Pawlica Paulina, Berthoux Lionel
Laboratory of Retrovirology, Department of Medical Biology and BioMed Research Center, Université du Québec à Trois-Rivières, 3351 Boulevard des Forges, Trois-Rivières, QC G9A 5H7, Canada.
Viruses. 2014 Nov 4;6(11):4195-211. doi: 10.3390/v6114195.
Retroviral capsid (CA) cores undergo uncoating during their retrograde transport (toward the nucleus), and/or after reaching the nuclear membrane. However, whether HIV-1 CA core uncoating is dependent upon its transport is not understood. There is some evidence that HIV-1 cores retrograde transport involves cytoplasmic dynein complexes translocating on microtubules. Here we investigate the role of dynein-dependent transport in HIV-1 uncoating. To interfere with dynein function, we depleted dynein heavy chain (DHC) using RNA interference, and we over-expressed p50/dynamitin. In immunofluorescence microscopy experiments, DHC depletion caused an accumulation of CA foci in HIV-1 infected cells. Using a biochemical assay to monitor HIV-1 CA core disassembly in infected cells, we observed an increase in amounts of intact (pelletable) CA cores upon DHC depletion or p50 over-expression. Results from these two complementary assays suggest that inhibiting dynein-mediated transport interferes with HIV-1 uncoating in infected cells, indicating the existence of a functional link between HIV-1 transport and uncoating.
逆转录病毒衣壳(CA)核心在其逆行运输(朝向细胞核)过程中,和/或在到达核膜后会发生脱壳。然而,HIV-1 CA核心脱壳是否依赖于其运输尚不清楚。有一些证据表明,HIV-1核心的逆行运输涉及在微管上转运的细胞质动力蛋白复合体。在这里,我们研究了动力蛋白依赖性运输在HIV-1脱壳中的作用。为了干扰动力蛋白的功能,我们使用RNA干扰耗尽了动力蛋白重链(DHC),并过表达了p50/动力蛋白抑制因子。在免疫荧光显微镜实验中,DHC的耗尽导致HIV-1感染细胞中CA病灶的积累。使用生化分析来监测感染细胞中HIV-1 CA核心的解体,我们观察到在DHC耗尽或p50过表达时,完整(可沉淀)CA核心的数量增加。这两种互补分析的结果表明,抑制动力蛋白介导的运输会干扰感染细胞中HIV-1的脱壳,表明HIV-1运输与脱壳之间存在功能联系。