BIOLOGICAL EFFECTS OF HIV-1 GENETIC VARIABILITY

HIV-1 遗传变异的生物学影响

• 批准号: 3142540
• 负责人: Maureen M Goodenow
• 金额: $ 13.15万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-12-01 至 1994-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3142540
• 关键词: AIDS; AIDS /HIV diagnosis; HIV infections; cell type; child (0-11); circular DNA; gene expression; genetic promoter element; genetic regulatory element; human genetic material tag; human immunodeficiency virus 1; human subject; latent virus infection; nucleic acid repetitive sequence; nucleic acid sequence; pediatric AIDS; polymerase chain reaction; tissue /cell culture; virus DNA; virus genetics; virus infection mechanism; virus replication

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.

人类免疫缺陷病毒 1 型 (HIV-1)，其病原体 获得性免疫缺陷综合症（艾滋病）和相关复合物，显示出 高度的遗传和生物变异性。 HIV-1的多样性 对于跟踪艾滋病的流行病学、了解艾滋病具有重要意义 病毒的病理学以及治疗方法的开发， 预防和诊断试剂。 有一些重要的问题需要解答 回答了将遗传差异与病毒生物学联系起来的问题， 包括其潜伏期及其在不同环境中感染和复制的能力 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尚未得到广泛研究 评价。 整合到宿主细胞基因组中涉及共价连接 HIV-1 LTR 和细胞 DNA 之间的关系。 一般来说，未积分的线性或 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。 然而，未整合的病毒形式在培养物中积累 感染 HIV-1 的细胞似乎与体外相关 细胞病变。 细胞内未整合的 HIV-1 DNA 的流行 感染者及其与 HIV-1 发病机制的关系（如果有） 尚未确定。 拟议的研究特别针对：（1） 直接在感染个体的细胞中评估感染的程度 HIV-1 LTR 内的序列异质性； (2) 评估患病率 感染者体内未整合的 HIV-1 形式； (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遗传评估 直接在感染个体中进行多样性，无需体外选择 HIV-1 分离株的生长。 研究将重点关注临床疾病 儿科艾滋病患者及其无症状、血清反应呈阳性的母亲 评估患有艾滋病毒的个体之间随时间推移产生的 HIV-1 多样性 感染相关病毒的不同临床状态。 测序 LTR 使用聚合酶链式反应 (PCR) 进行酶促扩增后 将评估 LTR 变异的程度。 瞬态表达 受变体 LTR 控制的指示基因将评估生物学特性 LTR启动子功能和细胞类型的序列多样性的影响 特定的基因表达。 所提出的实验旨在测试 假设 HIV-1 LTRS 中存在自然遗传变异；那个 LTR 感染病毒的个体内部和个体之间的体内多样性发展 相关病毒；遗传多样性不仅有助于 LTRS 的功能变异性，但也与临床相关 HIV-1 感染者的状况。