HIV-1 buds predominantly at the plasma membrane of primary human macrophages.
HIV-1 buds predominantly at the plasma membrane of primary human macrophages.
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DOI：
10.1371/journal.ppat.0030036
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发表时间：
2007-03
期刊：
影响因子：
6.7
通讯作者：
Krausslich, Hans-Georg
中科院分区：
文献类型：
--
作者：
Welsch, Sonja;Keppler, Oliver T;Habermann, Anja;Allespach, Ina;Krijnse-Locker, Jacomine;Krausslich, Hans-Georg
HIV-1的组装和释放被认为发生在大多数宿主细胞的质膜上，但初级巨噬细胞除外，据报道初级巨噬细胞只在晚期内体出芽。在这里，我们应用了一种新颖的超微结构方法以独立于免疫标记物的方式评估原代巨噬细胞中的 HIV-1 出芽。受感染的巨噬细胞用 BSA 金喂养，并用非膜渗透染料钌红染色，以分别识别内体和质膜。充满病毒的空泡结构似乎位于细胞内，显示出强烈的钌红染色，但缺乏内吞的 BSA-金，将其定义为质膜。此外，HIV出芽谱实际上被排除在金填充内体之外，而经常在钌红阳性膜上检测到。掺入 HIV-1 中的细胞标记蛋白的组成支持病毒包膜的质膜来源。因此，与目前的观点相反，在受感染的巨噬细胞中，质膜也是 HIV-1 出芽的主要部位。巨噬细胞是HIV-1感染的主要靶细胞之一，在病毒发病机制中发挥着重要作用。先前的研究表明，人类原代巨噬细胞中 HIV-1 颗粒形态发生的途径是不同的，这被认为在病毒持久性中发挥着作用。早期报告表明，HIV-1 在明显的细胞内液泡结构中积聚，后来被鉴定为晚期内体起源。因此，内体被认为是人类原代巨噬细胞中 HIV-1 的出芽和储存室，感染性病毒可以以受控的方式从其中释放。在本研究中，我们表明 HIV-1 出芽主要发生在原代人类巨噬细胞的质膜上。使用电子显微镜，我们观察到巨噬细胞的细胞表面呈现出意想不到的复杂形态，有许多突起和深度内陷。 HIV-1出芽主要发生在这些明显与细胞表面相连且不属于内吞区室的内陷处。成熟的病毒颗粒可以保留在这种内陷中，从而形成细胞内出芽室的外观。这些结果表明以质膜作为病毒出芽位点的 HIV-1 形态发生的一般途径。
HIV-1 assembly and release are believed to occur at the plasma membrane in most host cells with the exception of primary macrophages, for which exclusive budding at late endosomes has been reported. Here, we applied a novel ultrastructural approach to assess HIV-1 budding in primary macrophages in an immunomarker-independent manner. Infected macrophages were fed with BSA-gold and stained with the membrane-impermeant dye ruthenium red to identify endosomes and the plasma membrane, respectively. Virus-filled vacuolar structures with a seemingly intracellular localization displayed intense staining with ruthenium red, but lacked endocytosed BSA-gold, defining them as plasma membrane. Moreover, HIV budding profiles were virtually excluded from gold-filled endosomes while frequently being detected on ruthenium red–positive membranes. The composition of cellular marker proteins incorporated into HIV-1 supported a plasma membrane–derived origin of the viral envelope. Thus, contrary to current opinion, the plasma membrane is the primary site of HIV-1 budding also in infected macrophages. Macrophages are one of the major target cells for HIV-1 infection and play an important role in viral pathogenesis. Previous studies indicated that the pathway of HIV-1 particle morphogenesis is distinct in primary human macrophages, and this has been suggested to play a role in viral persistence. Early reports indicated that HIV-1 accumulates within apparently intracellular vacuolar structures, which were later identified as being of late endosomal origin. Endosomes were therefore suggested to comprise the budding and storage compartment for HIV-1 in primary human macrophages, from which infectious virus can be released in a regulated manner. In the present study, we show that HIV-1 budding occurs predominantly at the plasma membrane also in primary human macrophages. Using electron microscopy, we observed that the cell surface of macrophages displays an unexpectedly complex morphology with many protrusions and deep invaginations. HIV-1 budding occurs primarily at these invaginations that are clearly connected to the cell surface and do not belong to the endocytic compartment. Mature virus particles can remain trapped within such invaginations giving the appearance of an intracellular budding compartment. These results suggest a general pathway of HIV-1 morphogenesis with the plasma membrane as viral budding site.