Involvement of clathrin-mediated endocytosis in human immunodeficiency virus type 1 entry
Involvement of clathrin-mediated endocytosis in human immunodeficiency virus type 1 entry
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DOI：
10.1128/jvi.79.3.1581-1594.2005
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发表时间：
2005-02-01
影响因子：
5.4
通讯作者：
Kräusslich, HG
中科院分区：
文献类型：
--
作者：
Daecke, J;Fackler, OT;Kräusslich, HG
人类免疫缺陷病毒（HIV）的生产性进入被认为是通过直接融合在质膜上发生的。在几项研究中观察到HIV颗粒的内吞摄取，但被认为是非生产性的，导致病毒在溶酶体中降解。我们在这里表明，内吞作用通过使用动力蛋白和Eps15的反显性阴性突变体显著地促进了HIV进入HeLa细胞。在cd4阳性的HeLa细胞系中诱导表达动力蛋白的显性阴性突变体可减少40 - 80%的HIV感染。这种效应与感染剂量无关，并在三种不同的分离株中观察到。通过实时PCR对逆转录产物和通过递送病毒粒子相关的vpr - β -内酰胺酶融合蛋白对病毒进入的分析显示了类似的减少，表明阻断发生在进入阶段。短暂转染另一种不同的动力蛋白反式显性阴性变体后，也观察到HIV进入量的显著减少，这种减少与转铁蛋白内吞作用的相对抑制有关。Eps15的显性阴性变体的表达，特异于网格蛋白依赖的内吞作用，使HIV进入HeLa细胞减少了约95%，证实了内吞作用在生产性感染中的作用。相比之下，对caveolin的显性阴性变体没有观察到任何影响。我们得出结论，动力蛋白依赖性、网格蛋白介导的内吞作用可导致HIV高产地进入HeLa细胞，这表明该途径是病毒进入的另一种途径。
Productive entry of human immunodeficiency virus (HIV) is believed to occur by direct fusion at the plasma membrane. Endocytic uptake of HIV particles has been observed in several studies but is considered to be nonproductive, leading to virus degradation in the lysosome. We show here that endocytosis contributes significantly to productive HIV entry in HeLa cells by using trans dominant-negative mutants of dynamin and Eps15. Inducible expression of a dominant-negative mutant of dynamin in a CD4-positive HeLa cell line reduced HIV infection by 40 to 80%. This effect was independent of the infectious dose and was observed for three different isolates. Analysis of reverse transcription products by real-time PCR and of virus entry by delivery of a virion-associated Vpr-beta-lactamase fusion protein revealed a similar reduction, indicating that the block occurred at the entry stage. A strong reduction of HIV entry was also observed upon transient transfection of a different trans dominant-negative variant of dynamin, and this reduction correlated with the relative inhibition of transferrin endocytosis. Expression of a dominant-negative variant of Eps15, which is specific for clathrin-dependent endocytosis, reduced HIV entry in HeLa cells by ca 95%, confirming the role of endocytosis for productive infection. In contrast, no effect was observed for a dominant-negative variant of caveolin. We conclude that dynamin-dependent, clathrin-mediated endocytosis can lead to productive entry of HIV in HeLa cells, suggesting this pathway as an alternative route of virus entry.