BIOLOGICAL EFFECTS OF HIV-1 GENETIC VARIABILITY

人类免疫缺陷病毒1型（HIV-1）， 获得性免疫缺陷综合征（艾滋病）和相关的复合物，显示了一个 高度遗传和生物变异性。 HIV-1的多样性 对追踪艾滋病的流行病学，了解 病毒的病理学，以及治疗的发展， 预防和诊断试剂。 有一些重要的问题 将基因差异与病毒的生物学联系起来， 包括它的潜伏期以及感染和复制不同细胞的能力 细胞类型。 参与调节HIV-1大部分方面的序列 生命周期，特别是病毒基因表达和整合， 位于5'和3'末端的长末端重复序列（LTR）内， 线性病毒分子。 遗传变异的直接贡献 HIV-1 LTR内的cic作用转录调控元件， 体外和体内的表型变异性尚未得到广泛研究。 评估。 整合到宿主细胞基因组中涉及共价连接 HIV-1 LTR和细胞DNA之间的联系 一般来说，不可积分的线性或 逆转录病毒的环状DNA形式仅以短暂的复制形式出现， 中间体的 然而，未整合的病毒形式在培养的细胞中积累， 感染HIV-1的细胞，并似乎与体外 细胞病变 未整合的HIV-1 DNA在人细胞内的流行率 感染个体及其与HIV-1发病机制的关系（如果有的话） 尚未确定。 建议的研究特别针对：（1） 直接在受感染个体的细胞中评估 HIV-1 LTR内的序列异质性;（2）评估 感染个体中未整合形式的HIV-1;以及（3）确定 功能测定中自然遗传变异的生物学效应。 实验设计的一个重要方面是评估遗传学的影响。 直接在感染个体中进行多样性，而无需体外选择 HIV-1分离株的生长。 这些研究将集中在临床疾病 儿童艾滋病患者及其无症状、血清反应阳性的母亲， 评估HIV-1多样性随时间的产生， 不同临床状态感染相关病毒。 测序LTR 在使用聚合酶链式反应（PCR）的酶促扩增之后， 将评估LTR可变性的程度。 瞬时表达 在变体LTR控制下的指示基因将评估生物学特性， LTR启动子功能序列多样性与细胞类型的关系 特异性基因表达 建议的实验旨在测试 HIV-1 LTR发生自然遗传变异的假设， 在体内和感染的个体之间产生多样性 基因多样性不仅有助于 LTRS的功能变异性，但也与临床 艾滋病毒1感染者的状况。

Human immunodeficiency virus type 1 (HIV-1), the etiological agent of acquired immunodeficiency syndrome (AIDS) and related complexes, displays a high degree of genetic and biological variability. The diversity of HIV-1 has implications for tracking the epidemiology of AIDS, for understanding the pathology of the virus, and for the development of therapeutic, preventive, and diagnostic reagents. There are important questions to be answered relating genetic differences to the biology of the virus, including its latency and its ability to infect and replicate in different cell types. Sequences involved in regulating most aspects of the HIV-1 life cycle, in particular viral gene expression and integration, are located within the long terminal repeats (LTRs) at the 5' and 3' ends of linear viral molecules. The direct contribution of genetic variability in cic-acting transcriptional regulatory elements within HIV-1 LTRs to phenotypic variability in vitro and in vivo has not been extensively evaluated. Integration into the host-cell genome involves covalent linkage between HIV-1 LTRs and cellular DNA. In general, unintergrated linear or circular DNA forms of retroviruses occur only as short-lived replicative intermediates. However, unintegrated viral forms accumulate in cultured cells infected with HIV-1 and appear to correlate with in vitro cytopathicity. The prevalence of unintegrated HIV-1 DNA within cells of infected individuals and its relationship, if any, to HIV-1 pathogenesis has not been determined. The proposed studies are specially aimed at: (1) evaluating directly in cells from infected individuals the extent of sequence heterogeneity within HIV-1 LTRs; (2) assessing the prevalence of unintegrated forms of HIV-1 in infected individuals; and (3) determining the biological effects of natural genetic variability in functional assays. An important aspect of the experimental design is evaluation of genetic diversity directly in infected individuals without selection for in vitro growth of HIV-1 isolates. The studies will focus on clinically ill pediatric AIDS patients and their asymptomatic, seropositive mothers to assess the generation of HIV-1 diversity over time between individuals with different clinical status infected with related viruses. Sequencing LTRs after enzymatic amplification using the polymerase chain reaction (PCR) will assess the extent of LTR variability. Transient expression of an indicator gene under control of variant LTRs will evaluate the biological effects of sequence diversity of LTR promoter function and cell-type specific gene expression. Proposed experiments are designed to test the hypotheses that natural genetic variability occurs in HIV-1 LTRS; that LTR diversity develops in vivo within and between individuals infected with related viruses; and that genetic diversity not only contributes to functional variability of LTRS, but is also associated within the clinical status of individuals infected with HIV-1.