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1型人类免疫缺陷病毒DNA的体外环化

Circularization of human immunodeficiency virus type 1 DNA in vitro.

作者信息

Farnet C M, Haseltine W A

机构信息

Division of Human Retrovirology, Dana-Farber Cancer Institute, Boston, Massachusetts.

出版信息

J Virol. 1991 Dec;65(12):6942-52. doi: 10.1128/JVI.65.12.6942-6952.1991.

DOI:10.1128/JVI.65.12.6942-6952.1991
PMID:1834863
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC250802/
Abstract

Linear viral DNA present in cytoplasmic extracts of cells newly infected with human immunodeficiency virus type 1 can be induced to form 1-LTR and 2-LTR circles by incubation of the extracts in the presence of added nucleoside triphosphates. No circular DNA forms are detected when extracts are incubated in the absence of added nucleoside triphosphates. Restriction enzyme analysis and polymerase chain reaction analysis with selected primers, as well as DNA sequence analysis of the polymerase chain reaction products, show that most of the 2-LTR circles are the result of autointegration reactions, while 1-LTR circles result from recombination between the long terminal repeats on the linear viral DNA. In addition, a small amount of simple 2-LTR circles, formed by end-to-end joining of the linear viral DNA, is formed in vitro. Integration of the linear viral DNA into heterologous DNA competes effectively with the formation of 2-LTR circles by autointegration. However, concentrations of target DNA which completely block autointegration have no effect on the formation of 1-LTR circles or simple 2-LTR circles. Factors present in extracts of uninfected cells can mediate the formation of 1-LTR circles and simple 2-LTR circles from purified deproteinated linear viral DNA, indicating that viral proteins are not necessary for the formation of these two types of circular viral DNA. These experiments demonstrate that all the transformations of linear viral DNA which occur in the nuclei of cells infected with human immunodeficiency virus type 1 can be reproduced in vitro.

摘要

在新感染1型人类免疫缺陷病毒的细胞的细胞质提取物中存在的线性病毒DNA，通过在添加核苷三磷酸的情况下孵育提取物，可被诱导形成1-LTR和2-LTR环。当提取物在不添加核苷三磷酸的情况下孵育时，未检测到环状DNA形式。用选定引物进行的限制性内切酶分析和聚合酶链反应分析，以及聚合酶链反应产物的DNA序列分析表明，大多数2-LTR环是自整合反应的结果，而1-LTR环是线性病毒DNA上长末端重复序列之间重组的结果。此外，在体外形成了少量由线性病毒DNA端对端连接形成的简单2-LTR环。线性病毒DNA整合到异源DNA中有效地与通过自整合形成2-LTR环竞争。然而，完全阻断自整合的靶DNA浓度对1-LTR环或简单2-LTR环的形成没有影响。未感染细胞提取物中存在的因子可介导从纯化的脱蛋白线性病毒DNA形成1-LTR环和简单2-LTR环，这表明病毒蛋白对于这两种类型的环状病毒DNA的形成不是必需的。这些实验表明，在感染1型人类免疫缺陷病毒的细胞核中发生的线性病毒DNA的所有转化都可以在体外重现。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/64717391e18d/jvirol00055-0586-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/4cb73d3892ae/jvirol00055-0581-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/3f91b592f0ef/jvirol00055-0582-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/90b95e24eef7/jvirol00055-0582-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/f2732f31d02b/jvirol00055-0583-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/bf34c6baa963/jvirol00055-0584-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/07fe07885e98/jvirol00055-0585-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/3ea94ab7a21b/jvirol00055-0585-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/64717391e18d/jvirol00055-0586-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/4cb73d3892ae/jvirol00055-0581-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/3f91b592f0ef/jvirol00055-0582-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/90b95e24eef7/jvirol00055-0582-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/f2732f31d02b/jvirol00055-0583-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/bf34c6baa963/jvirol00055-0584-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/07fe07885e98/jvirol00055-0585-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/3ea94ab7a21b/jvirol00055-0585-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5540/250802/64717391e18d/jvirol00055-0586-a.jpg

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