Vink C, van Gent D C, Plasterk R H
Division of Molecular Biology, The Netherlands Cancer Institute, Amsterdam.
J Virol. 1990 Oct;64(10):5219-22. doi: 10.1128/JVI.64.10.5219-5222.1990.
An essential step in the life cycle of the human immunodeficiency virus (HIV) is integration of a DNA copy of the viral RNA into the genome of the infected cell. We show here that this step can be faithfully accomplished in vitro by the enzymatic machinery of another retrovirus, Moloney murine leukemia virus (MoMLV). Mini-HIV substrates, which are linearized plasmids with long terminal repeat sequences at their ends, were incubated with cytoplasmic extracts of MoMLV-infected NIH 3T3 cells and target DNA. The MoMLV integration apparatus carried out integration of the mini-HIV substrates correctly; the terminal nucleotides of the viral substrate were removed, and a 4-base-pair duplication of the target DNA flanked the inserted viral DNA (C. Shoemaker, S. P. Goff, E. Gilboa, M. Paskind, S. W. Mitra, and D. Baltimore, Proc. Natl. Acad. Sci. USA 77:3932-3936, 1980). Our experiments show that the substrate sequence requirements for integration in vitro were limited to a few nucleotides, as the similarity between HIV and MoMLV long terminal repeat ends is minimal.
人类免疫缺陷病毒（HIV）生命周期中的一个关键步骤是将病毒RNA的DNA拷贝整合到受感染细胞的基因组中。我们在此表明，这一步骤可以在体外由另一种逆转录病毒莫洛尼鼠白血病病毒（MoMLV）的酶促机制忠实地完成。将末端带有长末端重复序列的线性化质粒微型HIV底物与感染了MoMLV的NIH 3T3细胞的细胞质提取物和靶DNA一起孵育。MoMLV整合装置正确地完成了微型HIV底物的整合；病毒底物的末端核苷酸被去除，插入的病毒DNA两侧是靶DNA的4个碱基对重复序列（C. Shoemaker、S.P. Goff、E. Gilboa、M. Paskind、S.W. Mitra和D. Baltimore，《美国国家科学院院刊》77:3932 - 3936，1980）。我们的实验表明，体外整合的底物序列要求仅限于几个核苷酸，因为HIV和MoMLV长末端重复序列末端之间的相似性极小。
