Porcine circovirus 2 infection of epithelial cells is clathrin-, caveolae- and dynamin-independent, actin and Rho-GTPase-mediated, and enhanced by cholesterol depletion
Porcine circovirus 2 infection of epithelial cells is clathrin-, caveolae- and dynamin-independent, actin and Rho-GTPase-mediated, and enhanced by cholesterol depletion
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DOI：
10.1016/j.virusres.2008.09.005
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发表时间：
2009-01-01
期刊：
影响因子：
5
通讯作者：
Nauwynck, H. J.
中科院分区：
文献类型：
--
作者：
Misinzo, G.;Delputte, P. L.;Nauwynck, H. J.
上皮细胞是猪圆环病毒 2 型 (PCV2) 的主要体内靶细胞。尽管这些细胞用于大多数 PCV2 基因表达研究，但对 PCV2 在上皮细胞中的进入、附着和内化知之甚少。 PCV2 与上皮细胞的附着发生迅速且具有时间依赖性。与依恋相反，内化缓慢。免疫荧光染色显示，在内化过程中，PCV2 与网格蛋白共定位，但与小窝蛋白不共定位。阻断网格蛋白介导的内吞作用使 PCV2 感染细胞的数量增加而不是减少三倍，这表明它并不代表导致完全复制的主要内化途径。对不同抑制剂的进一步分析表明，巨胞饮作用、动力依赖性内化和膜胆固醇在 PCV2 进入导致感染的过程中不起作用。用艰难梭菌毒素 B 抑制小 GTP 酶可将 PCV2 感染的 PK-15、SK 和 ST 的数量分别减少至 63 +/- 25%、47 +/- 21 % 和 14 +/- 6%。最后，抑制肌动蛋白聚合也阻止了PCV2感染，表明PCV2感染期间需要肌动蛋白。总之，这些数据表明，独立于动力蛋白和胆固醇，但依赖于肌动蛋白和小GTP酶的途径，允许PCV2在上皮细胞中内化，从而导致感染，并且上皮细胞中网格蛋白介导的PCV2内化不会随后进行完全复制。 (C) 2008 Elsevier B.V. 保留所有权利。
Epithelial cells are the major in vivo target cells for porcine circovirus type 2 (PCV2). Although these cells are used for most studies of PCV2 gene expression and, little is known on PCV2 entry, attachment and internalization, in epithelial cells. PCV2 attachment to epithelial cells occurred rapidly and in a time-dependent manner. In contrast to attachment, internalization was slow. Immunofluorescent stainings revealed that during internalization, PCV2 co-localized with clathrin, but not caveolin. Blocking clathrin-mediated endocytosis increased instead of decreased the number of PCV2-infected cells by threefold, suggesting that it does not represent the main internalization pathway leading to a full replication. Further analysis with different inhibitors revealed that also macropinocytosis, dynamin-dependent internalization and membrane cholesterol play no role in PCV2 entry that leads to infection. Inhibition of small GTPases with Clostridium difficile toxin B reduced the number of PCV2-infected PK-15, SK and STs to 63 +/- 25%, 47 +/- 21 % and 14 +/- 6%, respectively. Finally, inhibiting actin polymerization also blocked PCV2 infection, showing the need for actin during PCV2 infection. Together, these data indicate that a dynamin- and cholesterol-independent, but actin- and small GTPase-dependent pathway, allows PCV2 internalization in epithelial cells that leads to infection and that clathrin-mediated PCV2 internalization in epithelial cells is not followed by a full replication. (C) 2008 Elsevier B.V. All rights reserved.