Entry of poliovirus into cells is blocked by valinomycin and concanamycin A
Entry of poliovirus into cells is blocked by valinomycin and concanamycin A
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DOI：
10.1021/bi002069p
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发表时间：
2001-03-27
期刊：
影响因子：
2.9
通讯作者：
Carrasco, L
中科院分区：
文献类型：
--
作者：
Irurzun, A;Carrasco, L
脊髓灰质炎病毒包含一种无脂质包膜的病毒粒子，其进入易感细胞不需要完整的pH值。因此，内体中产生的pH梯度阻断不足以阻碍脊髓灰质炎病毒粒子向细胞质的移位，这表明移位发生在质膜上。测量由脊髓灰质炎病毒蛋白酶2A介导的病毒蛋白合成和eIF4G - 1裂解已被用于监测脊髓灰质炎病毒进入细胞的有效过程。输入的脊髓灰质炎病毒RNA的翻译产生足够的2A（pro）来裂解eIF4G - 1，为估计脊髓灰质炎病毒RNA传递到细胞质及其随后的翻译提供了一种灵敏的检测方法。刀豆霉素A（一种液泡质子 - ATP酶抑制剂）和缬氨霉素（一种促进细胞内K⁺外流的离子载体）联合使用可有力地阻止脊髓灰质炎病毒感染。此外，改变培养基的离子条件（增加K⁺浓度并降低Na⁺浓度），再加上刀豆霉素A，也会显著干扰脊髓灰质炎病毒的进入。这些发现表明，脊髓灰质炎病毒RNA需要细胞内完整的K⁺离子浓度才能脱壳并进入细胞质。此外，刀豆霉素A（以及其他内吞作用抑制剂）对有效阻止脊髓灰质炎病毒进入的实际贡献表明，至少有一定比例的脊髓灰质炎病毒亚粒子从内体移位。
Poliovirus contains a virus particle devoid of a lipid envelope that does not require an intact pH to enter into susceptible cells. Thus, the blockade of pH gradient generated in endosomes is not sufficient to impede the translocation of poliovirus particles to the cytoplasm, suggesting that translocation takes place at the plasma membrane. Measuring both viral protein synthesis and eIF4G-1 cleavage mediated by poliovirus protease 2A has been used to monitor productive entry of poliovirus into cells. Translation of the input poliovirus RNA produces enough 2A(pro) to cleave eIF4G-1, providing a sensitive assay to estimate poliovirus RNA delivery to the cytoplasm followed by its translation. Combination of concanamycin A, a vacuolar proton-ATPase inhibitor, and valinomycin, an ionophore that promotes K+ efflux from cells, powerfully prevented poliovirus infection. Moreover, modifying the ionic conditions of the culture medium (increasing the concentration of K+ and decreasing the concentration of Na+), together with concanamycin A, also significantly interfered with poliovirus entry. These findings suggest that poliovirus RNA requires an intact concentration of K+ ions inside the cells to be uncoated and to gain access to the cytoplasm. In addition, the actual contribution of concanamycin A (as well as other inhibitors of endocytosis) to the total inhibition of productive poliovirus entry points to the idea that at least some percentage of polioviral subparticles translocates from the endosomes.