Schaeffer Evelyne, Soros Vanessa B, Greene Warner C
Gladstone Institute of Virology and Immunology, University of California, San Francisco, California 94141, USA.
J Virol. 2004 Feb;78(3):1375-83. doi: 10.1128/jvi.78.3.1375-1383.2004.
Virions of the type 1 human immunodeficiency virus (HIV-1) can enter target cells by fusion or endocytosis, with sharply different functional consequences. Fusion promotes productive infection of the target cell, while endocytosis generally leads to virion inactivation in acidified endosomes or degradation in lysosomes. Virion fusion and endocytosis occur equally in T cells, but these pathways have been regarded as independent because endocytosis of HIV virions requires neither CD4 nor CCR5/CXCR4 engagement in HeLa-CD4 cells. Using flow cytometric techniques to assess the binding and entry of green fluorescent protein (GFP)-Vpr-labeled HIV virions into primary peripheral blood mononuclear cells, we have found that HIV fusion and endocytosis are restricted to the CD4-expressing subset of cells and that both pathways commonly require the initial binding of HIV virions to surface CD4 receptors. Blockade of CXCR4-tropic HIV virion fusion with AMD3100, a CXCR4-specific entry inhibitor, increased virion entry via the endocytic pathway. Similarly, inhibition of endosome acidification with bafilomycin A1, concanamycin A, or NH(4)Cl enhanced entry via the fusion pathway. Although fusion remained dependent on CD4 and chemokine receptor binding, the endosome inhibitors did not alter surface expression of CD4 and CXCR4. These results suggest that fusion in the presence of the endosome inhibitors likely occurs within nonacidified endosomes. However, the ability of these inhibitors to impair vesicle trafficking from early to late endosomes in some cells could also increase the recycling of these virion-containing endosomes to the cell surface, where fusion occurs. In summary, our results reveal an unexpected, CD4-mediated reciprocal relationship between the pathways governing HIV virion fusion and endocytosis.
1型人类免疫缺陷病毒（HIV-1）的病毒粒子可通过融合或内吞作用进入靶细胞，其功能后果截然不同。融合促进靶细胞的有效感染，而内吞作用通常导致病毒粒子在酸化的内体中失活或在溶酶体中降解。病毒粒子的融合和内吞作用在T细胞中同等发生，但这些途径被认为是独立的，因为HIV病毒粒子的内吞作用在HeLa-CD4细胞中既不需要CD4也不需要CCR5/CXCR4参与。使用流式细胞术技术评估绿色荧光蛋白（GFP）-Vpr标记的HIV病毒粒子与原代外周血单核细胞的结合和进入，我们发现HIV融合和内吞作用仅限于表达CD4的细胞亚群，并且这两种途径通常都需要HIV病毒粒子与表面CD4受体的初始结合。用CXCR4特异性进入抑制剂AMD3100阻断趋化因子受体CXCR4的HIV病毒粒子融合，增加了通过内吞途径的病毒粒子进入。同样，用巴弗洛霉素A1、 concanamycin A或NH(4)Cl抑制内体酸化增强了通过融合途径的进入。尽管融合仍然依赖于CD4和趋化因子受体结合，但内体抑制剂并未改变CD4和CXCR4的表面表达。这些结果表明，在内体抑制剂存在下的融合可能发生在非酸化的内体中。然而，这些抑制剂在某些细胞中损害从早期内体到晚期内体的囊泡运输的能力也可能增加这些含有病毒粒子的内体向细胞表面的再循环，在细胞表面发生融合。总之，我们的结果揭示了HIV病毒粒子融合和内吞作用途径之间意想不到的、由CD4介导的相互关系。
