Differential Incorporation of Cholesterol by Sindbis Virus Grown in Mammalian or Insect Cells
Differential Incorporation of Cholesterol by Sindbis Virus Grown in Mammalian or Insect Cells
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DOI：
10.1128/jvi.00755-09
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发表时间：
2009-09-15
影响因子：
5.4
通讯作者：
Hernandez, Raquel
中科院分区：
文献类型：
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作者：
Hafer, Amanda;Whittlesey, Rebecca;Hernandez, Raquel
胆固醇已被证明是甲病毒与人工膜（脂质体）融合所必需的。胆固醇也被认为在甲病毒进入和离开活细胞的过程中起着重要和关键的作用。奇怪的是，甲病毒的替代宿主昆虫是胆固醇营养缺陷型，并且含有非常低水平的这种固醇。为了进一步评估胆固醇在甲病毒生命周期中的作用，测量了由三种不同的蚊子（白纹伊蚊）细胞系;另一种昆虫细胞系，来自草地贪夜蛾的Sf 21;和BHK（哺乳动物）细胞产生的甲病毒辛德毕斯的胆固醇水平。辛德毕斯病毒在正常培养条件下在昆虫细胞中生长，并且通过在使用Cab-O-sil脱脂的血清、用甲基-β-环糊精处理的培养基或无血清培养基中生长而在胆固醇耗尽的细胞中生长。测定掺入细胞膜和从这些细胞产生的病毒中的胆固醇水平。将从这些处理的和未处理的细胞产生的病毒与在标准条件下在BHK细胞中生长的病毒进行比较。昆虫细胞在脱脂后产生辛德毕斯病毒的能力被发现是高度细胞特异性的，并且不依赖于细胞膜中胆固醇的水平。从脱脂细胞中产生野生型水平的感染性辛德毕斯病毒需要非常低水平的胆固醇。数据表明，病毒膜的一个作用是结构性的，提供了感染性所需的稳定性，而在某些细胞中，脱脂膜可能无法提供这种稳定性。这些数据表明，宿主细胞膜中胆固醇的量本身对病毒组装过程或病毒感染细胞的能力没有影响。相反，这些数据表明，辛德毕斯病毒的感染性和组装报告的胆固醇依赖性是所用昆虫细胞系和脱脂方法差异的反映。
Cholesterol has been shown to be essential for the fusion of alphaviruses with artificial membranes (liposomes). Cholesterol has also been implicated as playing an essential and critical role in the processes of entry and egress of alphaviruses in living cells. Paradoxically, insects, the alternate host for alphaviruses, are cholesterol auxotrophs and contain very low levels of this sterol. To further evaluate the role of cholesterol in the life cycle of alphaviruses, the cholesterol levels of the alphavirus Sindbis produced from three different mosquito (Aedes albopictus) cell lines; one other insect cell line, Sf21 from Spodoptera frugiperda; and BHK (mammalian) cells were measured. Sindbis virus was grown in insect cells under normal culture conditions and in cells depleted of cholesterol by growth in serum delipidated by using Cab-O-sil, medium treated with methyl-beta-cyclodextrin, or serum-free medium. The levels of cholesterol incorporated into the membranes of the cells and into the virus produced from these cells were determined. Virus produced from these treated and untreated cells was compared to virus grown in BHK cells under standard conditions. The ability of insect cells to produce Sindbis virus after delipidation was found to be highly cell specific and not dependent on the level of cholesterol in the cell membrane. A very low level of cholesterol was required for the generation of wild-type levels of infectious Sindbis virus from delipidated cells. The data show that one role of the virus membrane is structural, providing the stability required for infectivity that may not be provided by the delipidated membranes in some cells. These data show that the amount of cholesterol in the host cell membrane in and of itself has no effect on the process of virus assembly or on the ability of virus to infect cells. Rather, these data suggest that the cholesterol dependence reported for infectivity and assembly of Sindbis virus is a reflection of differences in the insect cell lines used and the methods of delipidation.