MOLECULAR CLONING AND CHARACTERIZATION OF HUMAN RETROVIRUSES

HIV-1分离株可根据以下情况分为两个主要亚组： 它们在体外的细胞宿主范围：巨噬细胞（MT）和T细胞系 热带的 嗜MT分离株感染巨噬细胞和外周血 但是，单个核细胞（PBMC）布塔雷不能在转化的CD 4+细胞中复制。 T细胞系 T细胞系嗜性分离物感染PBMC和CD 4 + T细胞。 细胞系，但在原代巨噬细胞中复制较差或根本不复制 （MDM）。 虽然T细胞是HIV-1复制的主要目标， 外周血中，巨噬细胞代表主要的HIV-1感染者 大多数组织中的细胞类型。 巨噬细胞可能是 HIV-1的储存库，可能对维持持续的 多年来，个人感染。 大多数HIV-1分离株 克隆的是T细胞嗜性的。我们成功地获得了一个完整的 从嗜巨噬细胞病毒分离物的分子克隆。初步 生物化学和物理分析表明， 包膜蛋白（gp 120）的脱落与 典型的T细胞系变体。 还有几种辅料的效果 基因（体外感染中的非必需基因）似乎是不稳定的。 VPU、VPR或NEF的突变适度降低了病毒复制。 任一PBMC巨噬细胞中的AD 8 -2克隆。 通过与其它全长感染性嗜巨噬细胞克隆的比较 在HIV-1中，pAD 8定向的HIV-1AD 8储备物在2000年生长至高滴度。 人MDM培养物，如通过p24抗原捕获测定监测的。后代 在HIVAD 8感染的原代人巨噬细胞中的病毒体产生也是 通过不太敏感的逆转录酶（RT）容易测量 比色法

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) butare 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 in 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 macrophages. By comparison with other full-length infectious macrophage-tropic clones of HIV-1, the pAD8-directed HIV-1AD8 stock grew to high titers in cultures of human MDM, as monitored by p24 antigen capture assay. Progeny virion production in HIVAD8-infected primary human macrophages was also readily measurable by the less sensitive reverse transcriptase (RT) assay.