Du Z, Ilyinskii P O, Lally K, Desrosiers R C, Engelman A
New England Regional Primate Research Center, Harvard Medical School, Southborough, Massachusetts 01772, USA.
J Virol. 1997 Nov;71(11):8124-32. doi: 10.1128/JVI.71.11.8124-8132.1997.
Sequences at the left terminus of U3 in the left long terminal repeat (LTR) and at the right terminus of U5 in the right LTR are important for integration of retroviral DNA. In the infectious pathogenic molecular clone of simian immunodeficiency virus strain mac239 (SIVmac239), 10 of the 12 terminal base pairs form an imperfect inverted repeat structure (5' TGGAAGGGATTT 3' [nucleotides 1 to 12] and 3' ACGATCCCTAAA 5' [nucleotides 10279 to 10268]). Nineteen different mutant forms of SIVmac239 proviral DNA with changes at one or more of the positions in each of the 12-terminal-base-pair regions were constructed. Viral replication was severely or completely compromised with nine of these mutants. Revertants appeared 40 to 50 days after transfection in two independent experiments with mutant 7, which contained changes of AGG to TAC at positions 5 to 7 in U3 and TCC to GAA at positions 10275 to 10273 in U5. Virus produced at these times from mutant 7 transfection replicated upon reinfection with only a slight delay when compared to the wild type. Sequence analysis of the LTR and integrase regions from infected cultures revealed two predominant changes: G to A at position 10275 in U5 and Glu to Lys at position 136 in integrase. Derivatives of clone 7 in which these changes were introduced individually and together were constructed by site-specific mutagenesis. Each change individually restored replication capacity only partially. However, the combination of both mutations restored replicative capacity to that of the original revertants. These results indicate that changes in integrase can compensate for mutations in the terminal nucleotides of the SIV LTR. The results further indicate that resistance to integrase inhibitors may include both integrase and LTR mutations.
左长末端重复序列（LTR）中U3的左末端序列以及右LTR中U5的右末端序列对于逆转录病毒DNA的整合至关重要。在猿猴免疫缺陷病毒株mac239（SIVmac239）的感染性致病分子克隆中，12个末端碱基对中的10个形成了一个不完全的反向重复结构（5'TGGAAGGGATTT 3'[核苷酸1至12]和3'ACGATCCCTAAA 5'[核苷酸10279至10268]）。构建了19种不同的SIVmac239前病毒DNA突变形式，这些突变形式在12个末端碱基对区域的每个区域中的一个或多个位置发生了变化。其中9种突变体的病毒复制受到严重损害或完全受损。在两个独立的实验中，用突变体7转染后40至50天出现了回复体，突变体7在U3的第5至7位包含AGG到TAC的变化，在U5的第10275至10273位包含TCC到GAA的变化。与野生型相比,这些时候由突变体7转染产生的病毒在再次感染时仅稍有延迟就能复制。对感染培养物的LTR和整合酶区域进行序列分析发现了两个主要变化：U5中第10275位的G变为A以及整合酶中第136位的Glu变为Lys。通过位点特异性诱变构建了单独引入和一起引入这些变化的克隆7衍生物。每个变化单独仅部分恢复了复制能力。然而，两种突变的组合将复制能力恢复到了原始回复体的水平。这些结果表明，整合酶的变化可以补偿SIV LTR末端核苷酸的突变。结果还进一步表明，对整合酶抑制剂的抗性可能包括整合酶和LTR突变。
