MOLECULAR CLONING AND CHARACTERIZATION OF HUMAN RETROVIRUSES

人类免疫缺陷病毒(HIV-1)的基因组大小为 约9.7kB，带有编码几种病毒的开放阅读框 蛋白质。基因产品(结构、调节和辅助病毒 蛋白质)由剪接和非剪接的mRNAs产生，并可影响病毒 在各种细胞类型中复制。由于HIV-1之间异质性 逆转录病毒基因组是慢病毒的一个明显特征，分子 克隆可以为研究它们的生物化学提供有用的试剂 和物理特性。 根据HIV-1分离株可分为两个主要亚组 其体外细胞宿主范围：巨噬细胞(MT)和T细胞系 北极星。嗜甲氧西林金黄色葡萄球菌可感染巨噬细胞和外周血细胞 血液单个核细胞(PBMC)，但不能在 转化的CD4+T细胞系。T细胞系热带分离株感染这两种 PBMC和CD4+T细胞株，但复制很差或根本不原代 巨噬细胞MDM)。尽管T细胞是HIV-1的主要靶点 在外周血中复制，巨噬细胞是主要的 大多数组织中存在HIV-1感染细胞类型。巨噬细胞可能是 是HIV-1的主要宿主，可能对维持 在个人身上持续感染多年。大多数HIV-1病毒 我们克隆的菌株是嗜T细胞的。我们已经成功地 从嗜巨噬细胞病毒中获得完整的分子克隆 隔离。初步的生化和物理分析表明 被膜蛋白(Gp120)的自发脱落 与典型的T细胞系变种不同。此外，还包括 出现了几个辅助基因(在体外感染中不是必需的) 是可有可无的。VPU、VPR或NEF突变略有减少 AD8-2克隆在PBMC或巨噬细胞中的病毒复制。

The genomic size of Human Immunodeficiency Virus (HIV-1) is approximately 9.7 kB with open reading frames encoding several viral proteins. Gene products (structural, regulatory and accessory viral proteins) arise from spliced and unspliced mRNAs and can affect viral replication in various cell types. Since heterogeneity among HIV-1 retroviral genomes is a distinct feature of Lentiviruses, molecular clones could provide useful reagents for studying their biochemical and physical properties. HIV-1 isolates can be divided into two major subgroups on the basis of their cellular host range in vitro: macrophage (MT) and T-cell line tropic. MT-tropic isolates infect both macrophages and peripheral blood mononuclear cells (PBMC) but are unable to replicate in transformed CD4+ T-cell lines. T-cell line tropic isolates infect both PBMC and CD4+ T-cell lines but replicate poorly or not at all primary macrophages MDM). Although T-cells are the major target for HIV-1 replication in peripheral blood, macrophages represent the predominant HIV-1 infected cell type in most tissues. Macrophages are probably the primary reservoir of HIV-1 and may be important for sustaining a persistent infection in individuals for many years. Most HIV-1 isolates we have cloned are T-cell tropic. We have succeeded in obtaining a complete molecular clone from a macrophage-tropic viral isolate. Preliminary biochemical and physical analyses have shown that the spontaneous shedding of the envelope protein(gp120) is drastically different from the typical T-cell line variants. Also the effects of several accessory genes (non-essential in in vitro infections) appear to be dispensable. Mutations in VPU, VPR, or NEF modestly reduced viral replication of the AD8-2 clone in either PBMC or macrophages.