West nile virus entry requires cholesterol-rich membrane microdomains and is independent of αvβ3 integrin
West nile virus entry requires cholesterol-rich membrane microdomains and is independent of αvβ3 integrin
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DOI：
10.1128/jvi.00008-08
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发表时间：
2008-06-01
影响因子：
5.4
通讯作者：
Nelson, Jay A.
中科院分区：
文献类型：
--
作者：
Medigeshi, Guruprasad R.;Hirsch, Alec J.;Nelson, Jay A.
自1999年以来，西尼罗河病毒（WNV）一直是美国病毒性脑炎的主要原因。感染性WNV内化的各种细胞类型的内吞过程没有得到很好的表征，并参与富含胆固醇的膜微区，或脂筏，在WNV的生命周期中，以前没有调查。在这项研究中，我们发现通过用甲基-β-环糊精短暂处理细胞胆固醇水平的消耗导致释放到培养上清液中的感染性WNV的滴度降低100倍，以及胆固醇消耗细胞中WNV基因组拷贝数的减少。将外源性胆固醇添加到胆固醇耗尽的细胞中逆转了这种效应。胆固醇消耗感染后并没有影响西尼罗河病毒的增长，这表明效果发生在西尼罗河病毒进入的水平。我们还表明，虽然西尼罗河病毒进入不需要α v β 3整合素和黏着斑激酶，西尼罗河病毒颗粒未能被胆固醇耗尽的细胞内化。最后，我们显示了共定位的WNV包膜蛋白和霍乱毒素B，这是内在的一个脂筏依赖性途径，在细胞膜上的微区簇。这些数据表明，西尼罗河病毒在内化的初始阶段利用脂筏，脂筏可能含有一种可增强西尼罗河病毒内吞作用的因子。
West Nile virus (WNV) has been the leading cause of viral encephalitis in the United States since 1999. The endocytic processes involved in the internalization of infectious WNV by various cell types are not well characterized, and the involvement of cholesterol-rich membrane microdomains, or lipid rafts, in the life cycle of WNV has not been investigated previously. In this study, we found that the depletion of cellular cholesterol levels by brief treatment with methyl-beta-cyclodextrin resulted in a 100-fold reduction of the titers of infectious WNV released into the culture supernatant, as well as a reduction in the number of WNV genome copies in the cholesterol-depleted cells. The addition of exogenous cholesterol to cholesterol-depleted cells reversed this effect. Cholesterol depletion postinfection did not affect WNV growth, suggesting that the effect occurs at the level of WNV entry. We also showed that while WNV entry did not require alpha v beta 3 integrin and focal adhesion kinase, WNV particles failed to be internalized by cholesterol-depleted cells. Finally, we showed the colocalization of the WNV envelope protein and cholera toxin B, which is internalized in a lipid raft-dependent pathway, in microdomain clusters at the plasma membrane. These data suggest that WNV utilizes lipid rafts during initial stages of internalization and that the lipid rafts may contain a factor(s) that may enhance WNV endocytosis.