Section of Infectious Diseases, Department of Internal Medicine, Yale University School of Medicine, New Haven, Connecticut, USA.
J Virol. 2012 Nov;86(22):12053-68. doi: 10.1128/JVI.01970-12. Epub 2012 Aug 29.
Productive replication of human immunodeficiency virus type 1 (HIV-1) occurs efficiently only in humans. The posttranscriptional stages of the HIV-1 life cycle proceed poorly in mouse cells, with a resulting defect in viral assembly and release. Previous work has shown that the presence of human chromosome 2 increases HIV-1 production in mouse cells. Recent studies have shown that human chromosome region maintenance 1 (hCRM1) stimulates Gag release from rodent cells. Here we report that expressions of hCRM1 in murine cells resulted in marked increases in the production of infectious HIV-1 and feline immunodeficiency virus (FIV). HIV-1 production was also increased by hSRp40, and a combination of hCRM1 and hSRp40 resulted in a more-than-additive effect on HIV-1 release. In contrast, the overexpression of mouse CRM1 (mCRM1) minimally affected HIV-1 and FIV production and did not antagonize hCRM1. In the presence of hCRM1 there were large increases in the amounts of released capsid, which paralleled the increases in the infectious titers. Consistent with this finding, the ratios of unspliced to spliced HIV-1 mRNAs in mouse cells expressing hCRM1 and SRp40 became similar to those of human cells. Furthermore, imaging of intron-containing FIV RNA showed that hCRM1 increased RNA export to the cytoplasm.By testing chimeras between mCRM1 and hCRM1 and comparing those sequences to feline CRM1, we mapped the functional domain to HEAT (Huntingtin, elongation factor 3, protein phosphatase 2A, and the yeast kinase TOR1) repeats 4A to 9A and a triple point mutant in repeat 9A, which showed a loss of function. Structural analysis suggested that this region of hCRM1 may serve as a binding site for viral or cellular factors to facilitate lentiviral RNA nuclear export.
人类免疫缺陷病毒 1 型（HIV-1）的有效复制仅在人类中发生。HIV-1 生命周期的转录后阶段在小鼠细胞中进行得很差，导致病毒装配和释放出现缺陷。以前的工作表明，人类染色体 2 的存在会增加小鼠细胞中的 HIV-1 产量。最近的研究表明，人类染色体区域维持 1（hCRM1）刺激鼠细胞中的 Gag 释放。在这里，我们报告 hCRM1 在鼠细胞中的表达导致感染性 HIV-1 和猫免疫缺陷病毒（FIV）的产量显著增加。hSRp40 的表达也增加了 HIV-1 的产生，hCRM1 和 hSRp40 的组合对 HIV-1 的释放产生了超过加性的效果。相比之下，鼠 CRM1（mCRM1）的过度表达对 HIV-1 和 FIV 的产生影响很小，并且不会拮抗 hCRM1。在存在 hCRM1 的情况下，释放的衣壳量大量增加，与感染滴度的增加相平行。与这一发现一致，在表达 hCRM1 和 SRp40 的鼠细胞中，未剪接的 HIV-1 mRNA 与剪接的 HIV-1 mRNA 的比例变得类似于人细胞中的比例。此外，包含内含子的 FIV RNA 的成像表明 hCRM1 增加了 RNA 向细胞质的输出。通过测试 mCRM1 和 hCRM1 之间的嵌合体，并将这些序列与猫科 CRM1 进行比较，我们将功能域映射到 HEAT（亨廷顿、伸长因子 3、蛋白磷酸酶 2A 和酵母激酶 TOR1）重复 4A 到 9A 和重复 9A 中的三点突变，该突变显示出功能丧失。结构分析表明，hCRM1 的这一区域可能作为病毒或细胞因子的结合位点，促进慢病毒 RNA 的核输出。
