suggesting that Nef is exerting its activity on some cellular pathway. In addition, we describe the construction and properties of hitherto unreported infectious molecular clones of the ROD strain of HIV-2.
虽然已经证明猿猴免疫缺陷病毒的Nef蛋白对于恒河猴体内高病毒载量的建立和猿猴艾滋病的发展是必不可少的，但在组织培养中证明人类免疫缺陷病毒（HIV）Nef蛋白的关键作用却一直难以实现。关于Nef对体外病毒复制是产生负面影响还是正面影响，数据一直相互矛盾。为了确定Nef在组织培养中病毒传播过程中的作用，并获得能够区分Nef突变病毒和野生型病毒的病毒-宿主系统，我们对三种HIV-1毒株和两株HIV-2 ROD毒株的感染性分子克隆的nef基因进行了突变，并研究了由此产生的病毒感染多种CD4阳性T细胞系和外周血单核细胞（PBMC）的能力。突变所有病毒的nef基因对活化的PBMC中的病毒产生有适度的负面影响。在一些病毒感染的某些T细胞系中，影响很严重，几乎检测不到或根本检测不到Nef突变病毒。在其他细胞系中，nef基因突变的结果要么对复制动力学没有影响，要么有轻微的负面影响。因此，Nef活性丧失的后果是否能在体外得到证明取决于所使用的特定病毒和宿主细胞，这表明Nef正在对某些细胞途径发挥其活性。此外，我们描述了HIV-2 ROD毒株迄今未报道的感染性分子克隆的构建和特性。
