PDF(Pubmed)
Abstract：
HIV-1 relies on host RNA polymeraseII (Pol II) to transcribe its genome and uses multiple transcription start sites (TSS), including three consecutive guanosines located near the U3-R junction, to generate transcripts containing three, two, and one guanosine at the 5\' end, referred to as 3G, 2G, and 1G RNA, respectively. The 1G RNA is preferentially selected for packaging, indicating that these 99.9% identical RNAs exhibit functional differences and highlighting the importance of TSS selection. Here, we demonstrate that TSS selection is regulated by sequences between the CATA/TATA box and the beginning of R. Furthermore, we have generated two HIV-1 mutants with distinct 2-nucleotide modifications that predominantly express 3G RNA or 1G RNA. Both mutants can generate infectious viruses and undergo multiple rounds of replication in T cells. However, both mutants exhibit replication defects compared to the wild-type virus. The 3G-RNA-expressing mutant displays an RNA genome-packaging defect and delayed replication kinetics, whereas the 1G-RNA-expressing mutant exhibits reduced Gag expression and a replication fitness defect. Additionally, reversion of the latter mutant is frequently observed, consistent with sequence correction by plus-strand DNA transfer during reverse transcription. These findings demonstrate that HIV-1 maximizes its replication fitness by usurping the TSS heterogeneity of host RNA Pol II to generate unspliced RNAs with different specialized roles in viral replication. The three consecutive guanosines at the junction of U3 and R may also maintain HIV-1 genome integrity during reverse transcription. These studies reveal the intricate regulation of HIV-1 RNA and complex replication strategy.
摘要：
HIV-1依靠宿主RNA聚合酶II（PolII）转录其基因组并使用多个转录起始位点（TSS），包括位于U3-R交界处附近的三个连续的鸟嘌呤，生成包含三个的转录本，两个,在5'末端有一个鸟苷，被称为3G。2G,和1GRNA，分别。1GRNA被优先选择用于包装，这表明这些99.9%相同的RNA表现出功能差异并突出了TSS选择的重要性。这里,我们证明了TSS选择受CATA/TATA框和R的开始之间的序列调节。我们已经产生了两个HIV-1突变体，它们具有不同的2核苷酸修饰，主要表达3GRNA或1GRNA。两种突变体都可以产生感染性病毒并在T细胞中进行多轮复制。然而,与野生型病毒相比,两种突变体均表现出复制缺陷。表达3G-RNA的突变体表现出RNA基因组包装缺陷和延迟复制动力学，而表达1G-RNA的突变体表现出降低的Gag表达和复制适应性缺陷。此外,经常观察到后一种突变体的回复，与逆转录过程中通过正链DNA转移进行的序列校正一致。这些发现表明，HIV-1通过篡夺宿主RNAPolII的TSS异质性来生成在病毒复制中具有不同专门作用的未剪接RNA，从而最大化了其复制适应性。在U3和R的连接处的三个连续的鸟嘌呤也可以在逆转录期间维持HIV-1基因组完整性。这些研究揭示了HIV-1RNA的复杂调控和复杂的复制策略。
