Hepatitis C Virus NS5A Activates the Mammalian Target of Rapamycin (mTOR) Pathway, Contributing to Cell Survival by Disrupting the Interaction between FK506-binding Protein 38 (FKBP38) and mTOR
Hepatitis C Virus NS5A Activates the Mammalian Target of Rapamycin (mTOR) Pathway, Contributing to Cell Survival by Disrupting the Interaction between FK506-binding Protein 38 (FKBP38) and mTOR
复制标题
丙型肝炎病毒 NS5A 激活哺乳动物雷帕霉素靶点 (mTOR) 通路，通过破坏 FK506 结合蛋白 38 (FKBP38) 和 mTOR 之间的相互作用来促进细胞存活
DOI：
10.1074/jbc.m110.112045
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发表时间：
2010-07-02
影响因子：
4.8
通讯作者：
Yuan, Zhenghong
中科院分区：
文献类型：
--
作者：
Peng, Lu;Liang, Dongyu;Yuan, Zhenghong
丙型肝炎病毒 (HCV) 通常会造成持续感染，很可能涉及复杂的宿主-病毒相互作用。我们之前报道，HCV 非结构蛋白 5A (NS5A) 与细胞蛋白 FKBP38 结合，导致人肝癌细胞系 Huh7 细胞凋亡受到抑制。在本研究中，我们进一步发现，在没有血清的情况下，NS5A 增加了 Huh7 中两种 mTOR 靶向底物 S6K1 和 4EBP1 的磷酸化水平。 mTOR 抑制剂雷帕霉素或 NS5A 敲低可阻断 NS5A-Huh7 和 HCV 复制子细胞中 S6K1 和 4EBP1 磷酸化的增加，表明 NS5A 特异性调节 mTOR 激活。 NS5A 和 FKBP38 突变体的过度表达或 FKBP38 敲低表明 mTOR 激活依赖于 NS5A-FKBP38 相互作用。 NS5A-Huh7 中的磷脂酰肌醇 3-激酶 (PI3K) 抑制剂 LY294002 处理表明 mTOR 激活不依赖于 PI3K。此外，NS5A 抑制 caspase 3 和聚（ADP-核糖）聚合酶的激活，而 NS5A 敲低或雷帕霉素可消除这种激活，表明 NS5A 特异性通过 mTOR 途径抑制细胞凋亡。进一步的分析表明，NS5A 通过 mTOR 发挥的细胞凋亡抑制作用也需要 NS5A-FKBP38 相互作用。谷胱甘肽 S-转移酶下拉和免疫共沉淀表明 NS5A 破坏了 mTOR-FKBP38 关联。此外，NS5A 或 FKBP38 突变体恢复了 mTOR-FKBP38 相互作用；这表明 mTOR-FKBP38 关联的损害依赖于 NS5A-FKBP38 结合。总的来说，我们的数据表明，HCV NS5A 通过损害 mTOR 和 FKBP38 之间的相互作用来激活 mTOR 途径来抑制细胞凋亡，这可能代表 HCV 持续存在和发病机制的关键机制。
Hepatitis C virus (HCV) often establishes a persistent infection that most likely involves a complex host-virus interplay. We previously reported that the HCV nonstructural protein 5A (NS5A) bound to cellular protein FKBP38 and resulted in apoptosis suppression in human hepatoma cell line Huh7. In the present research we further found that NS5A increased phosphorylation levels of two mTOR-targeted substrates, S6K1 and 4EBP1, in Huh7 in the absence of serum. mTOR inhibitor rapamycin or NS5A knockdown blocked S6K1 and 4EBP1 phosphorylation increase in NS5A-Huh7 and HCV replicon cells, suggesting that NS5A specifically regulated mTOR activation. Overexpression of NS5A and FKBP38 mutants or FKBP38 knockdown revealed this mTOR activation was dependent on NS5A-FKBP38 interaction. Phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 treatment in NS5A-Huh7 showed that the mTOR activation was independent of PI3K. Moreover, NS5A suppressed caspase 3 and poly(ADP-ribose) polymerase activation, which was abolished by NS5A knockdown or rapamycin, indicating NS5A inhibited apoptosis specifically through the mTOR pathway. Further analyses suggested that apoptotic inhibition exerted by NS5A via mTOR also required NS5A-FKBP38 interaction. Glutathione S-transferase pulldown and co-immunoprecipitation showed that NS5A disrupted the mTOR-FKBP38 association. Additionally, NS5A or FKBP38 mutants recovered the mTOR-FKBP38 interaction; this indicated that the impairment of mTOR-FKBP38 association was dependent on NS5A-FKBP38 binding. Collectively, our data demonstrate that HCV NS5A activates the mTOR pathway to inhibit apoptosis through impairing the interaction between mTOR and FKBP38, which may represent a pivotal mechanism for HCV persistence and pathogenesis.